regi.tankonyvtar.hu · Web viewApplied biochemistry. Applied biochemistry. Tárgymutató. 1. INTRODUCTION. 2. THE LIVING SYSTEMS. 3. BIOMOLECULES I. CARBOHYDRATES. 4. BIOMOLECULES

Applied biochemistry

Kincses, SándornéBalláne Kovács, Andrea

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Applied biochemistryírta Kincses, Sándorné és Balláne Kovács, Andrea

TÁMOP-4.1.2.A/1-11/1-2011-0009

University of Debrecen, Service Sciences Methodology Centre

Debrecen, 2013.


TartalomTárgymutató ......................................................................................................................................... 11. 1. INTRODUCTION ....................................................................................................................... 2

1. 1. 1. The object of biochemistry, its relationship with other sciences .................................... 22. 1. 2. Relationship between biochemistry and other sciences ................................................. 2

2. 2. THE LIVING SYSTEMS ............................................................................................................ 31. 2. 1. Characterization of living systems ................................................................................. 32. 2. 2. The composition of living matter ................................................................................... 3

3. 3. BIOMOLECULES I. CARBOHYDRATES ................................................................................ 81. 3. 1. Monosaccharides ............................................................................................................ 8

1.1. 3. 1. 1. Formation of cyclic monosaccharides ........................................................... 91.2. 3. 1. 2. Chemical reactions of monosaccharides ...................................................... 10

1.2.1. 3. 1. 2. 1. Redox reactions of monosaccharides .......................................... 101.2.2. 3. 1. 2. 2. Transformations of monosaccharides into each other ................. 11

2. 3. 2. Disaccharides ............................................................................................................... 122.1. 3. 2. 1. Reducing disaccharides ............................................................................... 132.2. 3. 2. 2. Non-reducing disaccharides ........................................................................ 13

3. 3. 3. Polysaccharides ............................................................................................................ 133.1. 3. 3. 1. Classification of polysaccharides ................................................................ 14

4. 4. BIOMOLECULES II. PROTEINS ........................................................................................... 161. 4. 1. Proteins can be classified ............................................................................................. 17

5. 5. BIOMOLECULES III. THE LIPIDS ................................ Error: Reference source not found1. 5.1. Classification of lipids: .................................................................................................. 19

1.1. 5. 1. 1. Saponifiable lipids ....................................................................................... 191.1.1. 5.1.1.1. Vaxes .............................................................................................. 201.1.2. 5.1.1.2. Neutral fats and oils (triglycerides) ................................................ 201.1.3. 5.1.1.3. Phosphoglycerides ........................................................................ 211.1.4. 5.1.1.4. Sphingolipids ................................................................................. 221.1.5. 5.1.1.5. Glycolipides ................................................................................... 22

1.2. 5.1.2. Insaponifiable lipids ...................................................................................... 221.2.1. 5.1.2.1. Steroids ........................................................................................... 221.2.2. 5.1.2.2. Carotenoids .................................................................................... 221.2.3. 5.1.2.3. Lipid soluble vitamins ................................................................... 23

6. 6. BIOMOLECULES IV. THE NUCLEIC ACIDS ...................................................................... 241. 6.1. The deoxyribonucleic acid (DNA) ................................................................................ 25

1.1. 6. 1.1. The primary structure of deoxyribonucleic acid (DNA) .............................. 251.2. 6.1.2. The secondary structure of DNA ................................................................... 251.3. 6.1.3. The biological function of DNA ............................................................ 26

2. 6. 2. The ribonucleic acids (RNA-s) .................................................................................... 262.1. 6. 2.1. The messenger RNA-s .................................................................................. 262.2. 6. 2. 2. Transfer RNA-s ............................................................................................ 272.3. 6. 2. 3. Ribosomal RNA-s ....................................................................................... 27

3. 6. 3. Nucleoside triphosphates .............................................................................................. 287. 7. BIOACTIVE COMPOUNDS I. VITAMINS ......................................................................... 29

1. 7.1. Physiological effects of vitamins: ................................................................................. 292. 7. 2. Lipid soluble vitamins .................................................................................................. 293. 7. 3. Water-soluble vitamins ................................................................................................. 32

8. 8. BIOACTIVE COMPOUNDS II. HORMONS ...................................................................... 37



1. 8.1. Classification of hormones: ........................................................................................... 381.1. 8.1.1. Hormones of hypophysis .............................................................................. 38

1.1.1. 8.1.1.1. The anterior pituitary (adenohypophysis)The defect or removal of anterior pituitary reflect in the operation of all organs to a smaller or larger extent. ......... 381.1.2. 8.1.1.2. Hormones of intermediate lobe (pars intermedia) ........................ 391.1.3. 8.1.1.3. Posterior lobe (neurohypophysis) hormones .................................. 39

1.2. 8.1.2. Hormones of pineal gland .............................................................................. 391.3. 8.1.3. Hormones of the thyroid glands .................................................................... 401.4. 8.1.4. The parathyroid gland .................................................................................... 401.5. 8.1 5. Hormones of the adrenal cortex ..................................................................... 411.6. 8.1.6. Hormones of adrenal medulla (catecholamines) ........................................... 411.7. 8.1.7. Hormones of pancreas ................................................................................. 411.8. 8.1.8. Hormones of the ovary .................................................................................. 411.9. 8.1.9. The testicular hormones (androgens) ............................................................. 42

2. 8. 2. Tissue hormones ........................................................................................................... 423. 8. 3. Plant growth hormones (Phytohormones) .................................................................... 43

9. 9. BIOACTIVE COMPOUNDS III. ENZYMES (BIOCATALIZATORS) ............................... 451. 9. 1. Structure of the enzymes .............................................................................................. 452. 9. 2. The function mechanism of the enzymes ..................................................................... 463. 9. 3. The specificity of the enzymes ..................................................................................... 464. 9. 4. Classification of enzymes ............................................................................................. 47

4.1. 9.4.1. Oxidoreductases ............................................................................................. 474.2. 9.4.2. Transferases ................................................................................................... 484.3. 9.4.3. Hydrolases ..................................................................................................... 484.4. 9.4.4. Lyases (Synthases) ......................................................................................... 494.5. 9.4.5. Isomerases ..................................................................................................... 494.6. 9.4.6. Ligases (synthetases) ..................................................................................... 49

5. 9. 5. Factors influencing the function of enzymes .............................................................. 4910. 10. THE METABOLIC PROCESSES I. CARBOHYDRATE METABOLISM ......................... 51

1. 10. 1. Carbohydrate biosynthesis in photosynthetic organisms ........................................... 521.1. 10.1.1. The light dependent phase (Hill reaction) ................................................... 521.2. 10.1.2 The light independent phase of photosynthesis (Calvin cycle) .................... 531.3. 10.1.3. The sucrose synthesis .................................................................................. 541.4. 10.1.4. The starch synthesis ..................................................................................... 54

2. 10. 2. Catabolic processes of carbohydrates ........................................................................ 552.1. 10.2.1. Cellular respiration ...................................................................................... 55

2.1.1. 10.2.1.1. Glycolysis ..................................................................................... 552.1.2. 10.2.1.2. Pyruvate decarboxylation ............................................................. 572.1.3. 10.2.1.3. Citric acid cycle ........................................................................... 572.1.4. 10.2.1.4. The terminal oxidation and oxidative phosphorylation ............... 58

2.2. 10.2.2. The pentose phosphate pathway ................................................................. 592.3. 10.2.3. Fermentation processes ................................................................................ 60

2.3.1. 10.2.3.1. The fermentation processes in the rumen of ruminants ............... 622.3.2. 10.2.3.2. The fermentation processes in silo .............................................. 63

3. 10.3. Gluconeogenesis (Glucose-resynthesis) ...................................................................... 644. 10.4. Glycogen metabolism .................................................................................................. 65

4.1. 10.4.1. Glycogen synthesis ...................................................................................... 654.2. 10.4.2. Glycogen mobilization, catabolism ............................................................. 66

11. 11. THE METABOLIC PROCESSES II. LIPID METABOLISM .............................................. 671. 11.1. Biosynthesis of lipids .................................................................................................. 67

1.1. 11. 1. 1. Biosynthesis of triglicerides ....................................................................... 671.1.1. 11.1.1.1. Biosynthesis of fatty acids ........................................................... 67



1.1.2. 11.1.1.2. The synthesis of glycerol ............................................................. 681.2. 11.1. 2. Biosynthesis of phospholipids ................................................................... 691.3. 11. 1. 3. The biosynthesis of carotenoids and steroid skeleton lipids ...................... 69

1.3.1. 11.1.3.1. The synthesis of steroids .............................................................. 702. 11. 2. The breakdown of lipids ............................................................................................. 70

2.1. 11. 2.1. The β-oxidation of saturated fatty acids .................................................... 712.2. 11.2.2. The catabolism of steroids ........................................................................... 73

3. 11. 3. The formation of ketone bodies (ketogenesis) ........................................................... 734. 11. 4. Glyoxylic acid cycle (Kornberg Krebs cycle) ............................................................ 74

12. 12. THE METABOLIC PROCESSES III. PROTEIN METABOLISM ...................................... 761. 12.1. The nitrogen fixation ................................................................................................... 762. 12.2. The synthesis of essential amino acids ........................................................................ 77

2.1. 12.2.1. The methionine and threonine biosynthesis ................................................ 772.1.1. 12.2.2.1. Methionine formation from homoserin ....................................... 78

2.2. 12.2.2. Lysine biosynthesis ...................................................................................... 782.3. 12.2.3. Arginine biosynthesis ................................................................................... 792.4. 12.2.4. Leucine, isoleucine and valine synthesis ..................................................... 792.5. 12.2.5. The phenylalanine and tryptophan biosynthesis .......................................... 79

3. 12.3. Protein Synthesis ......................................................................................................... 793.1. 12.3.1. The transcription .......................................................................................... 803.2. 12.3. 2. The translation ........................................................................................... 80

3.2.1. 12.3.2.1. Initiation ....................................................................................... 823.2.2. 12.3.2.2. Elongation .................................................................................... 823.2.3. 12.3.2.3. Termination .................................................................................. 83

4. 12.4. The fate of dietary proteins in heterotrophic organisms .............................................. 834.1. 12.4.1. The quality of proteins ................................................................................. 834.2. 12.4.2. The protein balance of the organism ........................................................... 854.3. 12.4.3. The digestion of proteins ............................................................................. 85

4.3.1. 12.4.3.1. Proteases occur in each cell. Their role is wide ranged. .............. 854.3.2. 12.4.3.2. The common features of amino acid degradation pathways ........ 864.3.3. 12.4.3.3. The catabolism of carbon skeleton of amino acids in the tricarboxylic acid cycle ............................................................................................................... 87

4.4. 12.4.4. Protein turnover ........................................................................................... 884.5. 12.4.5. Nitrogen excretion ....................................................................................... 89

4.5.1. 12.4.5.1. Nitrogen excretion in mammals, synthesis of urea (carbamide) .. 894.5.2. 12.4.5.2. Nitrogen excretion of birds and reptiles. Synthesis of uric acid . . 90

4.6. 12.4.6. Disturbances of amino acid metabolism ..................................................... 9013. 13. OTHER BIOCHEMICAL PATHWAYS ................................................................................ 92

1. 13.1. The biochemical bases of the function of skeletal muscle .......................................... 922. 13. 2. Factors influencing the quantity and quality of the urine ........................................... 923. 13. 3. The gastric juice and its separation ............................................................................ 93

3.1. 13. 3. 1. The mechanism of the hydrochloric acid production of the stomach ........ 934. 13. 4. The control of metabolic processes ............................................................................ 94

4.1. 13. 4. 1. The control of lipid metabolism ................................................................ 944.2. 13. 4. 2. The function of adenylate cyclase - cAMP system ................................... 95

4.2.1. 13.4. 2. 1. The presentation of adenylate-cyclase system operation through the mobilization of glycogen ...................................................................................... 964.2.2. 13. 4. 2. 2. Hormone control of carbohydrate metabolism ......................... 97

5. 13. 5. The role of liver in the intermediate metabolism ....................................................... 9814. 14. BIOCHEMICAL PATHWAYS IN THE FOOD INDUSTRY ............................................. 100

1. 14. 1. The application of the fermentation in the food industry ......................................... 1002. 14. 2. The biochemical processes of cereals germination ................................................. 100



3. 14. 3. Respiration during storage ....................................................................................... 1013.1. 14. 3. 1. Respiration of grain during storage ................................................ 1013.2. 14.3.2. The respiration of fruits and vegetables ................................................... 1013.3. 14. 3. 3. The ripening of fruits ............................................................................... 102

4. 14. 4. The biochemistry of meat ripening .......................................................................... 1035. 14. 5. Changes of colour through the meat processing ...................................................... 104

15. 15. RECOMMENDED REFERENCES .................................................................................... 10616. Questions ................................................................................................................................... 107


Az ábrák listája2.1. Table 1: The frequency of elements in the earth's crust and in the human body .......................... 32.2. Table 2. The chemical composition of Escherichia coli bacterium ............................................... 42.3. Figure 1: Molecular organizations in cells .................................................................................... 62.4. Table 3: Classification of organisms based on their mass sources ................................................ 73.1. Figure 2: The most important monosaccharides ........................................................................... 83.2. Figure 3: Formation of cyclic monosaccharides ......................................................................... 103.3. Figure 4: Redox reactions of monosaccharides, of D -glucose .................................................. 103.4. Figure 5: Transformations of monosaccharides .......................................................................... 113.5. Figure 6: Hexose-pentose transformation ................................................................................... 123.6. Figure 7: Disaccharides .............................................................................................................. 133.7. Figure 8: Polysaccharides ........................................................................................................... 154.1. Figure 9: Peptide bond ................................................................................................................ 164.2. Figure 10: Conformation of proteins .......................................................................................... 175.1. the core of gonane ....................................................................................................................... 226.1. Figure 11: Nucleobases ............................................................................................................... 246.2. Figure 12: From nucleotide monomers connected to polynucleotide ......................................... 256.3. Figure 13: The deoxyribonucleic acid. ....................................................................................... 266.4. Figure 14: The mRNA and the tRNA ......................................................................................... 277.1. Figure 15: Vitamin A .................................................................................................................. 307.2. Figure 16: Vitamin D, E, K. ........................................................................................................ 317.3. Figure 17: Vitamins (B1, B2, B3) ............................................................................................... 337.4. Figure 18: Water-soluble vitamins .............................................................................................. 347.5. Table 4: The role of vitamins in the function of enzymes .......................................................... 358.1. Figure 19: Regulation of hormone production ........................................................................... 378.2. Figure 20: The synthesis of melatonin ........................................................................................ 398.3. indole-acetic acid ........................................................................................................................ 438.4. gibberellic acid ............................................................................................................................ 448.5. zeatin ........................................................................................................................................... 449.1. Figure 21: Enzymes and activation energy ................................................................................. 459.2. Figure 22: The specificity of the enzymes .................................................................................. 469.3. Table 5: Coenzymes of transferases and transmitted chemical groups ....................................... 4810.1. Figure 23: The carbon, hydrogen and oxygen biological cycle ................................................ 5110.2. Figure 24: The nitrogen biological cycle .................................................................................. 5110.3. Figure 25: Hill reaction ............................................................................................................. 5310.4. Figure 26: Calvin cycle ............................................................................................................. 5410.5. Figure 27: Catabolic processes of carbohydrates ..................................................................... 5510.6. Figure 28: Glycolysis ................................................................................................................ 5610.7. Figure 29: Pyruvate decarboxylation ........................................................................................ 5710.8. Figure 30: Citric acid cycle and terminal oxidation ................................................................. 5810.9. Figure 31: The pentose phosphate pathway .............................................................................. 5910.10. Figure 32: Fermentation processes ......................................................................................... 6010.11. Figure 33: Alcoholic- and lactic acid fermentation ................................................................. 6110.12. Figure 34: Propionic acid and butyric acid fermentation ...................................................... 6310.13. Table 6: The fermentation processes in silo ............................................................................ 6310.14. Figure 35: Gluconeogenesis .................................................................................................... 6511.1. Figure 36: Biosynthesis of fatty acids ....................................................................................... 6711.2. Figure 37: The synthesis of triglycerid ..................................................................................... 68



11.3. Figure 38: The synthesis of isoprene ........................................................................................ 6911.4. Figure 39: The β-oxidation ....................................................................................................... 7111.5. Figure 40: Breakdown fatty acids with odd-numbered carbon ................................................. 7211.6. Figure 41: Ketogenesis ............................................................................................................. 7411.7. Figure 42: Glyoxylic acid cycle ................................................................................................ 7512.1. Figure 43: The nitrogen fixation ............................................................................................... 7612.2. Figure 44: The synthesis of essential amino acids .................................................................... 7812.3. Figure 45: Transcription ............................................................................................................ 8012.4. Figure 46: tRNA ....................................................................................................................... 8112.5. Figure 47: The steps of initiation complex formation .............................................................. 8212.6. Table 7: Biogenic amines .......................................................................................................... 8612.7. Figure 48: Oxidative deamination (amino acids) ..................................................................... 8712.8. Figure 49: Entering of carbon skeleton of amino acids the citric acid cycl .............................. 8812.9. Figure 50: Synthesis of carbamide ............................................................................................ 9013.1. Figure 51: Steps of gastric acid secretio ................................................................................... 9413.2. Figure 52: cAMP system .......................................................................................................... 9613.3. Figure 53: The outline of the neourohormonal control of the carbohydrate metabolism and the blood-sugar level .......................................................................................................................................... 9713.4. Figure 54: The glucose transport between organs and its hormonal regulation ....................... 9814.1. Figure 55: The biochemistry of meat ripening ....................................................................... 10414.2. Figure 56: Changes of colour through the meat processing ................................................... 105


Tárgymutató


1. fejezet - 1. INTRODUCTION1. 1. 1. The object of biochemistry, its relationship with other sciencesBiochemistry is a life science between biology and chemistry. Its development is connected to these two disciplines tightly. The new results of chemistry and biology also appear in biochemical research, thus they enhance the development of this discipline. Considering the content of biochemistry it is closer to biology, while based on its methods it is closer to chemical sciences.

Biochemistry deals with the physiology of living nature, and of the organisms in molecular terms. Emphasizing the unit of living world the aim of its discipline is to recognize chemical processes taking place in all living organisms in a multi-faceted way.

Biochemistry examines the following areas:

• The structures, organizations and functions of living matter in molecular terms,

• The chemical structures of molecules constructing the living matter,

• Interactions taking place during the formation of supramolecular structures, cells multicellular tissues and organisms.

• Material and energy transport between living matter and its surroundings,

• The storage and transmission of information that is needed for self reproduction in cells

• Chemical changes accompanying the reproduction, aging and death of cells and organisms,

• Self controlling processes of chemical reactions in living cells, influencing factors of the direction of these chemical reactions

2. 1. 2. Relationship between biochemistry and other sciencesBiochemistry is a distinct discipline . The structures of biomolecules, the directions of metabolic pathways, and their regulatory mechanisms by enzymes can be understood based on the chemical laws only. Biochemistry is an interdisciplinary science. It is tightly connected in animal and plant physiology and organic chemistry. It applies the results of mathematics, physics, physical-chemistry and colloid chemistry. The results of biochemistry provide basis for more applied sciences such as medical science, pharmacology and toxicology. Dietetics, food industry and forage doctrine technologies, plant production, animal husbandry and environment protection can also use the results of biochemistry.


2. fejezet - 2. THE LIVING SYSTEMS1. 2. 1. Characterization of living systemsLiving systems differ from lifeless ones qualitatively. Lifeless systems tend to an intense disorder (entropy is growing), by the combination of various elements. Lifeless systems are characterized by the sum of compounds with small molecular mass. Living systems tend to maintain dynamic equilibrium with their environment, maintaining order as an independent unit.

Metabolic processes are important processes of life. They can be divided into two parts: Catabolism is the decomposition of organic matter and anabolism is the construction of organic compounds. Living beings take up and lose substances and energy continuously from their environment to ensure their metabolism and their survival. They are characterized by constant renewal. Their macromolecules are formed by various combinations of a few elements.

Major features of living organisms:

• They are self maintaining,. They keep their inner stability (homeostasis), peculiarities and individuality in spite of the change of their environment; they are in dynamic balance with their environment.

• They follow the change of their environment by modifying their metabolism.

The velocity and direction of biochemical processes can be changed by enzymes.

• They are open systems →there is an energy and matter replacement between living matter and its environment.

• They are economical (end product→intermedier→precursor). Catabolism and anabolism are in relationship, the intermedier or end product of a certain process can be the precursor of another process.

• They are capable of self-reproduction (DNA). Due to their capability of forming successors similar to them, life can be maintained. They have information carrying,

• Storing, reading, and copying systems. Information can be passed to their successors (DNA, RNA).

• There is uniform material construction (ATP). The same biochemical processes take place in living beings independently on their state of development. (e.g.: ATP)

• They have been continuously developed by evolution.

2. 2. 2. The composition of living matterThe composition of living organisms (both in the quality and in the quantity of the elements) differ from that of lifeless environment and the earth’s crust. Table 1. shows the frequency of elements in the earth's crust and in the human body.

2.1. ábra - Table 1: The frequency of elements in the earth's crust and in the human body


2. THE LIVING SYSTEMS

Among the elements in the earth's crust, oxygen, silicon and metals occur in the largest quantity. Living matter consists of four elements in 99%, these are hydrogen, oxygen, carbon and nitrogen. These four elements are called organogenic elements. Organogenic elements rank among non-metallic elements. There is a wide variety of their molecules formed by covalent bond. The existence of the numerous varieties of their molecules can be explained by the specific properties of carbon atom (it can form single and double bond as well with itself, nitrogen and oxygen)

Phosphorus and sulphur are also essential in the construction of the living matters. Phosphorus can be found mostly in ester ulinkage, while sulphur is attached to the carbon atom by covalent bond. Sulphur and phosphorus together with organogenic elements are called biogenic elements

To the normal life function of living organisms of other elements are also essential, but these occur in a much smaller quantity in them. Na, K, Ca, Mg belong to macro elements as they are present in plant in quantities more than 0.1% on a dry matter weight basis, and in humans and animals more than 0,005%. Cl, I, Fe, Zn, Mn, Co, Cu, Mo, Se, B are micro elements while their amount in human organisms is smaller than the above-mentioned quantities. Apart from the listed ones living beings contain other elements but these are not essential.

Their biological role is not known yet. The composition of molecular constituents that can be found in the living organisms are represented through the example of Escherichia coli bacterium (Table 2.).

2.2. ábra - Table 2. The chemical composition of Escherichia coli bacterium



Living organisms contain water in the largest quantities. Among their organic compounds lining organisms contain proteins, carbohydrates, nucleic acids and lipids in a significant quantity. The amount of the other types of organic matter is negligible compared to the mass of these four biomolecule groups.

These biomolecules (carbohydrates, lipids, proteins, nucleic acids) are well-separable structurally and functionally, though they have common properties as well. Their common feature is, that

they consist of monomer units. These monomer units are connected to each other by water loss reactions (condensation) creating macromolecules.

The monomer units of carbohydrates are monosaccharides that of nucleic acids are nucleotides. Proteins formed by the attachment of a lot of amino acids (monomers), while most of the lipids consist of the fatty acid monomers.

They are the main characters of metabolism processes. There are wide variety of proteins and nucleic acids. Carbohydrates and lipids do not have so many variants, the number of their monomer units and the variations of the ulinks are fewer. The features of biomolecules differ from inorganic molecules.

biomolecules: molecules of lifeless matter:

complicated diverse construction,

ordered structure

simple construction, disordered mixtures

energy taken up from the environment ensures the maintenance of the organization

energy taken up from the environment increases disorder

their structure is suitable for specific functions specific function is not recognizable

contain information,

are capable of self-reproduction

do not contain information,

are not capable of self-reproduction

Water has numerous functions in living organisms, due to its specific properties. (V-shape, polar molecule, hydrogen bonds between their molecules, amphoteric character, great specific heat and great vaporization heat.)

The role of water in biological systems:

• Water molecules hydrate macromolecules. Hydrogen bonds can be formed between water and macromolecules or macromolecules can possess charges thus polar water molecules surround them forming a hydration shell.



• It is solvent. Water dissolves many kinds of substances such as salts. Among non dissociating substances, water dissolves polar substances. Amphipathic molecules (containing polar and apolar parts) form micelles in water.

• It is a transfer medium, due to the fact that it is a good solvent, it takes part in mass transport between cells, tissues and organs

• It participates in many chemical reactions. It can be reactant or end product in biochemical reactions.

• It plays role in the regulation of heat balance. Due to its high specific heat and vaporization heat, water can buffer the changes of temperature in environment, thus the cells of living organisms can maintain a relatively constant temperature. Due to its high vaporization heat organisms can lose heat during sweating, protecting themselves from warming

In living organisms, inorganic ions are present in a small amount, although they are of great importance also. They have several functions.

The role of inorganic ions in biological systems:

• Enzyme activators: They influence the velocity of metabolic processes by activating or blocking catalysts of biochemical processes,

• Components of enzymes: Their deficiencies can inhibit the metabolism processes, which cause the accumulation of intermedier products,

• Participants in stimulus transfer,

• Regulators of osmotic potential, influencing water uptake and loss.

• Regulators of the acid-base equilibrium. Biochemical processes take part at certain pH values. Enzymes have pH optimum. Maintaining the acid-base balance in the living organisms is an essential function.

• Components of the compounds taking part in the oxidation-reduction processes,

• Components of hormones, they can influence the effect of hormones on metabolism.

• Hormone regulators,

• Constituents of multi cellular tissues and organs. They are often attached to organic matters.

Living beings can construct their own organic substances during metabolism (Figure 1). Reactants of Anabolism processes are always simple inorganic compounds (autrophic living beings) or organic compounds (heterotrophic organisms). From precursors intermediate products are formed in biochemical processes, which then transform to monomers. As monomers attach to

each other they form macromolecules that are the building blocks of the cells. During the interactions of macromolecules cell constituents with special functions are formed. They are the so called supra molecular systems, that are required for the special life processes of cells. Catabolism processes take place through similar steps (supra molecular systems, precursors). Anabolism processes require energy, while catabolism processes provide energy for the cell and the organism.

2.3. ábra - Figure 1: Molecular organizations in cells



Living organisms provide energy and precursors for their anabolism processes from their environment. They can be classified based on the source of matter and energy (Table 3.)

2.4. ábra - Table 3: Classification of organisms based on their mass sources


3. fejezet - 3. BIOMOLECULES I. CARBOHYDRATESCarbohydrates are organic compounds that contain carbon, hydrogen, and oxygen. Oxygen can occur in the groups of hydroxyl, ether or oxo-group. Hydrogen is usually in 2:1 ratio to oxygen.

Carbohydrates belong to the most important organic compounds from chemical and biological aspect too. They can be found in flora and fauna as well. Among nutrients, the carbohydrates are important energy sources.

One part of carbohydrates with low molecular weight (e.g. glucose, fructose and beet sugar) is nutrient, while carbohydrates with high molecular weight are either reserve nutrients (e.g. starch and glycogen), or support and frame substances (e.g. cellulose).

Being attached to other substances, carbohydrates create various compounds: e.g. nucleotides, alkaloids, heparin, and glycoprotein. Based on their structure carbohydrates are polyhydroxy-aldehydes or polyhydroxy- ketones, and their derivatives (e.g. their condensed products)

Classification of carbohydrates:

• monosaccharides (sometimes called simple sugars: glucose, fructose, etc.)

• di- and oligosaccharides (contain 2-8 monosaccharide units: sucrose, maltose, etc,)

• Polysaccharides (Carbohydrates with large molecular weight, containing hundreds or thousands of monosaccaharide units. They are not sweet: starch, cellulose, glycogen, pectin, etc)

1. 3. 1. MonosaccharidesMonosaccharides are monomer molecules, the simplest carbohydrates. They cannot be hydrolyzed to smaller carbohydrates. The monosaccharides are polyhydroxy-aldehydes or polyhydroxy-ketones without side chains. The formers called aldoses and the latter ketoses.

According to the number of carbon atoms monosaccharides can be classified as triose, tetrose, pentose, hexose and heptose.

They have sweet taste, they are crystalline substances solving well in water.

Except for dihydroxyacetone all of the m onosaccharides have at least one asymmetric carbon atom, thus m onosaccharides have optical stereoisomers. Stereoisomers have the same molecular formula, they differ only in their spatial arrangement, optical stereoisomers rotate the plane of polarized light in opposite directions.

Most of the naturally occurring monosaccharides have D configuration. D and L nomination refers to the configuration of the asymmetric carbon atom that is farthest from carbonyl group. If the OH group (in Fisher projection) attached to that asymmetric carbon atom points to the right the monosaccharide has D configuration, if it points to the left it has L configuration.

Figure 2 shows the structural formula of the most important trioses, pentoses and hexoses.

3.1. ábra - Figure 2: The most important monosaccharides


3. BIOMOLECULES I. CARBOHYDRATES

1.1. 3. 1. 1. Formation of cyclic monosaccharidesIn aqueous solutions, the open-chain form of a monosaccharide often coexists with a cyclic (ring) form. The ratio of open chain form is very small compared to cyclic form. For instance, more than 99% of D-glucose molecules are in cyclic form, while less than 1% are in open chain form.

Formation of ring structure is a reversible, nucleophilic addition reaction. The aldehyde or ketone carbonyl group carbon (-C=O) and the hydroxyl group (-OH) (the hydroxyl group is bound to the farthest chiral carbon atom from the oxo-group) react forming a hemiacetal with a new C-O-C bridge. Figure 3 shows the ring formation of D glucose. The hydroxyl group shown with red in the cyclic form called hemiacetal hydroxyl group.

Two types of stable rings can be formed during the conversion from open-chain form to the cyclic form:

• the five membered ring structure is called furanose ,

• the six-membered ring is called pyranose .

Aldohexoses form six-membered ring, while ketohexoses form five membered hemiacetal formation. Figure 3 represents the formation of cyclic D-ribose.

The atoms of pyranose ring are not in a plane, they have a chair structure.

By forming the pyranose ring a new chiral centre (the C-1 carbon atom) appears, that was not there in the open-chain structure.

The OH group on C-1 of the hemiacetal atom has two possible orientations: above or below the plane of the



ring, namely axial or equatorial. The axial group is perpendicular to the mean plane, while equatorial is parallel to the mean plane (Fig. 3.).

3.2. ábra - Figure 3: Formation of cyclic monosaccharides

1.2. 3. 1. 2. Chemical reactions of monosaccharides1.2.1. 3. 1. 2. 1. Redox reactions of monosaccharides

Compounds containing oxo-group or free hemiacetal-hydroxyl group are oxidized easily thus they are reductants. Reductant monosaccharides give positive Fehling’s test result and silvering process.

Sorbite can be found in fruits /e.g. apple/. It has sweet flavour, and it is suitable for diabetic people as sweetener, since its decomposition does not increase the blood-sugar level. In simple carbohydrates, as an effect of mild oxidation (e.g. aldehyde) the hydroxyl group on the end of the chain oxidizes to COOH- group. With the oxidation of the aldehyde group of the aldoses polyhydroxyacids form, while through oxidation of primaryhydroxyl group to carboxyl group uronic acid forms.

Uronic acids contain formyl group beyond carboxyl and hydroxyl groups. When both C atoms on the end of the chain transform to carboxyl group, glucaric acids will form.

Among uronic acids D-glucuronic acid, D-galacturonic acid, and D-mannuronic acid can be found in nature (Fig. 4).

3.3. ábra - Figure 4: Redox reactions of monosaccharides, of D -glucose



1.2.2. 3. 1. 2. 2. Transformations of monosaccharides into each other

The transformations of monosaccharides into each other are very important steps of the carbohydrate metabolism /e.g. anaerobic glycolysis, pentose phosphate cycle, Calvin cycle, etc./.

The main types of transformations catalysed by enzymes are the following:

• During the transformation, the number of the carbon atoms does not change.

• Epimerization: during this reaction, substituents restructure sterically around one single carbon atom (e.g. transformation of D-ribose to D-xylose.

• Isomerization: the aldose-ketose transformation, where the carbonyl group shifts onto the adjacent carbon atom (e.g. transformation of D-glucose to D-fructose.).

• The transfer of C3 unit /active dihydroxy-acetone/ or C2 unit /active glycol aldehyde / from one of the sugars onto another one.

Through these reactions we get trioses, tetroses, pentoses and heptoses. The donor of C3 and C2 units is always the ketose, while the acceptor is the aldose.

During the transformation, the sum of the carbon atoms in the monosaccharides does not change.

• Transaldolase reaction: in this process the enzyme splits off fructose or sedoheptulose and transfers the C3 unit onto the appropriate aldehyde.

(e.g.) C7 + C3 → C4 + C6

• Transketolase reaction:

C2 unit is transferred (The donor is the ketose) from one monosaccharide to another one.

(e.g. C5 + C5 → C3 + C7)

To the formation of C2 unit ketose phosphate is needed, which steric arrangement on C3 atom is equal to that of fructose. In this way, it is possible, that ribulose -5-P epimerizes to xilulose-5-P and than becomes C2 unit donor.

• Aldolase reaction: Hexoses are transformed to trioses or the reverse (Fig. 5).

3.4. ábra - Figure 5: Transformations of monosaccharides



• During the transformation, the chain is shortened with a carbon atom.

• Hexose-pentose transformation

During the oxidation, the aldehyde group is converted to carboxylic-group then the molecule was decarboxylase (Fig. 6).

3.5. ábra - Figure 6: Hexose-pentose transformation

2. 3. 2. DisaccharidesIn disaccharides, two monosaccharide units are attached together by splitting a water molecule off. The glycosidic ulinkage can be split by acidic or enzymatic hydrolysis. In disaccharides one of the components is always glucose. Disaccharides can be reducing or non-reducing. In reducing disaccharides, the hemiacetal hydroxyl group of a monosaccharide unit reacts with the alcoholic OH group of the other monosaccharide unit, with the loss of a water molecule. Thus, the molecule contains a hemiacetal hydroxyl group, which can reduce Fehling solution.

In non-reducing disaccharides both of the hemiacetal hydroxyl groups are involved in the glycosidic ulinkage,



therefore they can not reduce Fehling reagent.

2.1. 3. 2. 1. Reducing disaccharidesMaltose

In maltose the hemiacetal OH group of an a β-D glucose molecule reacts with the alcoholic OH group of another β-D glucose molecule. α (1→ 4) glycosidic ulinkage is formed between the two monomer units, as hemiacetal OH group has a position and it is on C-1 while the alcoholic OH group that is involved in the reaction is on C-4. There is a sharp bend at the glycosidic ulinkage in maltose. Maltose is formed by enzymatic decomposition of starch and glycogen. Maltose can be splitted to there monomers by maltase enzyme.

Cellobiose

In cellobiose the hemiacetal OH group of a b -D glucose molecule reacts with the alcoholic OH group of another b -D glucose molecule. β (1→ 4) glycosidic ulinkage is formed between the two monomer units. In cellobiose the second monomer unit is rotated 180o .

Lactose

In lactose a β-D galactose molecule combines with α-D- glucose through a β(1→ 4) ulinkage. The hemiacetal OH group of the glucose molecule is retained; therefore lactose is a reducing sugar. Lactose can be found in the milk of mammals (5,5- 8% in breast milk, 4,5-5,5% in cow milk. Lactose molecules are broken down by lactase enzyme in the gut. Lactose is of great importance in the production of dairy products made with fermentation (yoghurt, sour cream).

2.2. 3. 2. 2. Non-reducing disaccharidesSucrose

One of the most important nutrients. It occurs in sugar beet or sugar cane in dissolved state, it can be extracted from them.

In sucrose an α-D- glucose combines with a β-D- fructose molecule forming an α,β (1→ 2) ulinkage. This molecule differs from the above-mentioned ones, as t he glycosidic bond is formed between the reducing ends of both units , therefore it is a non-reducing sugar (Fig. 7).

3.6. ábra - Figure 7: Disaccharides

3. 3. 3. PolysaccharidesPolysaccharides are macromolecules of repeated monomers units (monosaccharide) joined together by glycosidic bonds. Polysaccharides do not have sweet taste, and crystallic structure. They do not solve in water or



their aqueous solution result in a colloidic suspension.

3.1. 3. 3. 1. Classification of polysaccharidesPolysaccharides can be classified according to there chemical structure and there biological function.

• Chemical structure: Polysaccharides can be divided into two classes, based on how many C atoms their monomer units have.

• Pentosans are composed of pentoses (e.g. xylan, araban)

• Hexosans are composed of hexoses (e.g. cellulose, starch)

• Biological function: Polysaccharides are of great importance as food reserves in plants or as structural components of plants. Some examples for the most important polysaccharides

• food reserves polysaccharides: starch, glycogen, inulin,

• structural polysaccharides: cellulose, chitin, pectin.

Starch

Starch is a plant reserve nutrient that is stored in plant seeds and tubers. Potato contains cca. 2o % starch, while grain seeds 55-7o %. Starch is the principal carbohydrate source for human nourishment and it is utilized by decomposing to glucose. Starch does not dissolve in cold water, swells up in hot water, and then forms colloid solution. Colloid solution turns into gel due to cooling.

Starch is a mixture of two types of molecules, the linear amylose (~ 20%) and the branched amylopectin (~ 80%).

Amylose

It consists of 100-300 α-D-glucose subunits involving exclusively α(1→4) ulinks, as they are in maltose.

The α(1→4) structure promotes the formation of a helix structure. In the helix spiral six glucose units are in a thread. The spiral is stabilized by intramolecular hydrogen-bonds. Amylose molecules consist of single unbranched chains.

Amylopectin

Amylopectin has greater molecular weight than amylose, it consists of more than thousand glucose units. In amylopectin α (1→4) bonds are dominant also, but 12-20 glucose units are always followed by α (1→6) acetal ulinkage, that causes branches in the molecule.

Glycogen:

Glycogen is a reserve polysaccharide in animals. It is synthesized and stored in the cells of the liver and the muscles. Glycogen has a similar chemical structure and size to amylopectin but it is more extensively branched and compact than starch.

Inulin

Inulin is a fructose polymer and it naturally occurs in many types of plants. In an inulin molecule 30-35 fructose units attached with 2→1 ulinks. There are some glucose molecules attached to the end of a fructose chain.

Cellulose

Cellulose is the structural component of the primary cell wall of green plants (trees, grasses). Trees comprise of 40-50% cellulose. The purest cellulose source in nature is cotton. Cellulose is a linear polymer of 1000-14000 ß-D-glucose molecules that are connected by β (1→4) glycosidic bonds (cellobiose). Cellulose is a straight chain polymer, no coiling or branching occurs.

In cellulose, the hydroxyl groups on the glucose from one chain form hydrogen bonds with oxygen atoms on the



same or on a neighbour chain, holding the chains firmly together side-by-side and forming microfibrils.

Chitin

Chitin constitutes a major structural material in the exoskeletons of many arthropods and mollusks. Chitin is a homopolymer of N-acetyl-β-D-glucosamines that are connecting with ß-1→4 bonds.

Pectin

Pectin is a structural heteropolysaccharide, a constituent of the primary cell wall of plants. Pectin is a polymer of D-galacturonic acid monomers with α-1→ 4 glycosidic bonds. Pectin is usually used as gelling agent in jams, jellies or fruit juice. Certain fruits contain especially large amount of pectin (e.g. currant, gooseberry, quince) (Fig. 8)

3.7. ábra - Figure 8: Polysaccharides


4. fejezet - 4. BIOMOLECULES II. PROTEINSProteins are one of the most important building blocks of living organisms. They have broad task. Their monomeric units are amino acids. Amino acids contain acidic carboxyl group and basic amino group as well.

In proteins 20 kinds of α-L-amino acids can be found. In these molecules, the amino group is ulinked to that carbon atom (α.) that is closest to the carboxyl group. Amino acids of proteins differ from each other in their groups that are attached to the α-carbon atom. The α-carbon atoms are chiral. Among the enantiomers amino acids with L-configuration occur in proteins. Amino acids (monomers) joined together by peptide bonds and create a polypeptide chain (Fig. 9).

4.1. ábra - Figure 9: Peptide bond

Atoms in peptide bond (C, O, N, H) are coplanar. Due to the π-bond and the delocalization of the nitrogens non-bonding electron pair the rotation is inhibited, and there is trans-spacing. The trans-spacing is of great importance in the stabilization of secondary protein structure

Proteins can be classified chemically and biologically as well. By chemical classification the nature of the R groups are taken into account. Thus we can distinguish monoamino monocarboxylic acids with non-polar side chains, etc.

Based on biological properties, essential amino acids can not be produced by our organism, thus we must use for protein synthesis those amino acids that are produced by other organisms.

We are able to produce non-essential amino acids from precursors. The ones that can be produced only by adults in an appropriate amount, from other essential amino acids belong to the semi-essential amino acids.

Polypeptide chains consisting of more than 100 amino acids are called proteins, and their structure is organized in four stages.

The primary structure of proteins is the amino acid sequence in it. Primary structure determines the properties of other structures as well.

Two types of secondary structure of the proteins are known: these are α-helices and beta-pleated structure. Secondary structure is stabilized by hydrogen bonds formed among the peptide bonds (amide groups). The formation of hydrogen bonds is facilitated by the trans spatial position of-peptide bonds.

The polypeptide chain’s three-dimensional structure (conformation) is called the tertiary structure of the peptide. From the aspect of tertiary structure there are globular and fibrillar proteins. Fibrillar proteins can be characterized by pure α-helix or beta-pleated structure. In general, fibrillar proteins contain only a few types of amino acids. Tertiary structure is stabilized by primary - and secondary bonds between the R-groups of amino acids. (Among primary bonds the ionic and the disulfide bonds can be found while among secondary bonds all kinds of them can be found).

The quaternary structure of proteins is a complex structure formed by the interaction of more proteins. (For example, enzyme complexes) Proteins are able to perform their biological function by formation of their


4. BIOMOLECULES II. PROTEINS

quaternary structure (Fig. 10).

4.2. ábra - Figure 10: Conformation of proteins

1. 4. 1. Proteins can be classifiedAccording to their shape and structure:

• Fibrillar (fiber, fibrous) structure is characteristic for the frame, the supporting connective tissue proteins (e.g. proteins of the muscles, wool, hair).

• Globular (spherical) structure: is characteristic for the enzyme proteins, blood proteins (albumin, globulins). Not only simple but complex proteins also belong to here (e. g. myoglobin, casein).

There are proteins that do not strictly fall into one of these groups, they are characterized by the transition between the two types. There are also proteins that can be found both in globular and fibrillar forms.

According to their composition:

• Simple proteins only consist of amino acids

• Complex proteins also contain other components beside amino acids They can be classified based on their non-protein part

The main groups of complex proteins:

GROUP NON-PROTEIN PART TASK

Hemproteins iron-porfirin oxygen and electron transport

Metalloproteins Metallic ions biocatalysts

Phosphoproteins Phosphoric acid Reserve nutrient

Lipoproteins lipids Lipid transport

Glycoproteins Carbohydrates protection

Nucleoproteins Ribonucleic acids protein synthesis


4. BIOMOLECULES II. PROTEINS

According to their solubility:

GROUP SOLVENT

Albumins → water, dilute salt solution

Globulins → dilute salt solution

Prolamins → dilute alcohol

Glutelins → dilute acids or alkalis

Histones → dilute acids

Frame proteins → insoluble

Albumins and globulins can be found in plants and animals as well. Prolamins and glutelins are characteristic proteins for the cereal grains. Histones are nuclear constituents. Frame proteins can be found in animals.

According to their biological function

GROUP TASK

Enzymes Catalyze biochemical processes

Hormones Regulate biochemical processes

Receptor proteins Bind and transport stimuli

Protective proteins Protect against injuries and infections

(thrombin, immunoglobulins)

Transport proteins Material transport

(hemoglobin: oxygen transport, transferrin iron transport)

Structural proteins Construct frame and connective tissue

(collagen, elastin, keratin)

Reserve proteins Fundamental sources of embryonic development (zein, casein, ovalbumin)


5. fejezet - 5. BIOMOLECULES III. THE LIPIDSLipids are organic compounds; they carry out important functions in the living organism. Some lipids are used for energy storage, since the higher animals and oilseed plants store their reserve energy in fats, in oils. Large fraction of lipids together with proteins are the components of membranes, so directly or indirectly can influence some vital functions of the cells.

Lipids are chemically diverse group of compounds that are classified together because of their apolar structures, which give them high solubility in apolar solvents. These compounds have very low solubility in the aqueous environment of the cell.

1. 5.1. Classification of lipids:1. Saponifiable lipids:

• Simple lipids: (Alcohol and carboxylic acid(s) are obtained by hydrolyzing them.)

• vaxes

• neutral fats and oils (triglycerides)

• Compound lipids: (Besides alcohol and carboxylic acid other compounds are also obtained by hydrolyzing them.)

• phospholipids

• sphingolipids

• glycolipids

2. Insaponifiable lipids:

• Steroids

• sterols (cholesterol, ergosterol)

• bile acids

• hormons

• steroidal glycosids (digitoxine, strophantin)

• steroid alkaloids (tomatine, solanine)

• Carotenoids (tetraterpenoids and derivatives)

• Lipid soluble vitamins (A, D, E, K)

1.1. 5. 1. 1. Saponifiable lipidsLipids treated with concentrated alkali such as NaOH or KOH, give sodium or potassium salt of the fatty acid, which are soluble in water.

The fatty acids:

The fatty acids are the simplest lipids. These carboxylic acids are constituents of many complex lipids.


5. BIOMOLECULES III. THE LIPIDS

The classification of fatty acids:

• saturated fatty acids

• lauric acid (C11H23 – COOH);

• myristic acid (C13H27 - COOH);

• palmitic acid (C15H31– COOH);

• stearic acid (C17H35– COOH);

• arachidic acid (C19H39– COOH);

• unsaturated fatty acids

• oleic acid (C17H33 – COOH);

The place of the double bond beyond carbon: C9

• linoleic acid (C17H31– COOH);

The place of the double bonds beyond carbons: C9, C12 ( Ώ 6)

• linolenic acid (C17H29– COOH);

The place of the double bonds beyond carbons: C9, C12, C15 ( Ώ 3)

• arachidonic acid (C19H31– COOH);

The place of the double bonds beyond carbons: C5, C8, C11, C14

Most of the naturally occurring fatty acids have a chain of an even number of carbon atoms, from 14 to 22. However, in milk fatty acids with lower number of carbon atoms and with odd number of carbon atoms can be found. In most of the naturally occurring unsaturated fatty acids the orientation around double bonds is cis (cis configuration).

The linoleic acid and the arachidonic acid are essential fatty acids, while the linolenic acid is semi-essential fatty acid, because the human body can synthesize it from linoleic acid.

1.1.1. 5.1.1.1. Vaxes

Waxes are esters of saturated or unsaturated long chain monocarboxylic acids (fatty acids) and long chain monohydroxy alcohols (fatty alcohols).

1.1.2. 5.1.1.2. Neutral fats and oils (triglycerides)



The neutral fats and oils are the triesters of glycerol and three molecules of fatty acids.

In fats the ratio of saturated fatty acids to unsaturated fatty acids are higher then in the oils. The higher the ratio of saturated fatty acids the higher the melting point of the fat.

The triglycerides with similar esterified fatty acids are called simple triglycerides, w hile two or three fatty acids are different in triglyceride, is called mixed triglyceride.

The melting point of the neutral fat of the animals living on the warmer climate is higher. The diet rich in carbohydrate favours the synthesis of the fats containing saturated fatty acids and having higher melting point.

Our organism stores the big part of energies in fat depots in the form of neutral fats. The fat is the major energy source in most cells. The metabolic oxidation of fat consumes more oxygen, (gram per gram) than oxidation of carbohydrate, so a correspondingly higher amount of energy is released.

The lipids need smaller space in the cell than carbohydrates, because opposed to carbohydrates all lipids are hydrophobic molecules and are not surrounded by a thin film of water.

1.1.3. 5.1.1.3. Phosphoglycerides

The phosphoglycerides are the major component of all cell membranes. These compounds are similar to the triglycerides with one important difference, namely one of the three fatty acids is substituted by a phosphate group. All these compounds can be considered to be derivatived of glycerol-3-phosphate. Compounds, which contain hydroxyl group (cholamine, choline, serine) can react with the phosphate group of phospholipid by an elimination of water molecule.

Cholamine (ethanol-amine): HO-(CH2)2-NH2

Choline: HO-(CH2)2-N+(CH3)

Serine: HO-CH2-(CH)(NH2)(COOH)

Inositol: C6H12O6

Phosphatides are amphipathic molecules. O ne end of the phospholipid molecule (the phosphate group with bounding alcohols) is hydrophilic (lipophobic)

and the other end (the long carbon chain part) is hydrophobic (lipophilic). Because of the amphipatic nature, they can form micelles, they are the components of membranes.



Two common phospholipids are cephalin (phosphatidylethanolamine) and lecithin (phosphatidylcholine). These two phosphpglycerides are the main components of membranes of the animal cells.

1.1.4. 5.1.1.4. Sphingolipids

The sphingolipids mainly occur in the brain, in the spleen, in the liver and in the blood. Sphingolipids contain sphingosine (long chain unsaturated amino-diol).

Beside the fatty acid, a phosphoric acid can also join to sphingosine. A fatty acid is ulinked via an amide bond to the amino group of sphingosine. The hydroxyl group is phosphorylated and this is esterified by other alcohol. Sphingosine is converted into a variety of derivatives to form the family of sphingolipids. Sphingosine and the long carbon chain of fatty acid are the apolar (hydrophobic) parts of the molecule, the other part is polar (hydrophylic).

1.1.5. 5.1.1.5. Glycolipides

Glycolipids occur mainly in the nerve tissue. Glycolipids are carbohydrate-attached lipids. In glycolipids besides the fatty acid, monosaccharide (often galactose), oligosaccharide molecule is connected by an O-glycosidic bond to the sphyngosine.

1.2. 5.1.2. Insaponifiable lipidsLipids treated with concentrated alkali such as NaOH or KOH, do not hydrolyze. If they hydrolize, the end products are not soluble in the water.

1.2.1. 5.1.2.1. Steroids

The common feature of the steroids is the gonane skeleton (four cycloalkane rings join to each other). The rings are not aromatic. The rings are non-planar, they exist in the chair or in boat conformation. Several substituents can be attached to the carbon atoms of the ring, these substituents vary by the configuration. Considering their chemical composition and their function exceptionally diverse molecules belong here.

5.1. ábra - the core of gonane

The sterols (cholesterol, ergosterol) are the precursor for the synthesis of the vitamin D. The cholesterol in a free state or in an ester form with fatty acids is a component of animal fats. It is the constituent of the animal fats. It appears in the bile and in the blood too.

The bile acids differ from each other in the substituents (containing carboxyl-group also) being attached to the sterane skeleton. In the bile, they can be found in the form of their salts. The bile acids emulsifies lipids, so facilitate their digestibility.

The steroid type hormones are the hormones of the adrenal cortex and the sexual hormones. These hormones are originated from the progesterone. We will deal with their task in a later chapter.

The steroidal glycosides have effect on cardiac muscle (digitoxine, strophantin). They enhance the contraction of the cardiac muscle. The steroidal glycosides are synthesized by plants. In these molecules, special sugars are bound to the steroid skeleton via a glycosidic bond.

The steroidal alkaloids (solanine, tomatine) are vegetal origin. A strong physiological effect characterizes them.

1.2.2. 5.1.2.2. Carotenoids



The carotenoids areproduced from isoprene molecules (five-carbon units). The tetraterpenoids (carotene) and their derivatives (beside C and H they contain heteroatom, xanthophyll) are important carotenoids. They are produced from 8 isoprene molecules and they contain 40 carbon atoms. The double bonds in these molecules are conjugated. That is the reason of their colours, so they provide one part of the colour molecules in plants.

1.2.3. 5.1.2.3. Lipid soluble vitamins

We will deal with lipid soluble vitamins in a later chapter in particular .


6. fejezet - 6. BIOMOLECULES IV. THE NUCLEIC ACIDSThe nucleic acids (DNA, RNA) are macromolecules. Their monomer units are nucleotides. The nucleotides can be hydrolysed into three parts: phosphoric acid, pentose, and N containing aromatic heterocyclic base (nucleobase) (Fig.11). The nucleotides joined by phosphodiester bonds between the 3’ hydroxyl of one sugar and the 5’ hydroxyl of the other.

DNA nucleotide RNA nucleotide

phosphoric acid phosphoric acid

pentose sugar deoxy-D-ribose D-ribose

nucleobase purine base: adenine, guanine;

pyrimidine base: cytosine, thymine.

purine base: adenine, guanine;

pyrimidine base: cytosine, uracil.

6.1. ábra - Figure 11: Nucleobases

Nucleobases (uracil, thymine, cytosine, and guanine) due to the migration of hydrogen atom and the double bond may undergo amide-imidic acid tautomeric shifts,

Which yields the lactam tautomer (lactim ↔ lactam tautomers). Due to the tautomerization process, the bases are capable to attach the 1’ carbon atom of the pentose by removal of water molecule. For the construction of hydrogen bonds, which are a major factor of stabilizing the spatial conformation of polynucleotide chain, the tautomerization process also is responsible.

The chemical structure and the connection of nucleotide monomers to polynucleotide chain is shown in Fig. 12.

In each nucleotide the nucleobase is attached to the 1’ carbon of the sugar via an N-glycosidic bond, while the phosphate group is attached to the 5' carbon atom of the ribose sugar via ester bond.

The connection between nucleotide units in chain is through a phosphate group attached to the hydroxyl on the 5’ carbon of one unit and the 3’ hydroxyl of the next one (by elimination of water). This forms phosphodiester bonds. In this way very long nucleic acid chain can be formed. At one end the nucleic acid always has free hydroxyl group on the 3’ carbon atom of the sugar (3’ end) and the other end of the molecule always has phosphoric acid connecting to the 5’ carbon atom of the sugar (5’ end), which is able to form other bonds.


6. BIOMOLECULES IV. THE NUCLEIC ACIDS

6.2. ábra - Figure 12: From nucleotide monomers connected to polynucleotide

1. 6.1. The deoxyribonucleic acid (DNA)1.1. 6. 1.1. The primary structure of deoxyribonucleic acid (DNA)The primary structure of nucleic acids (DNA, RNA) is determined by the sequence of connected nucleotides. The nucleotides containing different N-bases can be attached in an optional order. The sequence of the bases determines the information available for building the living material.

The DNA consists of two long antiparallel polynucleotide chains. The two strands form a spiral called a double helix. The strands run in opposite directions and they are coiled in a right-handed manner about the same axis. The DNA double helix is stabilized by hydrogen bonds between nucleobases of nucleotides opposite each other. In the DNA the mole ratio of purine bases (adenine, guanine) and pyrimidine bases (cytosine, thymine) is equal (1:1).

A + G = T + C, which is possible when opposite of the nucleotide containing purine base can settle only nucleotide containing pyrimidine base. The consistent space filling may evolve in that way. The distance of the two chains is constant throughout the entire DNA molecule. The diameter of DNA is 2 nm.

In DNA the number of nucleotides containing adenine and thymine and the number of nucleotides containing guanine and cytosine are equal (A=T; G=C). This law can be explained by the secondary structure of DNA.

1.2. 6.1.2. The secondary structure of DNAIn DNA the sequence of the nucleotides of one chain defines the sequence of the nucleotides in the other chain. Opposite of the nucleotide containing adenine can settle only nucleotide containing thymine, while opposite of nucleotide containing guanine can settle only nucleotide containing cytosine. Adenine and thymine are forming two hydrogen bonds, while guanine and cytosine are forming three hydrogen bonds. The number of possible hydrogen bond is increased by forming of lactam tautomer.

The two strands are described as complementary to one another, following from the rule of base pairing. Two linear strands run in opposite direction to each other, the 5’ end of the one chain is in connection with the 3’ end of the other chain.

The double strand of DNA twists together to helical form. The phosphodiester-ulinked sugar residues form the backbone of the nucleic acid molecule and are on the outside of the helix.

The nucleobases are inside of the helix protected by the sugar phosphate groups (Fig. 13). In DNA ten base-pairs are in each turn of the helix.



6.3. ábra - Figure 13: The deoxyribonucleic acid.

The DNA in the nucleus of eucaryotes bounds to histone proteins forming the nucleosome (histon + a part of DNA’s chain = nucleosome). The nucleosomes are protected and stabilized by additional histone proteins from outside. The DNA inside the nucleus is organized by histone into DNA-histone nucleoprotein complex known as chromatin.

1.3. 6.1.3. The biological function of DNA DNA is responsible for self-reproduction of the living organism. DNA is able to produce an exact copy of itself and at cell division (mitosis, meiosis) information is passed from an organism to its descendents. Through this feature DNA is capable for transmitting of unique and racial characteristics of organism to successor generations.

Genetic information in DNA is stored in „codons” (in the form of a coded sequence of bases). The sequence of codons on a DNA tells the cell the sequence of amino acids in a protein. DNA directs the protein synthesis. DNA determines the number and the connection sequence of amino acids of the proteins in living organisms.

2. 6. 2. The ribonucleic acids (RNA-s)RNA-s consist of long, single-stranded, unbranched chain of nucleotides. RNA forms an A-form helix, but may have some double-strand regions too, as a consequence of self-complementary. The RNA the double-strand regions often folds RNA into three-dimensional structures. RNA-s take part in protein synthesis directly. According to their biological tasks and their role in the protein synthesis, there are three types of RNA-s:

• messenger ribonucleic acids (mRNA),

• transfer ribonucleic acids (tRNA),

• ribosomal ribonucleic acids (rRNA).

RNA-s differ in molecular weight, in composition of nucleotides, and their three-dimensional structure may also be different. The quantity of RNA-s in the cells are multiple (5-10 fold) compared to that of DNA.

2.1. 6. 2.1. The messenger RNA-sMessenger RNA-s (mRNA) carry coded genetic information about a protein sequence to the ribosomes for



synthesis of new proteins. In the cells hundreds of thousands of different proteins are synthesized, and it requires the same kinds of mRNA-s also. The mRNA (like other RNA-s) is produced in the nucleus in the process of transcription. (Transcription: is the process of creating an RNA copy of a sequence of DNA.)

It is characteristic to mRNA-s that they can be synthesized quickly, but they can break down also quickly so they have very short lifetime. The RNA molecule consists of 300-3000 nucleotide units. The sequence of nucleotides (bases) of all mRNA-s is complementary with the sequence of nucleotides (bases) of the appropriate section of DNA, so it determines the amino acid sequence of protein (translation). The RNA synthesized from DNA may contain non-informal parts (pre-mRNA).

The pre mRNA during its migration in the cytoplasm (the pre-mRNA must get out of the nucleus to the place of protein synthesis, being in the ribosomes in the cytoplasm) goes through a ripening process, when the non-informal parts are torn out of it.

2.2. 6. 2. 2. Transfer RNA-stRNA-s have the smallest molecular weight of all nucleic acids. They consist of 70-100 nucleotide units. tRNA-s have highly ordered structure, the single strand is folded into a three-dimensional structure stabilized by hydrogen bonds between base pairs. The secondary structure of tRNA usually is visualized as the cloverleaf structure. At the developing of three-dimensional structure, the loops are created where no hydrogen bonds are formed. Each loop or chain’s end has an important role. Free nucleotides (nucleotide triplet) remain in the loops, which ones and also the chain’s ends have important role in the biological tasks of tRNA-s. The tertiary structure is described as L-shaped.

The function of tRNA-s is to carry the activated amino acids to the site of protein synthesis. There are specific tRNA-s for each amino acids. There are at least 20 kinds of tRNA-s needed to the transport of 20 kinds of amino acids. (Some amino acids belong to diverse tRNA-s.)

For transportation to the ribosome, an amino acid is joined to its specific tRNA. This process is directed by a three-nucleotide sequence in one end of tRNA (anticodon).

The common feature of tRNA-s is that all of them have cytosine-cytosine-adenine nuleotide triplet at the 3’ end of the chain, where the amino acid is connected (binding site). (Fig. 14)

6.4. ábra - Figure 14: The mRNA and the tRNA

2.3. 6. 2. 3. Ribosomal RNA-sRibosomal RNA-s comprise up to 50-65% of the total ribonucleic acids in the cells. Several types of rRNA-s (5) are possible to distinguish. The rRNA molecules generally have single-stranded polynucleotide chain, but may contain unordered and double-strand regions too. The different rRNA types can be detached on the basis of their sedimentation coefficient. The ribosomal RNA with proteins forms the subunits of the ribosome (ribonucleoproteins). The ribosomes consist of two units, the large subunit and the small subunit. The two subunits are associated in the presence of Mg ions.

The most important task of ribosomes is to supply the corresponding structure for binding of mRNA and



binding of tRNA-s (carrying the amino acids) because the protein synthesis takes place on the surface of ribosomes.

3. 6. 3. Nucleoside triphosphatesThe nucleotides in the cells act not only as the monomers of the nucleic acid, but also as an energy source. In the nucleotides more phosphate groups can be joined together by phosphoanhydride bonds. To the formation of these bonds a great amount of energy is needed, which is produced in the metabolism. Energy stored in ATP is released upon the hydrolysis of the anhydride bonds and this energy is consumed in the anabolic pathway and in other energy-consuming processes (ATP ↔ ADP ↔ AMP).

Nucleotides: ATP (Adenosine-triphosphate, contains adenine), UTP (Uridine-triphosphate, contains uracil), GTP (Guanosine-triphosphate, contains guanine), CTP (Cytidine-triphosphate, contains citosine).

In the important biochemical processes, the enzymes carry out their activities together with the cofactors (NAD+, NADP+, FAD, CoA). These cofactors are analogous with the nucleotides or with the compounds containing nucleotides. We will write about them in the following section.


7. fejezet - 7. BIOACTIVE COMPOUNDS I. VITAMINS Vitamins are biologically active organic compounds that are essential to the biological processes of animal and human beings. Humans and many other organisms can not synthesize these essential compounds or can synthesize in small amounts and therefore must obtain from their diet, either the vitamin itself or a compound from which the required vitamin can be synthesized.

Vitamins do not provide energy, but they are essential for the material and energy flows. Sometimes one compound is essential vitamin for one organism, while another species is able to produce it. For example, ascorbic acid (vitamin C) is a vitamin for humans, monkeys and guinea pigs, but is not vitamin for most other animals. Other species studied to date can be synthesized vitamin C from glucose. Vitamins as a unified group can be formed only biologically because their chemical structures are very diverse and different.

If a particular vitamin is permanently missing from the diet, the organism will suffer from a deficiency and problems will appear in its functioning. Mostly growth disorders may occur. This phenomenon is named hypovitaminosis.

The more severe version of this is when (usually) typical clinical picture develops, which can be cured by dosage of vitamin. This phenomenon is named avitaminosis.

Dysfunction, hypervitaminosis may also occur if very large amount of vitamin is introduced into the body.

Food often does not contain the active form of vitamin, but also contains the precursor, from which the body is able to produce the active form of vitamin by using the converter mechanism. These compounds are called provitamins in the literature.

Organic compounds that inhibit the absorption or actions of vitamins are called antivitamins. The structures of antivitamins are very similar to the structure of vitamin.

Classification of vitamins

1. 7.1. Physiological effects of vitamins:The vitamins can be absorbed from the digestive system and as catalytic or regulating factors can join in the vital processes.

One part of vitamins is ulinked to proteins and affect as a component of coenzyme or prosthetic group of an enzyme. These vitamins are called prosthetic vitamins.

The inductive vitamins are identified as those of the vitamins that are essential for living organisms but their physiological role is not fully understood.

The vitamins can be divided into two classes based on their solubility. Thus, we distinguish lipid-soluble vitamins (vitamins A, D, E and K) and water soluble vitamins (B vitamins, vitamins H, C, P, pantothenic acid, folic acid).

2. 7. 2. Lipid soluble vitaminsPhysiological effects of lipid-soluble vitamins have been known, but their function in metabolic processes of the cells is only poorly understood.

In their absence, the activity of enzymes changes in certain animal tissues. It is believed that the fat-soluble vitamins regulate the biosynthesis of certain proteins. The lipid-soluble vitamins can be stored by the organism, so their lack develops later or takes shape more rarely. Hypervitaminosis also occurs, especially in the case of vitamin A and D.


7. BIOACTIVE COMPOUNDS I. VITAMINS

Today people often try to cover their vitamin needs by synthetically produced vitamin products. In this case an overdose may often occur.

At the time of the natural foods consumption, the appearance of the hypervitaminosis can hardly be imagined.

Vitamin A (retinol)

Vitamin A or retinol is a primary, unsaturated alcohol of molecular formula C 20H30O. They consist of β-ionone ring and the connected side chains of isoprene unit. An alcoholic hydroxyl group is attached to side chain. Vitamin A aldehyde or carboxylic acid may be formed by the chemical oxidation of this group (-OH), which are also bioactive compounds. The hydroxyl group (-OH) of vitamin A can form an ester with fatty acids. The resulting compounds often outweigh the biological activity of vitamin A.

Vitamin A occurs only in the animal world, while in plants the precursor or provitamin is found in the pigments called carotenes. The vitamin A1 may be formed from 12 different carotenoid compounds (for example: α-, β-, γ-carotene, criptoxantine) in animals by the enzyme of carotinase. The conversion is the most efficient from β-carotene.

The recommended daily intake of vitamin A is ranged from 0.8 to 1.5 mg, or 5-9 mg of β-carotene per day.

Vitamin A is necessary for cell growth and differentiation, skin and cellular health. With the protection of mucosa of respiratory organs protects the body against invading pathogens. Vitamin A affects the development of bones and teeth and is responsible for their protection.

One of the most obvious consequences of deficiency of vitamin A is the night-blindness. This is the inability to see in dim light. The effect of vitamin A in this process is accurately known.

The rhodopsin in photoreceptor cells of the retina is decomposed to opsin and 11-trans-retinal. This change generates nerve impulses in the central nervous system, which develops a sense of vision. In order to the resynthesis of rhodopsin the 11-trans –retinal have to be retransformed into 11-cis retinal. This process is multi-stage, first 11-trans retinol is formed by reduction, than in the isomeration process 11-cis retinol is issued. In the following oxidation process, 11-cis-retinal is re-produced again by the retinol-dehydrogenase enzyme and with connecting to opsin forms rhodopsin. This reaction sequence does not proceed quantitatively; the resulting loss can be replaced from the 11-cis-retinal derived from vitamin A of the blood (Fig. 15).

The best sources of vitamin A are cod-liver oil and other fish-liver oils, animal liver and dairy products (milk, butter), egg-yolk. The fruits and vegetables (carrot, spinach, pumpkin, cantaloupe) contain large quantities of carotenoids.

The overdose of vitamin A may cause hair loss, skin inflammation, fatigability beside pain in limb, malaise, upset stomach, lips and skin dryness, cracking.

7.1. ábra - Figure 15: Vitamin A

Vitamin D (Calciferols)



Calciferol is a collective noun. Several compounds with the same biological effects but with different chemical structure belong to the group of calciferols.

Vitamin D1 is called calciferol and lumisterol, vitamin D2 is called ergocalciferol, while vitamin D3 is called cholecalciferol.

Vitamin D controls the absorption and incorporation of calcium and phosphorous into the bones. In the absence of vitamin D „rachitis” symptom may occur. The patient's bones become soft, bent under the weight of the body. The joints become thick, the teeth break off.

Sources of vitamin D are fish liver oils, egg-yolk, yeast, wheat germ, champignon, oat-flake, where mainly provitamins of vitamin D can be found in large quantities.

Vitamin D is produced by the action of ultraviolet light from ergosterine (with plant origin) and from 7-dehydro-cholesterol (with animal origin), therefore vitamin D can be said to be produced from its provitamins (also in the skin).

The recommended daily intake of vitamin D is 10 μg (up to 20 years) and for adults 5μg.

The overdose of vitamin D may cause brittle bones, calcium level become higher in the blood, or may cause atherosclerosis.

Vitamin E (tocopherols)

In the body Vitamin E functions as an antioxidant, inhibits the oxidation of fatty acids, membrane lipides, provitamins and vitamins (vitamins A, C, carotene).

It plays a role in neurological functions. Vitamin E is needed for race preservation, because it promotes the fertilizing ability and fecundity, promotes the germ cell proliferation, growth and development of the fetus.

Vitamin E has anti-inflammatory effect, reduces the permeability of capillary blood vessels, and influences collagen formation as well.

Tocopherols consist of a chromanol ring and a hydrophobic side chain, which is a phytil (allows for penetration into biological membranes). Some versions differ in the groups (their number and location) connecting to chromanol ring.

Tocotrienols have the same structure as the tocopherol, but there is a double bond in the hydrophobic side chain, which consists of three isoprene units.

Vitamin E occurs only in the plant world. Tocopherols are synthesized in the chloroplasts of green plants and are transported to the seeds, to the lipid storage part of plant.

The recommended daily intake of vitamin E is 5-15 mg. The main sources of vitamin E are wheat germ oil, sunflower oil, safflower oil, nuts, nut oils (almond, hazelnuts), leafy green vegetables (spinach, turnip, beet), and avocados. (Fig. 16)

7.2. ábra - Figure 16: Vitamin D, E, K.



Vitamin K (Phylloquinones)

Vitamin K deficiency can cause severe bleeding, it is essential for blood clotting. Vitamin K helps the synthesis of the coagulation factors II., VII., IX. and X.

The Vitamin K1 and K2 are 2-methyl-naphthoquinone derivatives. In the vitamin K1 there is a phytil, while in the vitamin K2 there is a prenyl side chain.

The recommended daily intake of an adult of vitamin K is 1-4 mg, what the diet and the synthesized quantity (vitamin K2) by the intestinal flora living in the digestive system is usually able to cover. Vitamin K1 mainly occurs in the plants, whereas the vitamin K2 is found in animal food. The main sources of vitamin K are green leafy vegetables, cabbages and liver.

The fat-soluble vitamins have different sensitivity to the environmental effects. We have to pay attention to the reactivity of the vitamins during the storage and processing of food in order to get appropriate amount of vitamins into our body.

3. 7. 3. Water-soluble vitaminsThe group of prostethic vitamins belongs to water-soluble vitamins. Binding to proteins, they can influence the biological processes in the organism as the component of enzymes, coenzymes or prosthetic groups. They can not be stored in the body, so the vitamin deficiency can develop quickly, but for this reason, symptoms of hypervitaminosis do not occur. The water-soluble vitamins do not have provitamins, therefore we have to put on the active compounds in our food.

Vitamin B1 (thiamine)

Thiamine was the first of the B vitamins to be identified and that is why it is called vitamin B1. Thiamine contains a substituted pyrimidine ring as well as a five-membered ring containing nitrogen and sulphur. It plays important role in carbohydrate metabolism; in the absence of vitamin B1 intermediate metabolites (lactic acid, pyruvic acid) accumulate in the tissues and in the blood. Vitamin B1 is a constituent or prosthetic group of several enzymes such as pyruvate decarboxylase and transketolases as thiamine-pyrophosphate (TPP). Vitamin B1 enhances nervous system functions, and has an important role in normal functions of muscles and heart.

Deficiency of vitamin B1 may cause “Beriberi” disease (disease of the nervous system include weight loss, emotional disturbances, peripheral neuritis, muscular dystrophy).

The recommended daily intake of vitamin B1 of an adult is 1.5-2 mg.



The main source of vitamin B1 are yeast, whole grains, vegetables, fruits, meat (pork, fish) liver, milk, egg, and butter.

Vitamin B2 (riboflavin)

Vitamin B2 as a central component of the flavin enzymes (cofactors) influences the metabolic processes. Vitamin B2 acts as a hydrogen transfer in the oxidation processes. Vitamin B2 is an isoalloxazine derivative, contains a fused three-ring molecule called flavin. One of the nitrogen atoms of this ring system is substituted with a five-carbon sugar alcohol, ribitol.

Vitamin B2 can be found as riboflavin-5’-phosphate (FMN: flavin mononucleotide or as FAD: flavin adenine dinucleotide) in the prosthetic group of flavin enzymes classified in the dehydrogenases.

The FAD is formed with a connection of two nucleotide units through anhydride bond.

The functional part of FAD is the isoalloxazine ring, which serves a two-electron acceptor (reversible process).

The recommended daily intake of vitamin B2 of an adult is 1.5-2 mg. Source of vitamin B2 are venison, yogurt, soybean, milk, mushroom and spinach. The lack of vitamin B2 may cause cracked and red lips, inflammation of the lining of mouth and tongue; the eyes may become bloodshot, itchy and sensitive to bright light.

Vitamin B3 (nicotinic acid amid, niacinamid, vitamin PP)

Natural materials include nicotinic acid amide. Vitamin B3 expounds its biological effect as a component of oxidoreductase enzymes. It is a part of NAD+ and NADP+. The uptake and release of electrons and protons happens at the nicotinic acid amide part of the molecule.

The NAD + and NADP+ as a coenzyme of many dehydrogenase enzymes take part in the oxidation or reduction processes of a wide variety of compounds (Fig. 17).

The recommended daily intake of vitamin B3 is 10-20 mg. The lack of vitamin B3 besides pellagra disease may cause stomach and intestinal disorders.

7.3. ábra - Figure 17: Vitamins (B1, B2, B3)

Vitamin B6 (pyridoxine group)

The pyridoxol, the pyridoxal and the pyridoxamin are three natural forms of vitamin B6. All of these forms can



be found in phosphoester form in the body and can be easily transformed into each other. They are converted in the human body into a single biologically active form, pyridoxal 5-phosphate.

The phosphoric acid is bound to the CH2-OH group of the 5’ carbonic atom of the ring trough ester bond. Vitamin B6 has biological effect on the amino acid-protein metabolism and carbohydrate metabolism. It is a part of many enzymes (amino transferase, amino acid decarboxylase, phosphorylase).

The recommended daily intake of vitamin B6 of an adult is 2-3 mg. The lack of vitamin B6 may cause dermatologic and neurologic changes (seborrhoeic dermatitis-like eruption, ulceration, conjunctivitis, somnolence, confusion, neuropathy).

Vitamin B5 (pantothenic acid),

Pantothenic acid is the amide between pantoate and β-alanine.

Vitamin B5 has biological effect as the component of transacylase (CoA-SH). The CoA-SH is also a type of dinucleotide coenzyme. In the CoA-SH the pantothenic acid is connected as panthetein-4-phosphate to adenosine-3, 5-diphosphate.

The functional part of the coenzyme is the –SH group, which reacts with one or other carboxyl group of organic acid and produces thioester and water. In the absence of vitamin B5 fatigue, restlessness and muscle spasms may occur.

Vitamin B9, Folic acid (pteroil-glutamic acid)

The biologically active form of folic acid is tetrahydrofolic acid (FH4), which is a multi-reduced derivative. The biochemical role of tetrahydrofolate coenzymes is in the methylation processes.

Vitamin B9 (together with vitamin B12) has effect on forming of red - white blood cells, platelets. In the absence of vitamin B9 anemia may occur. This vitamin plays an important role in the forming of mucosa of the digestive system and in the synthesis of nucleic acids.

Vitamin B12 (cyanocobalamin)

Vitamin B12 has biological effect (often with folic acid) as the coenzyme component of enzymes (e.g.: methyl-malonyl-CoA mutase). In the absence of vitamin B12 complaints of the nervous system, pernicious anemia, lack of appetite, weakness and digestive complaints may occur.

Vitamin –H (Biotin)

Biotin is composed of a sulphur-containing ring part and valeric acid side chain. In nature, its amide (formed with lysine), biocytin occurs.

Biotin takes part in carboxyl-transfer as a prosthetic group of enzymes, which catalyse the transfer of CO 2 and HCO3

-. These enzymes require energy (ATP) and Mg2+-ions during their operation, thus CO2 temporarily connects to the N atom of the biotin. The activated carbon-dioxide connects to the proper acceptor in the form of carboxyl group as a result of the function of carboxylases.

This vitamin influences the protein, carbohydrate and lipid metabolism as well.

The recommended daily intake of vitamin H is 100-300 μg. The lack of vitamin H may cause hair loss, conjunctivitis, dermatitis, neurological symptoms, like depression, lethargy.

7.4. ábra - Figure 18: Water-soluble vitamins



Vitamin C (Ascorbic acid)

Vitamin C is a carbohydrate derivative. Dienol group can be found in the molecule, which causes the acidic properties of the compound. It can be found in oxidized and reduced form in cells and it forms redox system. The oxidized form is the dehydroascorbic acid, the reduced form is ascorbic acid.

Vitamin C plays an important role in cell respiration. It is essential for the repair and maintenance of connective and bone tissues, for recovery of wounds and fractures (vascular function).

Its biological effect is related to its oxidation-reduction ability. Vitamin C promotes the absorption of iron and calcium in the digestive tract, acts as hydrogen donor in the biochemical processes, is responsible for maintaining the reduced state.

Vitamin C plays a role in synthesis of collagen, adrenal hormones, serotonin and at the breakdown of tyrosine.

Vitamin C requirement is 80 mg, according to literature data, while fivefold quantity is recommended by others.

In case of lack of vitamin C chronic pain in the limbs or joints may occur. Deficiency will tend to bruise easily, have a negative impact on general healing of wounds. Deficiency can cause deterioration of the gums, bleeding gums may occur.

The main sources of vitamin C are vegetables, fruits, some animal products, plucks (liver, muscle).

7.5. ábra - Table 4: The role of vitamins in the function of enzymes




8. fejezet - 8. BIOACTIVE COMPOUNDS II. HORMONS In general hormones are organic compounds produced in endocrine glands, which regulate the chemical processes in the organism. The endocrine glands release their secretion directly into the bloodstream or lymphatic system, thus hormones can also affect other exterior parts of the organisms. They regulate the function, development, mass transport and growth of other organs. It is characteristic of hormone-producing glands that their secretions enhance or inhibit one another, thus they can be synergists or antagonists to each other.

The maintenance of inner balance (homeostasis) is ensured by the integrated system of nervous system and hormone-producing glands (neuroendocrine system) by the following process:

stimulus → excitation → regulator compound → enzyme → biochemical reactions

The hormone production is usually a result of nerve impulses, and it starts in the endocrine glands. The produced hormones get to the target cells via bloodstream, which receptors are capable of recognizing and binding them. The hormone-producing glands are also under hormonal control. This control is carried out by a three-level hierarchical system.

The operation of endocrine system is controlled by the diencephalon. Nerve impulse causes the secretion of neosecretum by hypothalamus (the part of diencephalon) that stimulates hormone production of the pituitary gland. According to it glandotrop hormones produced in anterior pituitary induce and stimulate the hormone production of other endocrine glands. The excess hormones in the bloodstream have effect on the hormone production of hypothalamus and pituitary as well (negative feedback). (Fig.19)

8.1. ábra - Figure 19: Regulation of hormone production

Endocrine glands

• Pituitary gland (hypophysis)

• Pineal gland

• Thyroid gland

• Parathyroid glands

• Adrenal glands


8. BIOACTIVE COMPOUNDS II. HORMONS

• Pancreas (islets of langerhans)

• Ovary (female)

• Testes (male)

1. 8.1. Classification of hormones: Hormones produced by endocrine glands can be classified based on their chemical structure, the type of metabolism affected by them or their mechanisms.

According to their chemical structure hormones can be

• Derivatives of the amino acids (adrenaline, melatonin, tiroxine)

• Peptides, polipeptides, protein (hormones o pituitary)

• Steroids (adrenal cortex, hormones of testicle, ovary)

According to the type of metabolism

• lipid metabolism

• carbohydrate metabolism

• protein metabolism

According to their mechanism

• Amino acid and peptide type hormones: bind to the cell membrane and start up the synthesis of compounds. These materials stimulate enzymes in the cell.

• Hormones that after entering the cell react with molecules inhibiting DNA in the nucleus. The transcription from DNA and the enzyme protein synthesis can begin (usually steroid hormones).

1.1. 8.1.1. Hormones of hypophysisHypophysis consists of three lobes: the anterior pituitary (adenohypophysis), posterior pituitary (neurohypophyseal) and the middle (pars intermedia) pituitary.

1.1.1. 8.1.1.1. The anterior pituitary (adenohypophysis)The defect or removal of anterior pituitary reflect in the operation of all organs to a smaller or larger extent.

Their Hormones:

• Growth hormone (somatotropic hormone, STH), that is a long- polypeptide chain consisting of 191 amino acids. It is responsible for the formation of genetically determined body size and body shape. It has an effect on protein transfer too. It enhances the uptake of amino acids by tissues, decreases the blood sugar level. It mobilizes the breakdown of fat tissue, thus fatty acid concentration increases in the plasma. Heat production enhances also.

• Adrenocorticotropic hormone (Corticotropin, ACTH) consists of 39 amino acids

It stimulates the hormone production of adrenal gland, thus it influences the amount glucocorticoids (cortisol, corticosterone)

• Thyroid-stimulating hormone (thyrotropin, TSH)

It belongs to glycoproteins. It has large molecular weight and it is sensitive thermally.



It regulates the hormone production of thyroid (T3 and T4). It affects the growth of the thyroid gland and iodine metabolism. It also influences the metabolism of connective tissue (eye socket).

• Gonadotropic hormones (hormones affect sex glands)They have central role in the regulation of reproduction. They organize and control structural and functional changes in female and male body. .

Glycoproteins

• This group includes Follicle-stimulating hormone (FSH). It is responsible for the stimulation of follicular development and the maintenance of spermohistogenezis.

• Luteinizing Hormone (LH) affects the hormone production of ovary and testicule

• Prolactin (mammotrope hormone PRL) has an effect on mammary glands, it can increase milk supply.

• Lipotrop hormone (LPH) consists of 90-amino acid peptide and affects lipid metabolism.

1.1.2. 8.1.1.2. Hormones of intermediate lobe (pars intermedia)

The color adjustment of some animals is a protective response in the fight for survival. It is important for them whether their skin is light or dark, which is affected by hormone of the intermediate lobe.

The melanocyte stimulating hormone (MSH) is a polypeptide that affects the pigmentation of the skin.

1.1.3. 8.1.1.3. Posterior lobe (neurohypophysis) hormones

In the case of destruction of the posterior lobe urination increases. Posterior lobe is in close structural and functional relationship with the hypothalamus. The lobe stores hormones of the hypothalamus. These hormones are cyclic peptides constructed by 9 amino acids.

• Vasopressin (antidiuretin, ADH) maintains the water balance of the body, it increases water retention in the collecting ducts of the kidney nephron.

• Under pathological conditions it also plays role in the regulation of blood circulation. Oxytocin increases the contraction of uterine smooth muscle and the outflow of the milk from mammary glands. It helps the sperm to enter the uterus. Its effect on uterine musculature is enhanced by estrogens while decreased by progesterone.

1.2. 8.1.2. Hormones of pineal glandThe function of pineal gland declines from sex maturity.

Melatonin is its hormone (acetylamino metoxitriptamin). It is formed by serotonin (Figure 20). The synthesis of melatonin and the activity of enzymes required to it fluctuate with the daily changes of light and darkness. Melatonin has an effect on skin color and its change.

8.2. ábra - Figure 20: The synthesis of melatonin



1.3. 8.1.3. Hormones of the thyroid glandsThyroid plays important role in regulation of the organism energy transfer and in iodine household. It is the only endocrine gland that can store its secretion. Its hormones are T3 (triiodothyronine) and T4 (thyroxine).

• Thyroxine is formed in much larger quantities, but triiodothyronine has greater efficiency. When thyroid has an increased hormone production (hyper tireoidism) due to the increased basal metabolism the organism loses weight, the velocity of blood flow increases, the heart rate increases. Thyroid gland also increases.

Thyroid hormones affect the growth and the development of organs and organ systems. They enhance oxygen consumption and their basal metabolism.

They increase the breakdown of stored carbohydrates and glucose consumption of the cells. Protein degradation is also increased by them. They facilitate the conversion of amino acids to glucose (process of so called gluconeogenesis), which is reflected in the raise of blood sugar level. They affect the salt and water balance of the organism as well. In case of decreased thyroid activity water and salt content of the organism are increased.

They increase fat mobilization in fat metabolism, and enhance the construction of fatty acids from carbohydrates. They also stimulate the synthesis of cholesterol in liver.

• Calcitonin (tireokalcitonin, TCH) is a peptide hormone that consists of 32-amino acids. It regulates the level of calcium. It reduces the blood calcium level, thus assimilation of calcium into bones increases. The calcitonin is the antagonist of parathyroid hormone.

1.4. 8.1.4. The parathyroid glandParathyroid gland plays important role in the regulation of the body Ca and P household. It affects the mobilization of calcium ions and the selection of phosphate ions. Hormone production of the gland is influenced by blood calcium level.

• Its hormone is parathyroid hormone (PTH) that is a peptide consisting of 84-amino acids. The sequence of the first 34 amino acids in the peptide is almost identical to the amino acid sequence of calcitonin.

As parathyroid hormone is the antagonist of calcitonin, it increases blood calcium level and effects ossification of bones because it enhances the mobilization of Ca from bones together with phosphate ions.

The increased production of PTH can increase calcification of certain vital organs, as calcium phosphate is deposited due to the increased calcium and phosphate level in blood (e.g. in kidney). Bones become brittle as a result of calcium and phosphorus mobilization.



1.5. 8.1 5. Hormones of the adrenal cortexThe adrenal plays important role in the metabolism and regulation of salt and water balance in the organism. Steroidal hormones of the gland protect cells and tissues against external harmful stimuli. Corticoids are the hormones of adrenal, which are classified based on their physiological effect.

• Hormones influencing salt and water household (mineralocorticoids) (e.g. aldosterone, deoxycorticosterone)

They exert their effect on tubular cells of kidney. They increase the retention of sodium ion, together with chloride ion. They enhance the secretion of potassium and hydrogencarbonate ions. Thus they affect the ratio of Na/K-in urine. The maintenance of the desired osmotic concentrations can be achieved by them.

• Hormones that affect carbohydrate metabolism (glucocorticoids) (e.g. cortisol, corticosterone)

They are the key hormones of gluconeogenesis, they enhance the transformation of amino acids, lactic acid, propionic acid, glycerol and oxaloacetic to glucose and to glycogen. They stimulate protein breakdown and fat mobilization. They increase blood sugar level.

• Hormones that affect sexual behavior (sexual steroids)

These are androgens and estrogens that are responsible for the development of secondary sex characteristics. In addition to it adrenal hormones also affect lymphatic tissues and blood cellular elements and the connective tissue. They have anti-inflammatory effects. Stress increases there quantities.

1.6. 8.1.6. Hormones of adrenal medulla (catecholamines)In general they have a sympathetic effect on the organism. As a result of their effect blood pressure and the heart rate increase and the coronary and muscle veins loosen.

The red blood cells stored in the smooth muscles of the honeycomb get into the bloodstream by means of the contraction of that. Breathing also intensifies.

Blood sugar level rises, as sugar formation is also increased by glyconeogenesis.

These hormones also play a role in transmission of the stimulus of sympathetic nerve fibers. The adrenal medullary function is necessary for the Cannon's "emergency response". They are also responsible for increased wakefulness.

1.7. 8.1.7. Hormones of pancreas Insulin and glucagon are produced by means of the endocrine islets of Langerhans in pacreas.

• Insulin is formed by β-cells, while the glucagon by α-cells.

Insulin consists of two open polypeptide chains (from 51 amino acids) that is bound together by disulfide bridges.

Glucose permeability of cell membranes is increased by insulin, which is reflected in the rise of glucose utilization in peripheral tissues. It enhances glycogen synthesis and storage in liver and muscle. It inhibits gluconeogenesis that is the formation of glucose from amino acids and other intermediate products. It stimulates fatty acid synthesis from glucose in the liver. It also enhances protein synthesis, thus it inhibits its breakdown to amino acids. As a result of these processes it decreases blood glucose level.

• Glucagon is a polypeptide consisting of 29 amino acids, which has an opposite effect on the organism to that of insulin. It causes the rise of blood sugar level as it enhances the conversion of intermediates (e.g. amino acids) into glucose. It enhances protein breakdown, stimulates fat mobilization, glucose formation from fat.

1.8. 8.1.8. Hormones of the ovaryEstrogen and progesterone are the ovarian hormones with steroid skeleton.



• Estrogens are the collective names of hormones that are responsible for the healthy development of female characteristics. These include estradiol, estrone and estriol. Their physiological effect decreases in the listed order. Cholesterol is the starting material for their synthesis. They circulate in blood either bound to protein (2/3) or in free form. They are inactivated in the liver. They are responsible for follicular and ovocitogenesis. They stimulate the cyclic development of uterus and vagina. They are responsible for the development of female sexual characteristics.

• Progesterone (corpus luteum or pregnancy hormone)

Progesterone provides appropriate conditions for inseminated egg in uterus. During pregnancy it inhibits the uterine contractions. It prevents new ovum maturation.

1.9. 8.1.9. The testicular hormones (androgens)These steroidal hormones are formed by Leydig's interstitial cells of testicles. Among them dihydrotestosterone has the greatest, testosterone has lower and rosterone has the smallest physiological effects. They are responsible for the development of male secondary sex characteristics. They affect metabolism, particularly proteins synthesis. Cholesterol is the starting material for the biosynthesis.

2. 8. 2. Tissue hormonesTissue hormones are compounds with some characteristics similar to the hormones produced in endocrine secretion glands, but they are produced by tissue cells with other functions. Among tissue hormones there are some hormones that are secreted by epithelial cells lining the lumen of the stomach and small intestine. The hormones are important in controlling digestive function, they affect on function of stomach and other parts of the digestive tube.

These peptide hormones are called gastrointestinal (GI) hormones.

Secretin is a gastrointestinal hormone and is secreted in the S cells of the duodenum. Secretin is a peptide hormone, which is composed of 27 amino-acids, stimulates the formation and secretion of bile and pancreas. Secretin enhances the secretion of water and bicarbonate production.

Gastrin is produced by G cells of the duodenum and in the pyloric antrum of the stomach. Gastrin is composed by 17 amino acids. It effects on gastric juice secretion, such hydrochloric acid and pepsin production as well.

Cholecystokinin is arised in the duodenum as well. It consists of 33 amino acids and it stimulates the pancreatic and gastric juice production. Cholecystokinin promotes excretion of bile from the gallbladder and it has a positive effect on food intake.

In addition more GI hormones are known (enteroglucagon, motilin, somatostatin, etc), and it is possible to discover new ones. Their effect is largely similar to the ones above.

Angiotensin I is a tissue peptide hormone that affects the vascular system and smooth musculature. This protease decapeptide is produced in the kidney and is activated by renin from angiotensinogen. The Angiotensin II. is formed from the Angiotensin I. by the cleavage of two amino acids. They have regulating effect on the adrenal function and they increase the blood pressure.

Bradikinin consists of 9 amino acids, and have a strong vasodilatory effect. It stimulates the blood flow in the brain, kidneys, coronary arteries and in the skin. Bradikinin lowers blood pressure. It increases the permeability of the capillary wall, so helps the migration of white blood cells. It also influences the smooth muscle contraction.

There are also tissue hormones that they expound their effects on the place of their formation. These are biogen amines. Biogen amines are produced from amino acids.

Histamine is derived by decarboxylation of histidine. It can be found almost everywhere in the organism. It is stored bound to the protein in the cells. It is released by mechanical, thermal, toxic and allergic stimuli. It promotes the secretion of hydrochloric acid. It is a vasodilator, released locally in sites of inflammation or allergic reaction. It increases capillary permeability, decreases the blood pressure.



Serotonin (5-hydroxytryptamine) is derived by hydroxylation and then by decarboxylation of tryptophane. Serotonin is also produced in the enterochromaffin cells in the alimentary canal. It has vasoconstrictor effects, thus increases the blood pressure, increases the heart rate, causes the tracheal, bronchial stenosis. It stimulates intestinal peristalsis. In case of the injury of vein walls serotonin is released, that causes local vasoconstriction. Serotonin is a tissue hormone, but is also a neurotransmitter. It has effect on the central nervous system.

Acetylcholine acts as a neurotransmitter. The synthesis and the breakdown of acetylcholine are rapid enzymatical processes. These reactions are catalysed by acetylcholine esterase (mainly in the brain and nerve tissue) and cholinesterase (serum). The cholinesterase also catalyzes the hydrolysis of propionyl choline and butyryl choline.

Gamma-aminobutyric acid (GABA) is derived from the decarboxylation of the glutamic acid. It can be found in the brain. GABA is the major inhibitory neurotransmitter. It inhibits the ganglionic transmission.

3. 8. 3. Plant growth hormones (Phytohormones)In higher plants, like in animal body, the connection between cells is achieved by chemical messengers. The plant hormones are chemical mediators. They interact with specific proteins, or receptors. They can expound their effect away from the place of their formation. They occur in low concentration in plants. Mostly, they are effective together with other phytohormones. Their effects highly depend on their concentrations. Typically, plant growth hormones have low molecular weight, they are not peptide hormones.

They can not be grouped clearly by their physiological function so their classification is based on their chemical structure.

Auxins are indole skeleton hormones. Most important members of the auxin family are indole acetic acid (IAA) and phenylacetic acid. They have effects on plant growth and development. The precursors for the synthesis of auxin are the aromatic amino acids (tryptophan, phenilalanine). The auxin synthesis takes place in young leaves and developing crops. The degradation of auxin is catalyzed by auxin-oxidases. The process is enhanced by the light and ethylene. The auxins promote the effects of gibberellins and cytokinins on cells, and cooperate with ethylene in the regulation of plant growth.

8.3. ábra - indole-acetic acid

Gibberellins are derived from the ent-gibberellane skeleton. In higher plants their synthesis is intensive in



young, growing parts of the plant (in the youngest shoots). Their synthesis corresponds to the synthesis of terpenoids. The speed of the reaction sequence is affected by the length of the days. In long days gibberellins, while in short days abscisic acid is formed. Gibberellins promote flowering also.

8.4. ábra - gibberellic acid

Cytokinins are adenine derivatives containing isopentenyl side chain. The cytokinins stimulate cell division in the presence of auxin, enhance the nutrient transport. They can be found in every plant and they are needed in all developmental stages of plants. They help in maintaining the young status of plant.

8.5. ábra - zeatin

Ethylene stimulates the ripening of fruits and at the same time inhibits the growth, inhibits the auxin transport.

Abscisic acid is a terpenoid, and is an obstructive substance, which hurries the ageing, the defoliation. It plays a role in the fall of leaves in autumn and in ensuring the winter dormancy.


9. fejezet - 9. BIOACTIVE COMPOUNDS III. ENZYMES (BIOCATALIZATORS) The enzymes are proteins that direct and accelerate the biochemical processes in the living organisms by lowering the activation energy.

A + B → C + D, pl.: 2 H2O2→ 2H2O+ O2

The activation energy of this process without catalyst is 75 kJ/mol, and with inorganic catalizator (Pt) is 49 kJ/mol, while with catalase enzyme is 23 kJ/mol (Fig. 21).

9.1. ábra - Figure 21: Enzymes and activation energy

The data also show that the activation energy is decreased to a greater extent by the influence of an enzyme than by the effect of inorganic catalysts. The enzymes thus able to accelerate reaction rate dramatically.

1. 9. 1. Structure of the enzymesEnzymes are proteins considered simple or complex proteins. In many cases, enzymes consist of a protein part called apoenzyme and a combination of one or more parts (which may be a non-protein substance) called active group. The apoenzyme carries the active group, which catalyzes the reaction. The two parts together, the enzyme complex, is referred to as holoenzyme. An enzyme can perform its biological function only as a holoenzyme.

Many enzymes are only proteins. These enzymes are known as simple enzymes (e g hydrolases). The active site of an enzyme is created by functional groups of amino acid side chains. These groups have to be in proper spatial position relative to each other, and they are stabilized by bonds. If the spatial position changes because of alteration of the external circumstances, the enzyme loses its biological activity.

The active group (is bound to the protein part of an enzyme) is called coenzyme or prosthetic group.

The coenzyme is a non-protein active group, which is loosely attached to the apoenzyme. This group can be detached by dialysis from the protein part of the enzyme. During the reaction the coenzyme can be released from an apoenzyme and can get onto the other (NAD, ATP, UTP, CTP, CoA-SH).

Prostetic group is tightly bound (with primary or secondary bonds) to the apoenzyme. This group can not be


9. BIOACTIVE COMPOUNDS III. ENZYMES

(BIOCATALIZATORS) detached by dialysis. The prosthetic group is bound to the same apoenzyme in the process of catalysis (FAD, Hem, vitamin H, vitamin B6, metal ions).

Multi-enzymes consist of a number of enzyme proteins (with different catalytic activities), which are held together by secondary bonds. Many reactions take place with their help, where the specific sequence of reactions is determined. The substrate is transferred from one of the reaction site to another one.

2. 9. 2. The function mechanism of the enzymesThe reactions catalysed by the enzymes are two-step processes. First, the enzyme (E) reacts with the substrate (S) and a transient enzyme-substrate complex is formed (ES) at the active site of the enzyme. In the second step there is a transformation process in the enzyme-substrate complex and the enzyme-product complex (EP) is formed. Finally, the product is detached from the enzyme (Fig. 22)

In the catalytic processes, the task of apoenzymes is to keep the active group. Apoenzymes should provide the appropriate spatial arrangement for running the reactions.

The catalytic process takes place in the active centre of the active group. The active centre contains the binding site, where the substrate is bound and the catalytic site, where such groups may be found which react with the substrate. It may even occur that the binding site is the same as the catalytic site.

The name of an enzyme can be derived from its substrate with the word ending in –ase (e.g. lactose-lactase), or from the chemical reaction it catalyzes, also with the word ending in –ase (e.g. dehydrogenase). Enzymes often have trivial names (for example: pepsin, papain), too.

The accurate, clear name of an enzyme has to refer to the substrate and the type of catalytic process as well.

For example: the substrate of glucose-1,6-transphosphorylase enzyme is the glucose-1 phosphate. This enzyme cleaves the phosphate group from the first carbon atom of glucose-1 phosphate and transports it to the 6th carbon atom.

3. 9. 3. The specificity of the enzymesMost enzymes can catalyze only one reaction of one substrate. This feature is called specificity. There are two specificities known of enzymes, the substrate specificity and the reaction specificity.

The substrate specificity means that at the binding site of the enzyme only its substrate can bind. For example: the lactic acid can only bind to the binding site of the lactic acid dehydrogenase enzyme. The apoenzyme is responsible for this substrate specificity.

The reaction specificity means that the enzyme catalyses one chemical reaction (Fig. 22).

9.2. ábra - Figure 22: The specificity of the enzymes



(BIOCATALIZATORS)

4. 9. 4. Classification of enzymesEnzymes are principally classified according to the reaction they catalyse. Enzymes are generally classified into six main family classes:

1. Oxidoreductases

2. Transferases

3. Hydrolases

4. Liases (synthases)

5. Isomerases

6. Ligases (synthetases)

More sub-groups can often be distinguished within the main groups.

4.1. 9.4.1. OxidoreductasesEnzymes in this class catalyse oxidoreduction reactions, catalyse such a biochemical processes, where oxygen uptake, proton and electron transfer happens. The systematic name of this class is based on donor: acceptor oxidoreductase.

Their coenzymes are generally: NAD+, NADP+, FAD, Liponic acid, Coenzim Q.

The oxidoreductases can be divided into two sub-groups: dehydrogenases or oxidases. One group includes the dehydrogenases, which oxidize the substrate by transferring hydrogen from the substrate (electrons and protons) to an acceptor. The substrate is oxidized and the acceptor is reduced.

AH2 + B →A + BH2

Oxidases are enzymes involved when O2 acts as an acceptor of hydrogen or electrons. The donor is the substrate. In this process, the substrate is oxidized, while reactive hydrogen peroxide is formed from the oxygen.

AH2 + O2 → A + H2O2



(BIOCATALIZATORS) Hydrogen peroxide is an easily degradable compound; it is regarded as a cytotoxic agent. During its decomposition free radicals are formed, which attack the unsaturated bonds of lipid components of the membrane, they have destroying effects.

In the defence mechanisms of living organisms, catalase and peroxidase enzymes catalyse the decomposition of hydrogen peroxide.

Catalase enzyme catalyses the breakdown of hydrogen peroxide into water and oxygen.

2H2O2 → 2H2O + O2

Peroxidase also catalyzes the hydrogen peroxide decomposition. Peroxidase cleaves electrons and protons from another compound, while this compound is oxidized. From the hydrogen peroxide two water molecules are formed.

AH2 + H2O2 → A + 2H2O

4.2. 9.4.2. TransferasesTransferases catalyse the transfer of a functional group from the substrate (called the donor) to another compound (called the acceptor).

AR + B → A + BR R: chemical group

In the coenzyme part of transferases, water-soluble vitamins can often be found as we have discussed in a previous section (Table 5).

9.3. ábra - Table 5: Coenzymes of transferases and transmitted chemical groups

4.3. 9.4.3. HydrolasesHydrolases are simple proteins that catalyze biochemical processes, where – with the incorporation of water – decomposition occurs. Although these processes are reversible, they are strongly shifted towards degradation. For example, the macromolecules in the organism are decomposed to their monomer units by hydrolysis.

AB + H2O → AH +BOH

Within the hydrolases more sub-groups can be distinguished.

Esterases (cleave ester ulinkages) include lipases and ribonucleases and phosphatases.

Glicosidases (catalyse the hydrolysis of the glycosidic ulinkage) include amylase, lactase, sacharase, nucleosidase.

Proteases split peptide bonds of the polypeptide chains (carboxy, amino and dipeptidases). Proteases are pepsin, trypsin, kimotrypsin and papain.



(BIOCATALIZATORS)

4.4. 9.4.4. Lyases (Synthases)Lyases catalyse chemical reactions without taking up or losing water, where cleaving of carbon-carbon, carbon-oxygen and carbon-nitrogen bonds will appear, often forming a new double bound, a new ring structure. In the coenzyme of lyases, there are (in general) vitamin B6- and B1. For example: the pyruvic acid decarboxylase enzyme (as a lyase), is a central enzyme of the glucose anabolism.

4.5. 9.4.5. IsomerasesThe isomerases catalyze reactions in which the configuration of one chiral carbon atom of the substrate is changed and thus a new product is formed (epimerisation). In cis-trans isomerization processes enzymes modify the spacing of molecules along the double bonds of a substrate. Enzymes catalyzing aldose-ketose interconversion are also in this group. These enzymes modify the location of oxogroup and thus another compound is created. For example, hexose-phosphate isomerase catalyzes the conversion of glucose-6-phosphate to fructose-6-phosphate. Coenzymes of isomerases are UDP, vitamin B12.

4.6. 9.4.6. Ligases (synthetases)The ligases catalyse the joining of two large molecules by forming a new chemical bond. The development of the new compound needs energy. The energy required for the process is supplied by high-energy-ulinked nucleotides (ATP, UTP, GTP).

The acetyl-CoA synthetase is also a member of the ligase family and catalyses the following process:

CH3-COOH + CoA-SH + ATP = CH3 - CO~SCoA + AMP + PPi

The ligases take part in the synthesis of nucleic acids and proteins.

5. 9. 5. Factors influencing the function of enzymesThe factors influencing the function and activity of enzymes also modify the velocity of reactions.

The enzyme activity is influenced by:

• enzyme concentration,

• substrate concentration,

• pH,

• temperature,

• activators,

• inhibitors.

Initially with the increasing of enzyme concentration, the velocity of reaction enhances, until all enzymes can be in enzyme-substrate complex state.

With decreasing substrate concentration and with increasing product concentration the velocity of the reaction begins to decrease. In this case, the substrate is present only in small concentration, so can not tie up the total amount of enzyme. The decrease of reaction might be often caused by enriched products (final inhibition).

With increasing substrate concentration the velocity of reaction increases up to the saturation value, but at the depletion of free enzyme molecules with increasing substrate concentration the velocity of the reaction will not increase.

The excess of substrate often causes inhibition, because when too many substrate molecules are present, it does not only attach to the binding site of the enzyme, but also modifies its configuration leading to inactivation.

With increasing temperature the velocity of reaction enhances up to a maximum then begins to decrease.



(BIOCATALIZATORS) The increase of temperature, as any chemical reaction including the enzyme-catalysed processes, enhances the rate of reaction, because the energy of the reactant molecules increases. The temperature of maximum velocity is called the optimum temperature of the enzyme. When the temperature is higher, the configuration of enzyme changes and it will not catalyse the reaction any longer.

Above a given temperature, the conformation of the protein part is changed to such an extent, that the enzyme loses its biological activity.

Enzymes have optimum pH. This is the pH range where enzymes can perform their biological tasks. Any change in pH above or below the optimum will cause a change of conformation of protein part of the enzyme, which influences its function negatively.

The redoxpotential of the medium also influences the functions of enzymes. Oxidoreductases are particularly sensitive to this parameter.

The activators increase the catalysing ability of enzymes. The activators may be cations (K+, Na+, NH4+, Mg++, Mn++, Zn++). Single charged cations expound their positive effect with pulling up the hydrate layer of the enzyme, while double charged cations expound their positive effect with helping to bind either the substrate or the coenzyme.

The inhibitors decrease the enzyme activity. The inhibition may be reversible or irreversible.

Reversible inhibitors bind to the enzymes with non-covalent interactions, so after their removal the enzyme may become active. The reversible inhibitions might be competitive, non-competitive, uncompetitive and allosteric inhibitions.

• Competitive (competing) inhibition means: The inhibitor is similar to the substrate or coenzymes chemically, so either the substrate or the inhibitor competes for access to the enzyme's active site, or the inhibitor binds to the apoenzyme instead of the coenzyme. In both cases the catalytic process is blocked.

• Non-competitive inhibition means: The inhibitor does not affect the binding of substrate, but prevents the conversion of bound substrate. This happens in such a way that the inhibitor is attached to the catalytic site of the enzyme's active centre.

• Uncompetitive inhibition means: The inhibitor cannot bound to a free enzyme, but is able to bind to the enzyme-substrate complex only.

• Allosteric inhibition means: The inhibitor does not bind to an active site of the enzyme, but binds to other parts of the protein. The inhibitor binds to an allosteric site and so a change in the shape of the active site will occur. The activity of the enzyme will decrease. The substrate can not bind to the active site due to the fact that the active site has changed shape and the substrate no longer fits. The accumulated end-product, in a similar manner inhibits the activity of the enzyme.

At the reversible inhibition the denaturation of protein part of an enzyme is irreversible. The enzyme loses its activity. Then the inhibitor is called destructor.


10. fejezet - 10. THE METABOLIC PROCESSES I. CARBOHYDRATE METABOLISMLiving organisms are in dynamic relationship with their environment to maintain their life processes. They take up nutrients and energy from their surroundings – from the lifeless nature, they transform it in their bodies and empty the decomposition products that are useless to them, into their environment.

Material conversion processes taking place in living organisms are called intermediate metabolism. Intermediate metabolic processes consist of constructing (anabolism) and decomposing (catabolism) processes. During metabolism there are substance, energy and information flows. Living organisms acquire material and energy for their constructing processes from different sources. It can be an aspect for the classification of living organisms. There is close relationship between organisms and their metabolism. The autotrophic organisms produce their own organic material from simple materials (precursors: CO2, H2O) taken up from their environment by means of sunlight. The by-product of this process, oxygen is discharged into the environment. The heterotrophic organisms construct their own materials from (organic) products of autotrophic organisms and oxygen released by them into the environment.

The heterotrophic organisms first break down materials taken up from their environment, then during their constructing processes they produce their own new materials from one part of the products (precursors) and energy. They empty unnecessary materials formed during the metabolism. These materials can be precursors for autotrophic organisms.Figure 23 show the turnover of the four main constituent elements (C, H, O and N) in the biosphere.

10.1. ábra - Figure 23: The carbon, hydrogen and oxygen biological cycle

The carbon, hydrogen and oxygen biological cycle are self-sufficient considering the whole biosphere. Nitrogen occurs in large quantities in the atmosphere, but the majority of the organisms can not utilize it in elemental form. Only special bacterium species can build (fix) nitrogen from air in their bodies. From their dead organic material after transformation processes in the soil (aminization, ammonification, nitrification) those forms of nitrogen arise, which can be utilized by plants. Nitrogen entering the food chain in this way becomes part of the biological cycle. Nitrogen can be found in precursors of constructing processes of higher ranking animals. Animals excrete nitrogen during their metabolic processes. From the decomposition of their dead organic matter such inorganic nitrogen forms are formed in the soil, which is used by plants in their constructing processes. Those processes also take place in soil (denitrification), where elemental nitrogen is formed (nitrogen loss). This nitrogen is released in the atmosphere and it can return to the biological cycle only through nitrogen fixing bacteria (Fig. 24).

10.2. ábra - Figure 24: The nitrogen biological cycle


10. THE METABOLIC PROCESSES I.

CARBOHYDRATE METABOLISM

1. 10. 1. Carbohydrate biosynthesis in photosynthetic organismsCarbohydrate synthesis of the photosynthetic organisms, is known as photosynthesis. Here carbohydrates are formed from inorganic precursors by means of the energy of sun. Photosynthesis takes place in the chloroplasts of photoautotrophic organisms. The gross equation of photosynthesis is the following:

6 CO2+ 6 H2O → (CH2O)6 (monosaccharide) + 6 O2

Photosynthesis can be divided into two phases (light dependent and independent phases). These are characterized by different types of processes, and they take place in different places in the chloroplasts.

The light dependent phase takes place in the inner membrane system of the chloroplast. (thylakoid membrane). In the light reactions photons are absorbed and electrons are transported. As a result of the processes ATP, NADPH + H+ and O2 as by-products are formed at this stage. Light independent reactions take place in the intermembrane space of chloroplasts (stroma), where using the by-products of light dependent phase CO 2

fixation and carbohydrate synthesis occur.

1.1. 10.1.1. The light dependent phase (Hill reaction)Light energy is absorbed by photoreceptors. These are chromoproteins consisting of protein components and pigments.

Pigments such as chlorophyll -A and –B with porfirin frame, and carotenoids with isoprene frame are conjugated double bond systems. Due to there structures the conjugated double bond systems can be easily excited. During excitation their electrons enter into higher energy levels but they are still attracted by the atomic core. The excited state exists for a short period, after which the electrons jump back to the ground level. The difference of the energy level between the initial and the excited state is emitted or transferred to other systems.

Two kinds of pigment systems are involved in binding and concentrating light energy which are associated with electron transfer and are called photochemical system I and II.

Pigment systems contain light-gathering pigments and reaction centers.

The center of photochemical system I is called P700 and that of system II is called P680. The light absorption optimum of these centers are in 700 and 680 nm wavelength range. The difference is resulted from the quality of the pigments. While in photochemical system I carotene and chlorophyll-A dominate, in the system II xanthophyll and chlorophyll-B are characteristics.

The two photochemical systems are connected by electron transport chain. In the photochemical systems, electrons are transported from the more negative redox potential component towards the more positive one, except for two points in photosynthetic electron transport.




(I.: +0,4 → - 0,6V; II.: + 0,8 →0,0V).

In the process an electron is torn from photochemical system I by the absorbed and concentrated light energy and it attaches to a primary electron acceptor (FRA) and from there to ferredoxin. From ferredoxin the electron binds onto the oxidized NADP+ and reduces it.

The lack of electrons in photochemical system I is replaced by the electrons that leave photochemical system II and enter the electron transport chain, thus it will be able to absorb new light quantums. Then photochemical system II becomes electron-deficient, which is replaced by electrons derived from water hydrolysis. This form of electron transport is called non-cyclic electron transport.

Thus at non-cyclic electron transport both photochemical systems work. In photochemical system I NADPH+ + H+ is generated, in photochemical system II. water is split and oxygen gas is released. In the electron transport chain, during photophosphorylation ATP is formed. At cyclic electron transport one photochemical system is only in action. The electron from ferredoxin attaches not to oxidized NADP+, but to a member of the electron transport chain that is cytochrome b6. From here it binds to the cytochrome f, and then through plastocianin it returns to photochemical system I. The end product of cyclic electron transport is ATP. The cyclic path occurs when there is lack of oxidized NADP+ and there is excess ADP (Fig. 25).

10.3. ábra - Figure 25: Hill reaction

Mitchell theory of photophosphorylation

One part of the energy of electrons involved in transport process (between the two photochemical systems) is utilized for proton transfer. Plastoquinone has a central role in the proton transfer.

As a result of proton transfer their concentration increases on the inner surface of thylakoid membranes. Charge and voltage difference result proton transport from the inner space to outside through proton channels of the membrane. Proton channels are in connection with the ATPase enzyme. During proton transfer their energy is applied to form ATP by means of the ATPase enzyme. Light energy is converted to chemical energy.

1.2. 10.1.2 The light independent phase of photosynthesis (Calvin cycle)In the process carbon dioxide is transformed to carbohydrate from the end products of the light dependent phase. The cycle consists of three phases: these are carboxylation, reduction and regeneration phases. At carboxylation phase carbon dioxide is bound by ribulose 1,5-bisphosphate, from which two mol glyceric acid-3-phosphate is formed by the decomposition of the intermediate product. At the reductive stage first glyceric acid-1,3-diphosphate is formed using ATP, that was generated in the light dependent phase, then glyceraldehyde-3-phosphate is formed by means of NADPH + H+. This product is in equilibrium with dihydroxyacetone phosphate. Trioses combine and form fructose-6-phosphate from which glucose-6- phosphate is built by




isomerization.

At the regeneration phase ribulose 1,5-bisphosphate is formed, which is the starting compound of the cycle. During transaldolase, transketolase reactions, epimerization and isomerization processes ribulose 1,5-bisphosphate is reproduced by one part of the fructose-6-phosphate, glyceraldehyde-3-phosphate and dihydroxyacetone phosphate (Fig. 26).

The equation of the cycle

(6 ribulose 1, 5- diphosphate) + 6CO2+ 18 ATP + 12 NADPH+H+ →

(6 ribulose 1, 5- diphosphate) + 1 hexose + 18 ADP + 18Pi + 12 NADP+

10.4. ábra - Figure 26: Calvin cycle

1.3. 10.1.3. The sucrose synthesisGlucose is the end product of photosynthetic carbohydrate synthesis, but it does not accumulate in plants, as it is transformed to sucrose or different polysaccharides. UTP provides energy for sucrose synthesis and ATP for starch synthesis. The starting materials are sugar phosphates. During sucrose synthesis UTP ulinks to glucose-1-phosphate with a split of inorganic pyrophosphate. The newly created, intermediate product with high energy is uridine diphosphate-glucose that is capable of reacting with the fructose-6-phosphate. The process is catalyzed by sucrose phosphate synthase. After cleaving inorganic phosphate from sucrose phosphate by phosphatase enzyme sucrose is formed, while another product of the process is regenerated by UDP and ATP.

The reactions are the following:

glucose-1- phosphate + UTP → uridine-biphosphate-glucose +PPi;

uridine-biphosphate-glucose + fructose-6-phosphate ↔ sucrose-phosphate + UDP;

sucrose-phosphate → sucrose + Pi;

UDP + ATP→ UTP + ADP.

1.4. 10.1.4. The starch synthesisDuring starch synthesis ATP attaches to glucose-1-phosphate that is of the starting compound. Adenosine diphosphate-glucose that is formed by the cleavage of an inorganic pyrophosphate group ulinks to the acceptor by transglycolase enzyme.

The process of the reaction is the following: glucose-1-phosphate → ATP + adenosine diphosphate-glucose + PPi; adenosine diphosphate-glucose + acceptor → α-1 ,4-glycosyl acceptor + ADP.




2. 10. 2. Catabolic processes of carbohydratesHeterotrophic organisms use energy released during their catabolic processes for the constructing processes of their cells. Intermediate products are starting materials (precursors) of anabolic processes.

Among catabolic processes the greatest quantity of energy releases when nutrient molecules burn and form inorganic substances (CO2, H2O, NH3). The reaction sequence is called respiration (cellular respiration). Organisms use atmospheric oxygen from their environment and release carbon dioxide.

Catabolic processes begin with the hydrolysis of the macromolecules. During hydrolysis, they break into their monomeric units. The monomers decompose in individual reaction pathways forming the same intermediates (pyruvic acid, acetyl-CoA). Intermediates transform through common reactions to simple inorganic substances, and most of them are excreted by the body. The hydrolysis of macromolecules is catalyzed by hydrolases. The decomposition of fats and oils are catalyzed by lipases, that of proteins by proteases and that of nucleic acids by the nucleases. The hydrolysis of starch is enhanced by amylases, etc. The breakdown of starch and glycogen can also take place through hydrolysis with phosphoric acid. The process is catalyzed by phosphorylase enzyme. The end product of the reaction is glucose-1-phosphate (Fig. 27).

10.5. ábra - Figure 27: Catabolic processes of carbohydrates

In the following, the stages and steps of carbohydrate decomposition are described in detail, as the breakdown of other organic molecules also ends in this reaction sequence although it starts in different routes.

2.1. 10.2.1. Cellular respiration2.1.1. 10.2.1.1. Glycolysis

The first stage of glucose breakdown is glycolysis. The process takes place in the cytoplasm. The process does not require oxygen. At the end of the reaction sequence 2 M pyruvic acid is formed by 1M glucose. Pyruvic acid is decarboxylated at the second stage of the process. This process takes place already in the mitochondria. At the end of the reaction acetyl-CoA is formed by pyruvic acid.

The acetyl group with two carbon atoms enters into the third stage of the process from acetyl-CoA, which is called citric acid cycle. This takes place in the mitochondrial inner membrane within the matrix. In the process carbon dioxide molecules and reduced coenzymes (NADH + H+, NADPH + H+, FADH2) are formed from the acetyl group.

The last stage of complete breakdown is terminal oxidation that takes place in the mitochondrial inner membrane.




In this process, a large amount of water and energy are generated from reduced coenzymes and inhaled oxygen that is formed in the former stages of catabolic processes.

Figure 28 shows the reaction sequence of glycolysis that is the first stage of cellular respiration, Starting from glucose, two moles of ATP are used during the formation of fructose-1,6-diphosphate. The fructose-1,6-diphosphate breaks into sugar molecule with two or three carbon atoms in aldolase reaction. The glyceraldehyde-3-phosphate and dihydroxyacetone phosphate can be converted into each other. The subsequent step of the process continues with glyceraldehyde-3-phosphate. The decrease of its concentration enhances the transformation of dihydroxyacetone phosphate.

Glyceraldehyde-3-phosphate transforms to glyceric-1,3-diphosphate by incorporation of inorganic phosphate and oxidation with the formation of reduced coenzyme (NADH + H+). This molecule stores its energy in ATP and transforms to glycerin-3-phosphate. The next step in the reaction sequence is an isomerization process in which glyceric-2-phosphate is produced. From this pyruvic acid is formed besides ATP molecules, which is the end product of anaerobic glycolysis. In the process 2 moles of intermediates are formed from one mole of glucose from glyceraldehyde-3-phosphate.

10.6. ábra - Figure 28: Glycolysis




2.1.2. 10.2.1.2. Pyruvate decarboxylation

Pyruvate decarboxylation can be considered as the second stage of glucose combustion, which is closely related to the third section, the citric acid cycle.

In this process pyruvic acid formed in the cytoplasm translocates to the mitochondria. The pyruvate is transformed into acetyl-CoA by a process called oxidative decarboxylation. This process is catalysed by a multi-enzyme complex that is called pyruvate dehydrogenase complex (Fig. 29).

10.7. ábra - Figure 29: Pyruvate decarboxylation

The reaction is irreversible. Energy released during the oxidation reaction can be found in the ulinkage of acetyl-CoA and in the reduced cofactor (NADH + H+). Enzyme complex consists of three different enzymes and five cofactors.

These are:

• pyruvic acid decarboxylase enzyme with TPP (vitamin B1) coenzyme

• dihidrolipoil transacetylase enzyme with lipoic acid coenzyme

• the dihidrolipoil dehydrogenase enzyme with the FAD coemzyme

• CoA-SH,

• and NAD+.

Most of the produced acetyl-CoA molecules are utilized in citric acid cycle and the rest in fatty acid synthesis as a precursor.

2.1.3. 10.2.1.3. Citric acid cycle

Citric acid cycle is the third stage of glucose breakdown. The process takes place in the mitochondria. Enzymes catalyzing this reaction sequence can only be found in the mitochondria. In this cycle, the intermediate metabolic products are burned completely.

The acetyl group of acetyl-CoA joins the oxaloacetic citric acid and gives citric acid with water and CoA-SH loss. The process is catalyzed by the enzyme citrate synthase. Citric acid is transformed to isocitric acid by the enzyme aconitase. The intermediate is cis aconitic acid. In the next step oxalic succinic acid is formed catalyzed by dehydrogenase, which transforms to α keto glutaric acid through decarboxylation. CO2 and NADH + H+ leave the cycle.

The α-ketoglutaric acid transforms to succinyl-CoA in the next step. Then, through decarboxylation CO2 leaves the process and NADH + H+ are generated. Then Succinyl-CoA transforms to succinic acid, while the energy is stored in GTP. From succinic acid fumaric acid and FADH2 are formed by means of succinate dehydrogenase enzyme.

Fumaric acid is transformed to L-fumaric acid by fumarase enzyme, then in the final step of the cycle oxaloacetic acid and NADH + H+ are formed by means of malate dehydrogenase (Fig. 30).




10.8. ábra - Figure 30: Citric acid cycle and terminal oxidation

2.1.4. 10.2.1.4. The terminal oxidation and oxidative phosphorylation

This process is the fourth and final phase of glucose combustion. It takes place in the inner membrane system of mitochondria.

In the previous catabolism sections protons and electrons are formed through the oxidation of reduced coenzymes. They pass along redox enzyme complex systems in the mitochondrial inner membrane.

In the last step they react with the inhaled oxygen and form water. Proton transport results increased proton concentration at the external part of the membrane (Fig. 30).

The equalization of the charge difference occurs across proton channels (from outside to inside), the released energy transforms to ATP (oxidative phosphorylation).

Figure 30 also shows that while electrons and protons from NADP-and NAD- get to iron-sulfur proteins, from the reduced FAD electrons get directly to ubiquinone. During oxidative phosphorylation process from 1 mole of NAD and NADP 3 moles of ATP and from 1 mole reduced FAD 2 ATP molecules are generated.

The energy balance of the total breakdown of one mole glucose is the following.

Glycolysis: glucose → 2 pyruvate + 2 ATP + 2 NADH+H+ = 2 pyruvate + 8 ATP

Decarboxilation of pyruvate:

2 pyruvate → 2 acetyl-CoA +2 CO2 + 2 NADH+H+ = 2 acetyl-CoA + 6 ATP

Citric acid cycle:




2 acetyl-CoA → 4 CO2 + 4 NADH+H+ + 2 NADPH+H+ + 2 FADH2 + 2 GTP = 24 ATP

Total: 38 ATP

During the breakdown of one mole glucose (CO2 and H2O) 38 ATP molecule

2.2. 10.2.2. The pentose phosphate pathwayAn alternative pathway for glucose oxidation is the pentose phosphate pathway. The cycle enzymes can be found in a variety of animals, plants and microbes so these reactions are common in the living world.

The penthose phosphate pathway operates to varying extents in different cells. Due to the high physical load, with the aging of cells the breakdown of glucose may be shifted to this pathway.

The significance of the penthose phosphate pathway is that in addition to producing energy, also produces NADPH + H+, which is formed only in a few reactions, but is required to other processes such as the synthesis of fatty acid.

The major part of intermediate products of this process can be starting compounds of other processes.

The ribulose-5-phosphate is an important compound of the dark reaction of photosynthesis. The ribose-5-phosphate is a precursor for the synthesis of nucleic acids. Intermediates of the transaldolase - and transketolase reactions are involved in the synthesis of aromatic compounds through the shikimic acid pathway.

The pathway operates in the cytoplasm. The initial reaction is catalyzed by glucose-6-phosphate dehydrogenase. The specific electron acceptor of this enzyme is coenzyme NADP. In one turn of the pentose-phosphate cycle one mol CO2 is removed, thus six repeated turns result the complete oxidation of glucose-6-phosphate to CO 2

and water. In six turns of the cycle besides the 6 moles CO2, 12 moles NADPH+H+ are also generated, which can be further oxidized in terminal oxidation and this generates 36 mol of ATP.

1 mol glucose (C6H12O6) → 6CO2 + 12 NADPH+H+→ 36 ATP

10.9. ábra - Figure 31: The pentose phosphate pathway




2.3. 10.2.3. Fermentation processesThe role of fermentation is mainly important in the metabolism of microorganisms. Fermentation may also occur in higher organisms. For example, the breakdown process of glycogen may occur in the muscle due to an intense exercise where oxygen supply becomes limited. By the fermentation processes, the living cell obtains energy through the breakdown of carbohydrates to simple organic acids without requiring oxygen and by using hydrogen transfer coenzymes.

The end products are energy-rich compounds therefore little energy is released in this process. The end products might be the precursors of biosynthesis of other organic matter.

This process is called anaerobic dissimilation, because its products do not enter the citric acid cycle, but in additional anaerobic reactions, they are transformed into other compounds.

Figure 32 shows different fermentation pathways. The figure also shows that the pyruvic acid produced during anaerobic glycolysis can be converted further in a variety of ways, so it has central role in the fermentation processes as the starting compound.

10.10. ábra - Figure 32: Fermentation processes




The pyruvic acid can be decarboxylated to acetyl-CoA, which can be further converted to acetic acid or butyric acid. The pyruvic acid can be converted to acetaldehyde while CO2 is released. The acetaldehyde can be reduced to ethanol.

The pyruvic acid can also be transformed to lactic acid and propionic acid in the fermentation processes.

Bacteria are able to split or hydrolyse cellulose, hemicellulose and pectin too, therefore not only the starch can be the starting compound of the fermentation processes in the rumen of ruminants or in silos.

Many of the fermentation pathways are named for their end products. For example there can be acetic acid, butyric acid, lactic acid, propionic acid and mixed acid fermentation processes.

In alcoholic fermentation process, ethanol is formed from the pyruvic acid by reduction and after leaving carbon dioxide.

In lactic acid fermentation, process lactic acid is formed from the pyruvic acid (Fig. 33).

10.11. ábra - Figure 33: Alcoholic- and lactic acid fermentation

There are two pathways of propionic fermentation.

1. pathway: lactic acid → lactyl-CoA → acryloyl-CoA → propionyl-CoA → propionic acid.

2. pathway: oxalic acetic acid → malic acid → fumaric acid → succinic acid → succinyl-CoA → propionyl-CoA → propionic acid

The precursor of acetic acid fermentation is the acetyl-CoA. In this process, acetic acid is produced from acetyl-CoA.

The butyric fermentation takes place in the following pathway: acetyl-CoA → acetoacetyl-CoA → ß-hydroxy-butyryl-CoA → crotonyl-CoA → butyric acid.




In terms of ruminants the most important volatile acids are acetic acid, propionic acid and butyric acid. These organic acids are formed in the rumen or in the silo by fermentation processes. The volatile acids contain the most part of the energy of fodder, therefore plays a decisive contribution to the ruminant material and energy consumption.

During the fermentation process, the terminal oxidation is stopped and thus the reduced coenzymes can only transfer their electrons and protons to intermediates. In the process, the aforementioned volatile acids or in a bad case hydrogen and methane are formed. The energy content of these two gases is lost for the animal.

2.3.1. 10.2.3.1. The fermentation processes in the rumen of ruminants

Rumen microbes ferment dietary carbohydrates to organic acids to obtain energy for their anabolic processes. In rumen generally mixed acid fermentation takes place. Ruminants are able to cover their high percentage (60-70%) of energy demands from acetic, propionic and butyric acids generated in the rumen during fermentation. Besides volatile fatty acids methane and hydrogen gases are also generated during fermentation, which gases leave with rumen and intestinal gases. The quality of fodder influences the rate of end products of fermentation. During feeding acetic acid fermentation is promoted.

The ratio of volatile fatty acids, acetic acid:propionic acid:butyric acid are 6.5:2:1. The pH is close to neutral (pH 6-7).

During feeding the volatile fatty acid content increases, acetic acid content decreases, while propionic acid content increases. Due to the decrease of pH in the rumen the conditions are favourable for the synthesis of butyric acid. Butyric acid has unfavourable influence onto the inner part of the first stomach (epithelium). The absorption of fatty acids from the first stomach decreases, so the pH also decreases. If the pH reaches the 5 to 5.5 values, the lactic acid bacteria multiply and lactic acid fermentation is promoted. When the rumen pH becomes more acidic, acidosis may occur. The production of volatile fatty acids is reduced so that the energy supply of animals becomes insufficient, which can ultimately lead to the death of animals.

One part of volatile fatty acids is neutralized by the saliva (HCO 3-), while the higher part is absorbed across the

ruminal epithelium.

The volatile fatty acids passing through the epithelium can get directly or can get after transformation to the bloodstream. Velocity of absorption of volatile fatty acids is determined by their polarity and pH of the medium. If the pH is low, the dissociation of volatile fatty acids decreases, thus the molecule become apolaric, which helps their absorption.

15-20% of the acetic acid is able to be absorbed at pH=6; 30–35 % at pH=5; while 65–70 % at pH=4. Among volatile fatty acids butyric acid has the highest absorptive capacity at the same pH followed by the values of propionic and acetic acids. The fate of absorbed (and got to the blood) volatile fatty acids is different in the organism.

One of the products of mixed acid fermentation in the rumen is acetic acid, so it is understandable that the acetic acid concentration of ruminants’ blood is much higher than that of other animals. The absorbed acetic acid is mainly utilized by muscle and fat cells.

Acetic acid found in the muscle cells is converted to acetyl-CoA in mitochondria and is oxidized to CO 2 and water in the citric acid cycle and in the terminal oxidation while energy (ATP) is generated.

Acetic acid found in the fat cells is converted to acetyl-CoA in the cytoplasm which compound is a precursor for fatty acid synthesis.

Nearly 65% of propionic acid formed during fermentation can get directly into bloodstream. Propionic acid is removed from blood by the liver, where it is formed to propionyl-CoA and after carboxylation transformed to succinyl-CoA. This compound entering the citric acid cycle either is devoted to energy generation, or is formed to glucose from the intermediate of the cycle (from oxaloacetic acid) by the process of gluconeogenesis. Nearly half of the milk sugar is generated from propionic acid in this way.

The 35% of propionic acid formed in rumen is converted to lactic acid in epithelial cells and they can get into the bloodstream in this form.




Nearly half of the butyric acid formed during butyric acid fermentation, gets directly to the bloodstream and is transformed to acetyl-CoA in the liver. Acetyl-CoA is oxidized in the citric acid cycle and in the oxidative phosphorylation and supplies energy to animals.

The other part of the butyric acid is transformed to β-hydroxy-butyric acid in epithelial cells, and is transformed further to aceto-acetate by oxydation. In this process two moles of acetyl-CoA are generated, which produces energy in the mitochondria of cells.

Energetically, the best form of fermentation process in the rumen is the propionic acid fermentation (Fig. 34).

10.12. ábra - Figure 34: Propionic acid and butyric acid fermentation

2.3.2. 10.2.3.2. The fermentation processes in silo

The fermentation processes taking place in silo show significant differences (besides many similarities) compared to the processes taking place in rumen. These are presented in Table 6.

During ensiling (in preservation process) the plant is still alive, respirating, so beside CO2 and H 2O heat is also released. The CO2 has a higher density than air that is why it falls to the bottom of the silo and displaces the air. In the oxygen-free condition the fermentation starts.

In preservation process the lactic acid fermentation is the best, the process is influenced by many factors. The quality of product forming at the end of reaction chain is influenced by the quality and carbohydrate content of fodder and also influenced by temperature, the moisture content and the anaerobic-aerobic conditions.

10.13. ábra - Table 6: The fermentation processes in silo




The lactic acid bacteria are able to ferment mono and disaccharides. The concentration of sugars must be so high to let the bacteria to grow, meanwhile the pH has to fall below 4.5. At the beginning of the process heterofermentation takes place, lactic acid, ethyl alcohol and acetic acid are formed, while the pH begins to decrease. Because of the acidification the growth of proteolytic, putrefactive bacteria slow and fungi accumulate (moulds). Moulds are not sensitive to low pH. Lactic acid generated previously is used by moulds.

In poorly compressed silages volatile fatty acids are formed by the acetic acid-butyric acid mixed fermentation process.

Yeasts also grow at low pH and produce ethyl alcohol from pyruvic acid and beside this acetic acid may also be formed. The slow decrease of pH leads to the formation of unfavourable esters. The esters are detrimental to the quality of the silage.

If the pH ranges from 4.5 to 5 in silo, the lactic acid-producing Streptococci multiply. The decrease of pH (pH <4.5) favours the growth of Lactobacilli, which ferment lactic and acetic acids. Formation of acetic acid is not favourable because the fodder becomes quickly acidic.

The decrease in pH below 4 favours the growth of Clostridium species, producing mainly butyric acid (but butyl alcohol, ethyl alcohol, isopropyl alcohol and acetone as well). The life activities of these species in silage reduce the quality of silo. Their also harmful activities are that they hydrolyze proteins and with deamination of amino acids produce iso-acids.

The volatile fatty acids arising in mixed acid fermentation processes in large amounts are weaker acids than lactic acid. This causes an increase in pH, whereupon the keeping quality of silage decreases. The ammonia released by deamination of amino acids also increases the pH.

3. 10.3. Gluconeogenesis (Glucose-resynthesis)One of the possible way of glucose formation is the gluconeogenesis, which is an opposite process to glucose breakdown. The gluconeogenesis is defined as the biosynthesis of glucose (carbohydrate) from intermediates of catabolic processes. The principal substrates for this process are lactic acid, produced from glycolysis in skeletal




muscle and amino acids, generated from dietary protein or from the breakdown of muscle protein during starvation. The carbon framework of some amino acids entering the citric acid cycle can be transformed to an intermediate product, which might be the precursor for resynthesis. These amino acids may be arginine, aspartic acid, glutamic acid, histidine, methionine, proline, serine, threonine, tryptophan and valine. The resynthesis takes place in the liver. The principal substrate of the process is phosphoenol pyruvate (Fig. 35).

10.14. ábra - Figure 35: Gluconeogenesis

Gluconeogenesis (resynthesis) occurs in glycogen mobilization or in intense muscle workout, when sugar is generated from lactic acid. Cells also produce sugar from oxaloacetic acid during starvation, in the absence of insulin, or even in carbohydrate-free diet. In such cases, the concentration of oxaloacetic acid may decrease in cells, which may be the main cause of the formation of ketone bodies as well.

In the case of high input or input of bad quality protein, the organism is not able to build its amino acids and proteins, therefore sugars will be released from intermediates of these proteins breakdown. Then the energy efficiency of formation of glucose is not favourable, because energy is needed for deamination and for removing ammonia resulting from this process (urea synthesis). Significant precursor of glucose in ruminants is the propionic acid, from which the glucose can be formed with high efficiency. In this process propionyl-CoA is synthesized first from propionic acid using ATP and CoA, then after its carboxylation methyl-malonyl-CoA is synthesized. In the next step the succinyl-CoA is produced from methyl-malonyl-CoA by a mutase enzyme, which is transformed to oxaloacetic acid in the citric acid cycle. The oxaloacetic acid may transform to glucose as described above.

4. 10.4. Glycogen metabolism4.1. 10.4.1. Glycogen synthesisHigh amount of polysaccharide are stored in mammalian liver and muscle as glycogen. Mammals need to store energy in this form because the breakdown of glycogen and the release of its energy are faster than that of fats. The carbohydrates are absorbed in the form of monosaccharides. After meals the blood glucose level rises. Glucose enters the liver and it is converted to glucose-6-phosphate, which is in equilibrium with glucose-1-phosphate. Due to the high concentration of glucose-6-phosphate the synthesis of glycogen starts.

The reaction process is as follows:

The energy of the glucose-1-phosphate is not enough to connect to the starter molecule (more connected glucose units) by glycosidic bond, and therefore reacts with uridine triphosphate and thus takes up energy from UTP.




The uridine diphosphate-glucose (UDP-glucose) is an energy-rich precursor. This molecule is able to connect with 1 → 4 ulinkage, while UDP is released.

Glycogen is composed of glucose molecules ulinked together linearly by 1→4 glycosidic bonds with branches are created by 1→6-glycosidic bonds. Glycogen synthesis involves both polymerization of glucose molecules and branching from 1→6 ulinkages. These branches can be introduced by branching enzyme called amylo-(1,4→1,6)-transglycosylase.

Glycogen is synthesized mainly in the liver and in the muscles. In muscles, the precursor of glycogen synthesis is glucose, while in the liver beside glucose other organic compounds can also be precursors (lactic acid, oxaloacetic acid).

4.2. 10.4.2. Glycogen mobilization, catabolismThe stored carbohydrates, glycogen is broken down in the storage organs, in the liver and in muscle. The mobilization of glycogen starts when the blood sugar level of the body is reduced after exercise or when stressed state occurs.

The central enzyme of glycogen catabolism is the phosphorylase, which tears off monomers from the ends of the polysaccharide chain, while transforms them phosphoric-ester form. The end product of the breakdown process is the glucose-1-phosphate. Glycogen phosphorylase catalyses the removal of the terminal glucose of glycogen when the bond is 1 → 4 ulinkage and stops glucose residues from a branch point (1 → 6 bond), producing a limit-dextrin. The limit-dextrin is degraded by α-(1→ 6) glycosydase enzyme. The glucose-1-phosphate originated from the breakdown is rapidly converted to glucose-6-phosphate by isomerisation process.

The glucose-6-phosphatase enzyme can be found in the liver, which cleaves off phosphate group from glucose-6-phosphate and thus glucose is produced. The membrane of the liver is permeable for the glucose resulted in this process, so it can get into the bloodstream.

There are no free glucose molecules formed from muscle-glycogen in the muscle, because there is no phosphatase enzyme in the muscle, which could catalyze the process of forming glucose from glucose-phosphate. In the muscle, lactic acid is formed from the breakdown of glycogen. One part of the lactic acid can be oxidized to carbon-dioxide and water, while the other enters into the liver via the bloodstream and is converted to glucose and glycogen. The sugar formed in the liver gets into the muscles via the bloodstream, where it is involved in energy supply. The circuit between the muscles and liver is called the Cori cycle.


11. fejezet - 11. THE METABOLIC PROCESSES II. LIPID METABOLISM1. 11.1. Biosynthesis of lipids1.1. 11. 1. 1. Biosynthesis of trigliceridesThe biosynthesis of triglycerides can be divided into three inter-related reactions. In order to the formation of triglycerides, fatty acids and glycerin should be synthesized first, and after that, they have to be ulinked.

1.1.1. 11.1.1.1. Biosynthesis of fatty acids

The biosynthesis of saturated straight-chain fatty acid takes place in the cytosol of the cells. The precursor of the synthesis is the acetyl-CoA. This precursor can be derived from the degradation of fatty acids, it can also be derived from the decarboxylation of pyruvate, which is the end product of glycolysis or it can be originated from the degradation of amino acids (from their carbon chain). These processes take place in the mitochondria, thus acetyl-CoA needs to get to the cytosol, which is an energy-intensive process.

Another important precursor of this process is malonyl-CoA, which is also formed from acetyl-CoA by carboxylation. The reaction is catalyzed by acetyl-CoA carboxylase, which has a biotin prosthetic group. CO 2, Mg2+ and ATP also need to run the process and for the enzyme to work.

The biosynthesis of fatty acids is catalyzed by fatty acid synthase multi-enzyme complex. This enzyme complex contains six enzymes and an acyl carrier protein group, the ACP. The specific binding site of ACP is the SH-group, which is able to bind the acyl group. The ACP are surrounded by six enzymes. The ACP holds the molecules of the reaction by covalent bonds and forward them to one of the active sites of the enzyme to the other.

As shown in Figure 36 the ACP first binds the acetyl and malonyl starting materials. In the next step malonyl condenses with acetyl group to form acetoacetyl group. This condensation reaction is coupled with the loss of carbon dioxide (decarboxylation). One SH-group of the ACP enzyme complex becomes free. In the first reduction step acetoacetyl group forms β-hydroxy-butyryl group by NADPH+H+ and yields crotonyl group after dehydration. This step is followed by reduction with a second molecule of NADPH+H+ to form butyryl group. Then the free SH-group of the ACP binds another malonyl group and the process steps described above are repeated. One cycle results the lengthening of the chain with one C2 unit, and a total of seven such cycles leads to the formation of a molecule of 16-carbon palmitic acid. When palmitic acid is formed, the synthesis is ended.

The longer chain fatty acids are formed from palmitic acids torn from the multienzyme in the mitochondria or elongation of fatty acids may occur at edoplasmic reticulum by attaching other acyl groups.

11.1. ábra - Figure 36: Biosynthesis of fatty acids


11. THE METABOLIC PROCESSES II. LIPID

METABOLISM

The equation of biosynthesis of 16-carbon palmitic acid:

The unsaturated fatty acids are formed from saturated fatty acids. The desaturation reaction is catalyzed by microsomal oxygenase enzyme complex in the presence of oxygen. Its operation is membrane-bound. The production of polyunsaturated fatty acids (linoleic acid, linolenic acid) in plants happens by further oxidation of oleic acid. Animals are not able to synthesize these compounds, so they are essential for them.

1.1.2. 11.1.1.2. The synthesis of glycerol

The glycerol is originated from the dihydroxy-acetone-phosphate in the process of glycolysis. By reduction of this compound glycerol-1-phosphate is formed, which is the active form of the glycerol. The synthesis of triglycerides from glycerol-1-phosphate and activated fatty acids takes place in the liver and adipose tissues. The fatty acids are activated by ATP, AMP is incorporated (R-CO-AMP), while pyrophosphate is released. The energy-rich, activated fatty acid has ability to bound to CoA, thus acyl-CoA (R-CO-SCoA) is formed.

The activated fatty acids can react with the activated glycerol (glycerol-1-phosphate). In the first step of the process, lysophosphatidic acid is formed from the reaction of glycerol-1-phosphate and fatty acid-CoA, while CoA-SH is released. This process is catalyzed by glycerol-phosphate-acyl-transferase. Another acyl group is attached to lysophosphatidic acid catalyzed by glycerol-phosphate-acyl transferase enzyme, while phosphatidic acid is formed (Fig. 37).

The phosphate group of phosphatidic acid is hydrolyzed by phosphatidate-phosphatase. Another acyl group is connected to the free hydroxyl group. This process is catalyzed by acyl transferase. The triglycerid is thus formed.

11.2. ábra - Figure 37: The synthesis of triglycerid



METABOLISM

1.2. 11.1. 2. Biosynthesis of phospholipidsPhospholipids are the components of biological membranes and transport lypoproteids. Phospholipids are formed by the esterification of phosphatidic acid. A compound with alcoholic hydroxyl group is bounded to the phosphoric acid part of phosphatidic acid, while water is released.

The initial substance of the synthesis so is phosphatidic acid. Phosphatidic acid is activated by CTP. An energy-rich intermediate, CDP-diacyl-glycerol is formed in this process, while pyrophosphate group is released. The energy-rich CDP-diacyl-glycerol is able to bind kolamin, kolin, serine and inositol by ester ulinkage. The end products of this process are phosphoglyceride and CMP. The phosphoglycerides can be formed not only directly, but can also be transformed into each other by chemical processes.

Ethanolamine phosphatides (cephalins) may be formed by decarboxylation of phosphatidylserine, while from ethanolamine phosphatides with bonding three methyl groups yield choline phosphatides (lecithin).

There are two biosynthetic routes known to synthesize phosphatidylcholine. One pathway is the previously described process, when the ethanolamine part of phosphatidyl ethanolamine is directly methylated. Phosphatidyl ethanolamine undergoes three successive methylations, where methyl groups are derived from three S-adenosyl-methyonines. The reaction is catalyzed by phosphatidylethanolamine methyl transferase.

The other pathway is when choline is originated from food breakdown and is utilised. In the first step choline is phosphorylated by ATP and the resultant phosphocholine (in high-energy state) undergoes a cytidyltransferase reaction catalyzed by phosphocholine cytidyltransferase to give CDP-choline (beside pyrophosphate is released). The phosphocholine group is transferred to diacylglycerol catalyzed by phosphocholine transferase, yielding phosphatidylcholine and CMP as byproducts.

1.3. 11. 1. 3. The biosynthesis of carotenoids and steroid skeleton lipidsThe carotenoids and steroids are formed from isoprene molecules. The precursor of the isoprene units is the acetyl-CoA.

11.3. ábra - Figure 38: The synthesis of isoprene



METABOLISM

In the first step of the reaction, two acetyl-CoA molecules are connected to yield aceto-acetyl-CoA. In the next step, another acetyl-CoA is connected to aceto-acetyl-CoA to yield β-hydroxy-β-methyl-glutaryl-CoA. After reduction this compound is transformed to mevalonic acid. Mevalonate is phosphorylated by pyrophosphate. This will increase the energy of the compound, which is able to transform now to isopentenyl pyrophosphate in a chemical process. Isopentenyl pyrophosphate (active isoprene) is a precursor of the synthesis of carotenoids (Fig. 38).

1.3.1. 11.1.3.1. The synthesis of steroids

All the carbon atoms of cholesterol (the base compound of steroids) originate from acetate. The steroid skeleton is formed in several steps with coupling of 30 active isoprene molecules (isopentenyl pyrophosphate).

One of the intermediates of cholesterol synthesis is squalene. Squalene is an open chain triterpene, which after cyclization yields cholesterol.

The final stage of the cholesterol-synthesis is the sum of complicated processes:

• ring closing,

• migration of hydride,

• migration of methyl groups,

• saturation of unsaturated bonds.

Cholesterol serves as precursor to all of the steroid compounds, the corticoids, the sexhormones and vitamin D.

2. 11. 2. The breakdown of lipidsLipids are the major energy source in most cells. Lipids are apolar compounds; they do not absorb water, so their place claim is small. The metabolic oxidation of lipids yields large amount of metabolic energy. The neutral fats are the most abundant class of lipids in terms of the energy storage. Energy released during their metabolism gives half of the oxidative energy of liver, kidney, heart muscle, skeletal muscle. Under conditions of starvation the fat is almost the only source of energy. The brain, though it has high lipid content, is unable to use fatty acids as energy supply, therefore the brain gains energy from the oxidation of glucose.

The breakdown of lipids occurs through action of pancreatic lipase in the small intestine. These enzymes are produced in an inactive form and become active in the intestine. The phospholipase A hydrolyses the phospholipids, which is also produced in pancreas and in intestinal mucosa.

Bile emulsifies lipids, yields micelles and ensures a large surface area for the function of enzymes.



METABOLISMThe 1-3% of neutral fats as triglycerides, nearly half of them in the form of mono-and diglycerides, while the other half is completely hydrolyzed as fatty acids and glycerol, are absorbed from the gastrointestinal tract.

Glycerol originated from the hydrolysis of triglycerides is phosphorylated by using ATP in the liver. This process is catalyzed by glycerol kinase. The L-glycerol-3-phosphate yielded of this process may be the precursor of the biosynthesis of triglycerides or phosphoglycerides. After dehydrogenation L-glycerol-3-phosphate generates dihydroxyacetone phosphate, thus it can enter the glycolysis process.

2.1. 11. 2.1. The β-oxidation of saturated fatty acidsFatty acids arise in the cytosol, but their breakdown takes place in the mitochondria. The fatty acids must be activated to get through the mitochondrial membrane. It is an energy-intensive process (ATP). First, the fatty acid reacts with coenzyme A to form acyl-CoA. This is an activation step because the fatty acyl-CoA is an energy-rich compound and is more reactive. This reaction is coupled with the hydrolysis of ATP.

Since the mitochondrial membrane is impermeable to fatty acids or acyl-CoA-s, a specific transport system is needed to move them into the mitochondrial matrix, where oxidation occurs. This involves transfer of the fatty acyl moiety to a carrier called carnitine. The next step involves the formation of fatty acyl-carnitine, which can traverse the membrane.

Inside the mitochondrial matrix the acyl group of acyl-carnitine is passed through the mitochondrial CoA-SH. Fatty acid are oxidized in a series of repeating steps called β- oxidation. The steps of the β-oxidation of fatty acids are marked with 1-4 numbers (as the four steps) in Figure 39.

The first (1) of four steps is a dehydrogenation process and is catalysed by acyl-CoA dehydrogenase. In this process, the acyl thioester is oxidized by FAD to give an enoyl derivative. The dehydrogenation takes place between the α- and β-carbons and yields a trans-isomer (trans-enoyl-CoA). The next step is a hydration process. This reaction is catalyzed by enoyl-CoA hydratase. The product is 3-hydroxy-acyl-CoA, which contains a chiral carbon atom. In this reaction L-isomer is formed. Step 3 is the second oxidation where dehydroganation of the hydroxyl group takes place. In this process, 3-keto-acyl-CoA is formed from L-3-hydroxy-acyl-CoA. The process is catalyzed by L-3-hydroxy-acyl-CoA dehydrogenase and it is a NAD-dependent, stereospecific dehydrogenation. The enzyme can only carry out the conversion of L-isomers.

The last step (4) of the cycle is a cleavage by attack of a second molecule of coenzyme A on the β-carbon, to release acetyl-CoA and thus acyl CoA will be two carbons (C2) shorter than the original substrate. This process is catalysed by acetyl-CoA acyltransferase. Then the cycle starts again, and it will continue until the chain is completely reduced.

C2 units formed in the oxidation process can enter in the citric acid cycle.

The oxidation of unsaturated fatty acids occurs similarly as in the case of saturated fatty acids, until the breakdown is attained the unsaturated bond of chain. Here the reaction is stopped, because of the cis-configuration. Enoyl CoA hydratase can not be acted on, because it acts only on trans-compounds.

11.4. ábra - Figure 39: The β-oxidation



METABOLISM

The position of double bond is also wrong. During the oxidation of unsaturated fatty acids the double bond occurs between the C3-C4, while it has to be in C2-C3 position. To continue the process the isomerisation of double bond and the change of double bond position are required.

The problem is solved by two auxiliary enzymes. The enoyl-CoA isomerase catalyzes the reversible rearrangement of the double bond. The other auxiliary ezyme enoyl-CoA hydratase creates the L-3-hydroxy-acyl-CoA, from which the reaction can be continued as already described.

In cells, fatty acids with odd-numbered carbon chain can also be found in small amounts. Their breakdown occurs as the same method as the even-numbered ones. In the last step, propionyl-CoA (C3 units) is formed instead of the acetyl-CoA. The propionyl-CoA has to be converted into a form, which is capable to enter one of the sections of catabolic processes. The conversion steps are shown in Figure 40. In the first step, propionyl-CoA is transformed to methyl-malonyl CoA by carboxylation, which is an energy-intensive process. Then succinyl-CoA is formed by carboxyl mutase enzyme, which can now enter the citric acid cycle. Its breakdown will be continued there.

11.5. ábra - Figure 40: Breakdown fatty acids with odd-numbered carbon

The cells gain large amount of metabolic energy by breakdown of fatty acids. In the following, we can write the total energy balance of oxidation of stearic acid (C18).

Stearic acid oxidation yields: 9 acetyl-CoA, 8 FADH2, 8 NADH+H+



METABOLISMThe equation for catabolism of acetyl-CoA through the citric acid cycle:

2 CO2, 1 NADPH+H+, 2 NADH+H+, 1 FADH2, 1 GTP

9 acetyl-CoA: 18 CO2, 9 NADPH+H+, 18 NADH+H+, 9 FADH2, 9 GTP

In the terminal oxidation:

9 NADPH+H+ → 3 ATP*9= 27 ATP

26 NADH+H+ → 3 ATP*26= 78 ATP

17 FADH2 → 2 ATP*17= 34 ATP

Beside the 9 GTP-s (GTP=ATP) generated during the catabolism 139 ATP-s are also generated (the sum of ATP is 148), from which two ATP-s are necessary to use for the active transport of fatty acid to the mitochondrium.

As previously deduced the oxidation of one glucose (C6) molecule to carbon dioxide and water yields 38ATP-s. On the basis of data, it can be stated that a correspondingly larger amount of metabolic energy is released in the case of metabolic oxidation of fats than in the oxidation of carbohydrates.

2.2. 11.2.2. The catabolism of steroidsCholesterol is one of the most important steroids. The largest amount of cholesterol is excreted in the bile, after appropriate transformations it solves as salts of bile acids. It plays an important role in lipid metabolism, in the absorption. Most of the bile acids are absorbed from the intestine and utilized for emulsifying lipids (with a split from the chain substituted C17 atom).

90% of cholesterol taken up orally is converted to bile acids and a small amount of that is transformed to coprostanol by bacteria. It passes out of the body as faeces. Sebaceous glands of skin can also secrete 100 to 300 mg of cholesterol per day.

In the skin cholesterol can transform to calciferol (vitamin D) through enzymatic or photochemical reactions.

In the endocrine glands cholesterol transforms to pregnenolone that is the precursor for the steroidal hormone synthesis. Pregnenolone transforms to progesterone. It is a hormone compound and the precursor of the adrenal glands hormones progesterone as well. Pregnenolone and progesterone transform to oestrogens and androgens (sex hormones) in the adrenal cortex.

3. 11. 3. The formation of ketone bodies (ketogenesis)Collectively, β- hydroxybutyric acid, acetoacetic acid and acetone are called ketone bodies. Ketone bodies are created in the body in normal conditions. There are some amino acids (ketogenic amino acids; lysine; leucin) that broke down and their carbon skeleton enters the process of ketogenesis. Large quantity of ketone bodies can be formed in the metabolism of fats.

The acetil-CoA is produced from partial oxidation of fatty acids and transposted by the CoA-SH into the citric acid cycle mainly and it will be transformed further. It could also be happened to acetyl-groups as well which are derived from the catabolism of carbohydrates and some amino acids. The process requires large amounts of oxaloacetic acid as the acetyl CoA connects to this compound at the first step in the citric acid cycle.

When there is not enough oxaloacetic acid, it is unable to accept the acetyl-group and the ketogenesis will start, so the acetyl-CoA is then used in the synthesis of ketone bodies. The concentration of ketone bodies in the blood increases.

If due to starvation or diabetes the glucose level in the body remains low, the body generates glucose from one part of oxaloacetic acid to solve the problem. At the time of starvation or in case of diabetes the concentration of ketone bodies may multiple and the tissues are not able to digest them (ketosis).

In this case the ketone bodies can also appear in the urine (ketonuria). This condition is dangerous, because acetoacetic acid and acetone are harmful for the nervous system.



METABOLISMIn the first step acetoacetyl-CoA can be formed from 2 moles of acetyl-CoA by the CoASH transferase enzyme.

In the second step β- hydroxyl-β- methyl glutaryl-CoA forms from this compound. This process is catalyzed by hydroxymethyl-glutamyl-CoA synthetase and during the process another acetyl group enters.

These compounds are synthesized in the liver and the circulatory system delivers them to peripheral tissues and there they are oxidized.

The β- hydroxyl-β- methyl glutaryl-CoA is a precursor of steroid skeleton synthesis.

11.6. ábra - Figure 41: Ketogenesis

The β-hydroxybutyrate can be oxidized to acetoacetate and the opposite process can also occur.

β -hydroxybutyrate + NAD+ ↔ acetoacetate + NADH + H+

Acetoacetate can transform to acetone by decarboxylation. In peripheral tissues ketone bodies can join the catabolism.

In the mitochondria acetoacetate is activated from succinyl-CoA by means of CoA-transfer:

succinyl-CoA + acetoacetate → succinate + acetoacetyl-CoA

Succinate transforms to further compounds in the citric acid cycle. Acetoacetyl-CoA is converted into two mols of acetyl-CoAs with using of a CoA. These compounds then get into the citric acid cycle.

The brain converts large amounts of ketone bodies to gain energy if the glucose supply is inappropriate during starvation (Fig. 41).

4. 11. 4. Glyoxylic acid cycle (Kornberg Krebs cycle)This modified citric acid cycle occurs in some microorganisms and in germinating oilseeds. In this cycle anabolism dominates, thus it is contrary to citric acid cycle, where the main goal is catabolism. In the process, the starting amount of oxaloacetate is doubled in every turn of the cycle.

The balanced equation of the cycle:

2 acetyl-CoA + 3H2O + FAD + 2NAD+ → oxaloacetic acid + 2CoA + FADH2 + NADH+ +H+



METABOLISMThe oxidation of the intermediates differs from citric acid cycle in the trasformation of isocitric acid. In citric acid cycle the citric acid is decarboxylised oxidatively. However, here isocitrate lyase cleaves isocitrate to glyoxylic (C2) and succinic acid (C4). Glyoxylic acid reacts with the second acetyl CoA. The process is catalyzed by the enzyme malate synthase and malic acid forms. Malic acid is oxidized to oxaloacetic acid.

From succinic acid cleaved by the enzyme isocitrate lyase another oxaloacetic acid forms through the already known pathway: fumaric acid → malic acid → oxaloacetic acid. It is then ready to start another cycle again. The enzymes of the process at this stage are the same as with that of citric acid cycle. The glyoxylic acid cycle requires only one oxaloacetic acid, thus the other one is utilized for carbohydrates or amino acid synthesis (Fig. 42).

11.7. ábra - Figure 42: Glyoxylic acid cycle

Plants synthesize carbohydrates from fats by using glyoxylic acid cycle. In the process acetyl-CoA transforms oxaloacetic acid through glyoxylic acid cycle. Oxaloacetic acid is transformed to phospho-enol-pyruvate (PEP). The process is catalyzed by phospho-enol-pyruvate carboxylase enzyme. Glucose is built up from PEP via multi-step process. During the sequential reactions the steps of glycolysis take place inversely.


12. fejezet - 12. THE METABOLIC PROCESSES III. PROTEIN METABOLISMNitrogen can get into the food chain through nitrogen uptake by plants.

Plants can take up nitrogen from the soil as ammonium and nitrate ions. Although air contains nitrogen in large quantities, this form is not available for plants. Atmospheric nitrogen can be fixed by bacteria that live free in the soil or in symbiosis with certain plants. These plants use atmospheric nitrogen taken up and converted by bacteria. Nitrogen forms taken up by plants has to be reduced in plants.

1. 12.1. The nitrogen fixationThe symbiotic bacteria and host plants can fix and reduce nitrogen mutually. In there common metabolism bacteria are the N-source, while host plants are the C- source.

The nitrogen fixing bacteria contains nitrogenase enzyme that plays a central role in the nitrogen fixation. The enzyme consists of two subunits, which is a complex protein containing iron and molybdenum. One of the subunits only contains iron. The nitrogenase enzyme reduces molecular nitrogen to ammonium ion.

The process requires electrons (reduced coenzymes) and large amounts of energy (ATP) which are provided by the host plant. The transformation of 1 mole nitrogen into 2 moles ammonium ion requires 12 moles ATP.

Some parts of the reduced coenzymes from catabolic processes of the host plant enter the transformation directly by providing electrons. The other part enters the terminal oxidation and provides energy for the process. In the host plant the citric acid cycle works intensively, which requires oxygen-rich conditions. However anaerobic environment is needed for the reduction. This contradiction is resolved by the tissue structure of the root nodules.

Ammonium ion formed in the reduction process gets onto glutamic acid. The first product of the process is glutamine. It is transported as aspartic acid that forms in the process of trans-amination. The fate of ammonium ions taken up by the plant is the same as the previous one.

The plant must also reduce nitrate ions taken up from soil to ammonium ions. The process consists of two steps.

First, the enzyme nitrate reductase reduces nitrate ions to nitrite ions then nitrite reductase converts nitrite ions to ammonium ions.

The process requires electrons that can originate directly from the light phase of photosynthesis. In this case the electrons from ferredoxin do not get to oxidized NADP+, but to the nitrite ion. In the first stage of the reaction sequence (nitrate ion → nitrite ion) electrons are indirectly from the reduced coenzyme.

The amino group of the amino acids forms by the incorporation of ammonium ions. With the direct uptake of ammonium ion glutamic acid and glutamine can be synthesized. The reactions are shown in Figure 43. Ammonium ion ulinks to α-keto-glutaric acid by reductive amination.

In the process that is catalyzed by glutamate dehydrogenase glutamic acid forms by means of, NADPH + H +. Glutamic acid transforms to glutamine with energy input and ammonia bound. Glutamine synthetase catalyzes the amide biosynthesis.

Glutamic acid synthase synthesizes two moles of glutamic acids from α-keto glutaric and glutamin.

12.1. ábra - Figure 43: The nitrogen fixation


12. THE METABOLIC PROCESSES III. PROTEIN

METABOLISM

Glutamine stores ammonia in amide bond. During trans-amination reaction the stored ammonia get onto the corresponding oxo acid (carbon skeleton of amino acids) and form other amino acids.

pyruvic acid (1) + glutamic acid (2) → alanine (1) + α-keto-glutaric acid (2)

oxaloacetic acid(1) + glutamic acid (2) → aspartic acid (1) + α-keto-glutaric acid (2)

The carbon skeleton of amino acids comes from the intermediates of different metabolic pathways.

The α-ketoglutaric acid is an intermediate of citric acid cycle and it can convert to glutamic acid and glutamine. This compound is also the precursor of proline and arginine. These amino acids are also referred to as members of the glutamine family.

Oxaloacetic acid can also exit the citric acid cycle. It is trans-aminated and converted to aspartic acid that transforms to asparagine by binding a further ammonium ion with amide bond. The starting material of methionine and lysine synthesis is also oxaloacetic acid. Threonine also forms from oxaloacetic acid, which converts to isoleucine. The amino acids the synthesis of which starts with oxaloacetic acid are listed in aspartic acid family.

The synthesis of alanine, valine and leucine starts with pyruvic acid that is the final product of glycolysis. They are the members of pyruvic acid family.

Glyceric acid-3-phosphate can leave the process of glycolysis or photosynthesis and it can become the carbon skeleton of cysteine and serine. Glycine forms through the conversion of serine. These amino acids belong to the serine family.

Ribose-5-phosphate that is the intermediate product of the light independent phase of photosynthesis can be the carbon skeleton of histidine. Phospho enol pyruvic acid and erythrose-4-phosphate are also members of the reaction sequence of photosynthesis, and they together form the carbon skeleton of tyrosine, phenylalanine and tryptophan. They are cyclic amino acids.

2. 12.2. The synthesis of essential amino acidsThe human body and mammals can not synthesize some amino acids, they can only get them with nutrition. The majority of plants and bacteria are able to synthesize the amino acids, the synthesis routes are similar, but much more complicated than that of non-essential amino acids.

2.1. 12.2.1. The methionine and threonine biosynthesisThe common intermediate of methionine and threonine synthesis is homoserine. In mammals these amino acids are essential, as they have lack of that reaction way section in which homoserine is formed from aspartic acid.

The steps of the process:



METABOLISMAspartic acid beta-aspartyl phosphate → aspartate - semialdehyde → ß-homoserine.

From homoserine threonine is formed through homoserine phosphate by threonine synthase.

2.1.1. 12.2.2.1. Methionine formation from homoserin

The formation of methionine from homoserin starts with the formation of O-succinate. In this step succinyl group is attached to homoserine.

In the subsequent reaction cystathionine formed by means of the enzyme cystathionine γ-synthase. From this compound cystathionine β-lyase cuts water, ammonia and pyruvic acid, creating homocysteine. Homocysteine takes part in methylation process with methyl transferase in which the end product is methionine. Cystathionine is the starting material of both cysteine and methionine, but in mammals the cysteine and in plants the methionine can only be found, while in bacteria both of them are produced (Fig. 44).

12.2. ábra - Figure 44: The synthesis of essential amino acids

2.2. 12.2.2. Lysine biosynthesisIn bacteria and plants lysine biosynthesis passes through diaminopimelic acid, while in fungi through α-aminoadipic acid.

For the formation of diaminopimelic acid aspartate semialdehyde and pyruvate are required. They react each other during aldolcondensation then through a multi-stage reaction diaminopimelic acid is formed. After decarboxylation of this compound lysine is synthesized.



METABOLISM

2.3. 12.2.3. Arginine biosynthesisArginine is generated in the liver of mammals in large quantities, but this is immediately decomposed by the enzyme arginase. It follows that arginine produced by the liver can not be the arginine source of the body. The precursor of arginine synthesis is ornithine. Ornithine is formed by glutamic acid.

2.4. 12.2.4. Leucine, isoleucine and valine synthesisThe three branched-chain amino acid biosynthesis takes place in a similar route.

Ketoacid → active acetaldehyde + acetohydroxy acid

The processes are catalyzed by enzymes containing thiamine pyrophosphate. After further reactions, such as reduction, methyl-ethyl-migration, rearrangement, dehydration ketoanalog relevant to the amino acids is formed. From ketoanalogs through transamination amino acids are formed.

During the biosynthesis of valine from two molecules of pyruvic acid acetolactate synthase form α-acetolactate with carbon-dioxide release. In further steps α, β-dihydroxy isovalerate, and α-keto-isovalerate is formed. From it valine transaminase forms valin using the amino group from glutamic acid.

2.5. 12.2.5. The phenylalanine and tryptophan biosynthesisThe basic condition for the formation of these amino acids is the formation of 6-membered aromatic ring. The aromatic ring forms from aliphatic precursors. By plants the central intermediate in the formation of the aromatic ring is shikiminic acid. The lignin, ubiquinone and plastoquinone also form from shikiminic acid. The precursor of shikiminic acid is phospho-enol pyruvic acid and erythrose-4-phosphate.

The intermediate forms ring structure, then the process is followed by dehydration and reduction. After phosphorylation the shikiminic acid transforms to korizminic acid that is found in the branches of aromatic amino acid metabolism.

Korizminic acid is one of the products of the branches of metabolic pathways.

From it prephenic acid is generated, from which it is transformed to phenylalanine in a multi-stage process.

The korizminic acid can transform to antranilic acid as well, which synthesizes to tryptophan in additional transformations. The formation of the histidine starts with the bond of phospho-ribosyl pyrophosphate and ATP wherein the 5-phospho-ribosyl-glycosidic forms glycosidic ulinkage with first position nitrogen atom of the adenine part of ATP while pyrophosphate exits.

The 2nd carbon atom of phospho-ribosyl-2-pyrophosphate integrates to carbon skeleton of imidazole ring, whereas the 3rd carbon atom transforms to analin.

3. 12.3. Protein SynthesisDuring the biosynthesis of proteins information stored in DNA is transcribed into protein with appropriate amino acid sequence. The process takes place by means of the various types of RNA molecules.

The synthesis of proteins consists of two steps that can be divided to further sections.

1. During transcription information stored in the DNA is transcribed into RNA. The process takes place in the nucleus. (DNA → RNA)

The products move to the place of protein synthesis through nuclear pores.

2. During translation process information transcribed to RNA is translated into amino acid sequence on ribosomes. (RNA → protein)

The sequence of DNA nucleotides (N-bases) determines of the nucleotide sequence of the RNA. The sequence of RNA nucleotide triplets (base triplet) is the information that is responsible for the polypeptide amino acid sequence.



METABOLISM

3.1. 12.3.1. The transcriptionDuring the process RNA molecules (mRNA, tRNA, rRNA) are synthesized based on the DNA sample. The central enzyme complex of the process is the DNA-dependent RNA polymerase that consists of five subunits. The mRNA is the messenger RNA, the amino acid sequence of which determines the nucleotide sequence of the protein.

The tRNA is the transfer RNA, it transports the activated amino acids into the site of protein synthesis.

The rRNA is the ribosomal RNA, it is a part of the ribosome that is the center of the protein synthesis.

Transcription can be divided into three stages:

→ initiation is the start of the process

→ elongation is the prolongation of the chain

→ termination is the finish of the process.

Initiation begins with the recognition of promoter base composition on the initial section of the template DNA molecule. In the promoter section the RNA polymerase unscrews the double helix DNA molecule. In this section the two DNA strands are released. One of the strands of the DNA, the so called codic strand (starting with 3') serves as template. On the other strand there is no transcription – this strand remains 'silent'. The RNA synthesis occurs from the 5 'end to 3' end direction (Fig. 45).

12.3. ábra - Figure 45: Transcription

During elongation, monophosphate nucleosides attach themselves to the DNA in complementary order. The monophosphate nucleosides are formed from nucleoside triphosphate with pyrophosphate cleavage. The corresponding nucleotide monophosphate derivatives ulink to the appropriate 3'end.

At the final stage of chain construction (termination) the template DNA has palindromic structure. At this stage the sequence of bases is repeated as a mirror image. The end of RNA chain transcribed from them can form a hairpin loop. The RNA chain is cleaved by a protein from DNA. The site of termination is indicated with the hairpin loop RNA. The process requires energy (ATP). The resulting RNA molecules (pre-RNAs) transform during migration to the cytoplasm and they become mature. The non informal parts fall out. In the meantime the reactive groups at the ends of the RNA chain are protected from undesirable side reactions by compounds attached to them.

3.2. 12.3. 2. The translationThe second step of protein synthesis is the translation. The result of translation is the protein molecule. The process required for mRNA, tRNA, rRNA, ATP, GTP, factors, Mg ions and amino acids.

The general steps of the synthesis are also characteristic for the synthesis of other macromolecules. Monomers have to be activated. The activated units have to be connected to each other, than products have to break down from the apparatus of synthesis, and they have to form appropriate spatial conformation. These steps follow each other in the process of protein synthesis as well.



METABOLISMThe base sequence of messenger RNA (mRNA) determines the sequence of amino acids in the protein molecule. In an mRNA molecule three nucleotid units (base triplet) signifies an amino acid in the polypeptide chain. The majority of the amino acids are encoded by more than one base triplet.

There is a unique base triplet that is responsible for the start of the synthesis. AUG triplet is the start codon, which is the code of methionine. The synthesis can be finished by several base triplets (stop codons).

The ribosomal RNAs (rRNAs) connected with proteins form ribosomes in the endoplasmic reticulum. The protein synthesis takes place in ribosomes. Ribosomes consist of two subunits, one has small molecular weight and the other has larger molecular weight. During protein synthesis they are associated with Mg ions. Their task is to fix mRNAs, and the activated tRNAs that carry amino acids.

The biological role of transfer RNA (tRNA) is the transportation of activated amino acids into the place of synthesis, and inserting them into the protein chain. Amino acids can be transported by different types of tRNA molecules. (The 20 kinds of amino acids are transported by 60 kinds of tRNAs.) The tRNA molecules have a special cloverleaf conformation. There are three loops in their and the rest of the molecule is arranged in a double spiral.

The first loop (I) is the recognition site of the amino acids, i.e. that enzyme attaches here, which is responsible for amino acid binding.

The second loop (II) is the anticodon part. Anticodon is a triplet the sequence of which is complementary to the triplet of mRNA, thus they fit together. The anticodon part determines the amino acids binding to the tRNA binding site.

The third loop of tRNA (III) provides the binding to the ribosome. The activated amino acid binds to the free end of the molecule, which is the acceptor end. The end is the same (ACC) in all of the tRNA molecules. The hydroxyl group located at the 3rd carbon atom of nucleotide unit containing adenine is freely available (Fig. 46).

12.4. ábra - Figure 46: tRNA

The attachment of amino acids to tRNA

There are CCA base triplets on the binding sites of each tRNA (3 'end). Amino acids attach with ester bond to the 3’ carbon atom of ribose in the nucleotide containing adenine.

The nucleotide supplies the alcohol OH group, whereas the amino acid gives the carboxyl group for the ester bond. The process involving water loss is catalyzed by aminoacyl-tRNA synthetase enzyme. The amino acid has to be activated for the reaction. The activated amino acid is the aminoacyl-adenylate that ulinks to tRNA with a loss of AMP.

Three phases of translation can be separated: these are initiation, elongation and termination.



METABOLISM3.2.1. 12.3.2.1. Initiation

The initiation is signified with AUG that is the initiation codon of mRNA. In eukaryotes this codon is complementary to the anticodon part of tRNA bonding methionine.

For the initiation of the synthesis the initiation complex must be formed.

For the formation of initiation complex two subunits of the ribosome, mRNA and tRNA are required. The tRNA contains the initial amino acid in activated form that is the methionine-tRNA. GTP provides the energy required for the synthesis. Initiation factors and Mg ions (IF1, IF2, IF3,) play an important role. The steps of initiation complex formation are shown in the following figure (Fig. 47).

12.5. ábra - Figure 47: The steps of initiation complex formation

The IF3 factor binds to the smaller ribosomal subunit, thus it prevents the larger ribosomal subunit's association.

This is necessary to ensure the connection of the smaller ribosomal subunit to mRNA. In the second step, the initiation tRNA, IF2 and GTP arrive as a complex and reach the P site by means of IF1. In the third step the larger ribosomal subunit ulinks to the already formed complex. The process also requires magnesium ions.

During the connection GTP decomposes to GDP and phosphoric acid and the dissociation of initiation factors also takes place.

3.2.2. 12.3.2.2. Elongation

In the elongation stage of protein synthesis amino acids connect to the initiator amino acid one after the other in



METABOLISMa certain sequence specified by the mRNA. The polypeptide chain is growing. The information supplied by the mRNA transcribes in 5 '→ 3' direction forming polypeptide chain.

The rRNA moves along mRNA and it ensures the binding of tRNA. The involvement of new tRNA in the process takes place by means of elongation factors (EF-T and EF-G).

The new amino acid aminoacyl-tRNA - EF-T - GTP arrives to the ribosome as a complex and binds to the A binding site. The energy derived from GTP hydrolysis stabilizes the ribosome-aminoacyl-tRNA - mRNA complex.

The amino acids on the two tRNA molecules are attached via a peptide bond. The amino group of the newly arriving aminoacyl tRNA removes the amino acid from methionyl-tRNA by nucleophilic attack. The peptidyl transferase enzyme forms peptide bond with the carboxyl group of cleaved methionine and the amino group of newly arriving, but still RNA-bound amino acid. The formed dipeptide is located in A site and it is bound to the second t RNA. The amino acid-free (blank) tRNA remains in P site.

In order to continue the chain elongation tRNA containing the dipeptide has to shift to the P-site. The amino acid free tRNA has to leave the P-site. For the translocation of tRNA carrying dipeptide the factor EF-G and GTP are required. With the displacement of the tRRNA, according to the codon instruction another activated tRNA arrives into the vacant site. The subsequent steps of the process are the same as the previous ones.

In the final stage of protein synthesis (termination) the chain elongation is completed, the finished polypeptide chain is released from its apparatus. The completion of the synthesis is signified to the synthesizing system by mRNA via three nonsense codon (UAA, UGA or UAG). At this point the polypeptide chain is located at P site bound to tRNA. The tRNA corresponding to base triplets would arrive to the A-site, but there is not such tRNA molecule which anticodon part is complementary to these base triplet.

The role of terminator codons is to prevent the further elongation of the polypeptide chain. The recognition of these codons takes place by means of three release factors (RF1, RF2, RF3).

3.2.3. 12.3.2.3. Termination

In the first step of completion a releasing factor ulinks to the termination codon. The specificity of peptidyl transferase altered, thus it will be capable for hydrolyzing the ulinkage between the tRNA at the last site and the amino acid (polypeptide). The polypeptide chain and the tRNA are released and removed from the ribosome. In the next step, the mRNA and ribosomal disconnected. The ribosome dissociates and its subunits are ready to participate in the synthesis of a further polypeptide.

During synthesis the formation of the spatial structure of the polypeptide already begins. The formation of spatial structure does not need specific information, since it is determined by the amino acid sequence.

After termination the protein conformation is fully formed and meanwhile it undergoes through a maturation process. Here the starting amino acid can be cleaved, and the oxidation, methylation, phosphorylation of side-chains can occur. The amino group of N-terminal end is often acetylated.

Protein synthesis takes place only when the cells have adequate energy, because the construction of this macromolecule is a highly energy-inquiring process. This process consumes the most energy among all processes taking place in cells.

ATP (AMP + ATP → PPA) is required for the activation of amino acids, and for their binding to tRNA. GTP is used for the attachment of tRNA to the A-site. GTP is also required for the shift rRNA on mRNA. All in all the cleavage of four high-energy phosphate binding can result one peptide bond. 1/6 part of the invested energy only releases by the hydrolysis of this binding.

4. 12.4. The fate of dietary proteins in heterotrophic organisms4.1. 12.4.1. The quality of proteinsThe living organisms split dietary proteins to amino acids. The resulting amino acids as end products of



METABOLISMhydrolysis are utilized predominantly for the construction of its own body proteins. If necessary, they can be precursors of sugars (gluconeogenesis). They can be utilized by the organism for lipid or chemical energy production too.

In addition, they can be precursors of amino acids, hormones, porphyrins, purines, pyrimidines, alkaloids, coenzymes as well.

Excess amino acids not used by the body are degraded. Their nitrogen content empties in various forms. Their carbon skeleton decomposes into carbon dioxide and water. The various organisms have different amino acid requirement. Organisms can synthesize most of the required amino acids, but they can not produce one part of them, and they have to take it up by food. These types of amino acids are called essential amino acids.

Proteins have different values according to the satisfaction of amino acid requirement.

Proteins were classified on the basis of several aspects.

The biological value gives the percentage of the absorbed N that is incorporated into the organisms

It can be calculated as follows…

BV = ( ( Ni - Ne(f) - Ne(u) ) / (Ni - Ne(f)) ) * 100

Where:

• Ni = nitrogen intake in proteins on the test diet

• Ne(f) = (nitrogen excreted in faeces whilst on the test diet) - (nitrogen excreted in faeces not from ingested nitrogen)

• Ne(u) = (nitrogen excreted in urine whilst on the test diet) - (nitrogen excreted in urine not from ingested nitrogen)

The higher the biological value of the protein the more similar amino acid composition it has to human needs. Digestibility also affects its value.

Among raw materials that are often applied in nutrition the whole egg protein biological value is 100% that of milk is 93%. Among meat fish has 86%, beef has 85%, pork has 84% while chicken has 82% biological value. The biological value of the vegetable proteins is smaller than that of meats. The highest value has soy protein with 76% and potato with 74%. The BV of legumes is 66%, rice is 65%, wheat is 56%, while corn protein is 51%. Proteins of animal origin are called complete proteins while vegetable proteins are called incomplete proteins according to the biological value.

The chemical score of a protein is based on the relationship of its amino acid composition and its nutritional value. The examined protein is compared to the composition of a reference protein (FAO / WHO recommendations, eggs, meat, milk, etc..) To calculate a food’s chemical score, the amount of each essential amino acid provided by a gram of the food’s protein is divided by an “ideal” amount for that amino acid per gram of food protein.

This value does not provide information on the absorption of the protein and the fate of intermediates in metabolism. It is suitable for the determination of the limiting amino acids, thus this value can be used for the protein supplements.

The net protein utilization (NPU) gives the amount of retained nitrogen is related to consumed nitrogen percentage. This is the resultant of the certain protein’s digestibility, the absorption and utilization of the amino acids.

The protein efficiency ratio (PER) shows the amount of weight gain which 1g protein causes. This value is highly dependent on the quality of the protein.

The apparent digestibility can be calculated from the N content of the food and faeces, it is given in digestibility %.



METABOLISMDigestibility % = ((food N – faeces N)/food N)*100

The digestibility of milk, egg and meat are ~100%, while plant proteins are utilized in 80%.

The lower digestibility of plant proteins can be explained by the fact that there are their proteinase inhibitors in them that inhibit the function of protein decomposing enzymes.

4.2. 12.4.2. The protein balance of the organismThe proteins as the parts of every cell (plant or animal) are being destroyed and synthesized continually. The velocities of the intermediate metabolic processes are different in the different tissues. The lifetimes of the proteins are different:

The blood plasma proteins live for ~ 10 days, the muscle proteins exist for ~ 100 days.

The N balance of the organism, which is approximately equivalent to the protein balance is equal to the difference between the amount of intake N and emptied N.

Negative N - balance is in that body, which is in long-term shortage of protein. The organism does not receive the proper amount and quality of protein. This is often coupled with a lack of energy. In this case, the organism synthesizes sugar by protein breakdown complementing its energy in this way. After severe infections negative N balance can also occur, while there is an increased N-loss.

Building new tissues or muscles reflect in a positive N balance. During pregnancy and breast-feeding mothers have a positive protein (N) balance.

4.3. 12.4.3. The digestion of proteinsThe breakdown of proteins to amino acids is carried out by proteolytic enzymes (proteases). These enzymes break the peptide bonds between amino acids. They are hydrolases that hydrolyse bonds by water incorporation. The process is extracellular. Among proteases peptidases (exopeptidase) cleave amino acids from the end of the chain proteins, whereas proteinases are endopeptidases that cleave peptide bonds inside the chain.

The proteolytic enzymes are present in inactive form at the scene of the protein degradation that is the gastrointestinal tract by vertebrates. For their activation a smaller or larger- peptide chain must be cleaved by them. The activation is carried out by other enzymes, or the process is auto catalyzed.

Pepsin zymogen is a protein decomposition enzyme, the inactive form of which is pepsinogen. It works under acid conditions (pH 1-2). It can be activated by a cleavage of peptide chain from its N-terminal end, which has a molecular weight about 7000. This section contains the majority of basic amino acids of pepsinogen. Pepsin cleaves bonds mainly between amino acids with aromatic and other nonpolar side-chains.

The partially digested proteins leave the stomach and get to the small intestine. Here the pH is neutral. Further digestion of the proteins is carried out by chymotrypsinogen, trypsinogen, procarboxypeptidase A-B and proelastase which are produced in the pancreas in inactive form. The activation of trypsinogen is started by enteropeptidase, then it becomes autocatalytic process. In the process a hexapeptide is cleaved from the N-terminal end of trypsinogen. Trypsin cleaves peptide bond mainly next to carboxyl group in peptides. Chymotrypsin activates trypsin.

In the small intestine the mixture of exo-and endopeptidases hydrolyze the partially decomposed proteins to amino acids.

The amino acids are absorbed through the intestinal epithelium, which is an energy-requiring, active process. Amino acids from blood stream get into the tissues, where they are involved in metabolism. The metabolic pool of the cells is supplemented by amino acids formed from the degradation of tissue proteins. Intracellular protein breakdown is carried out by cathepsins.

Proteases vary widely in their optimum pH, they can be acid, neutral and alkaline proteases.

4.3.1. 12.4.3.1. Proteases occur in each cell. Their role is wide ranged.



METABOLISM• Proteases influence the composition of the cell protein substance.

• Proteases demolish the unnecessary proteins in order to build the necessary new protein molecules

• They promote the exit of secretory proteins from the cell by splitting the signal peptide.

• They activate inactive zymogens.

• They inactivate biologically active proteins that have already filled their functions.

• They inactivate defective proteins.

• They break down proteins to provide energy to the body during starvation.

4.3.2. 12.4.3.2. The common features of amino acid degradation pathways

The catabolic reactions of amino acids are catalyzed by multienzymes.

The substituents of α-carbon atom (carboxil-, amino groups) can be removed by similar catabolic pathway. The fate of reaction products are various during the metabolic processes.

One way of amino acids degradation is the cleavage of carboxyl group.

During decarboxylation carbon dioxide split off from the carboxyl group of the amino acid and bioactive amines are formed. The process is catalyzed by amino acid decarboxylases. They are characterized by narrow substrate specificity. Generally, they can only decarboxylate one amino acid. Pyridoxal phosphate is required for their operation. Glutamic acid decarboxylase breaks off carbon dioxide from L-glutamic acid. The resulting biogenic amines is the γ-aminobutyric acid.

After decarboxilation the residual biogenic amines are still after physiologically active compounds that are precursors of hormones, coenzymes or become building blocks of other materials. Some amino acids and biogenic amines resulted from them are shown in the Table 7.

12.6. ábra - Table 7: Biogenic amines



METABOLISMA small amount of amino acids only decomposes by decarboxylation to form biogenic amines. Another way of amino acid breakdown is the removal of amino groups. During deamination amino group is split off and oxo acids are formed.

The removal of amino group is carried out in the liver and kidneys in mammals. The process takes place by transamination or oxidative deamination.

In transamination reaction one part of the amino group of amino acids attaches to a new α -keto acid and forms another amino acid.

-amino acid + pyruvate ↔ α-keto acid + alanine

The process is catalyzed by transaminases. The process has a dual role. The amino-nitrogen of the amino acid is held back and built into another amino acid while the carbon skeleton of the amino acids is transformed in a way that it will be able to enter to the citric acid cycle.

During oxidative deamination ammonia is split off from amino acids (Fig. 48).

The process is catalyzed by NAD+, NADP+-specific amino acid dehydrogenase, amino acid oxidase or amine oxidases

12.7. ábra - Figure 48: Oxidative deamination (amino acids)

During the process catalyzed by glutamate dehydrogenase enzyme glutamic acid transforms to α-keto glutaric acid. The dehydrogenase enzyme is the only enzyme that operates under physiological conditions with appropriate intensity. Glutamic acid has a central role in the storage the amino nitrogen by transamination. Glutamic acid dehydrogenase enzyme has a role in the formation of nitrogen balance.

The amino acid oxidases catalyze the oxidative deamination of amino acids with using molecular oxygen as acceptor and producing ammonia.

amino acid + O2 + H2O → α-ketoacid + NH3 + H2O2

Amino acid oxidases are flavoproteins. These enzymes are present in animal cells (for example: in endoplasmic reticulum of the liver), appear in mushrooms, bacteria, but their occurence is not proved in higher ranked plants.

Plant amine-oxidases are less known, in the animal tissues mono- and diamine-oxidases are present.

amine + O2 → imine + H2O2

imine +H2O → aldehyde + NH3

In vertebrates the split amino-groups are emptied in the form of urea, uric acid or ammonium ion.

4.3.3. 12.4.3.3. The catabolism of carbon skeleton of amino acids in the tricarboxylic acid cycle

The carbon skeleton of amino acids are broken down via citric acid cycle into CO2 and H2O (ATP is generated).

Each of the 20 amino acids of proteins has a separate catabolic pathway. All 20 pathways converge into 5



METABOLISMintermediates, all of which can enter the citric acid cycle (Fig. 49).

• The acetyl-CoA pathway:

Alanine, glycine, serine, cysteine, isoleucine and thereonine are converted to pyruvate at first, then after decarboxylation it transforms to acetyl-CoA.

Leucin, lysine, tryptopan, phenylalanine and tyrosine are converted into acetoacetyl-CoA at first, then it transforms to acetyl-CoA.

Pyruvate converted from amino acids can also take part in the gluconeogenesis.

• The α-ketoglutaric pathway:

Glutamic acid, glutamine, proline, histidine and arginine are converted into glutamate that is then deaminated by a transaminase and forms α-ketoglutarate that enters the cycle.

• Succinyl-CoA pathway:

Four amino acids are broken down through propionyl CoA and methyl-malonyl-CoA intermediers. It transforms to succinyl-CoA that enters the cycle.

One part of the methionine, valine, threonine and leucine transforms through this pathway.

The sulphur content of methionine (when methionine is broken down) is built up into the cisteine.

• Fumarate pathway:

From a part of phenylalanine, tyrosine and fumaric acid aspartic acid forms which enters the cycle.

• Oxaloacetate pathway:

Aspartic acid and asparagine form oxaloacetate that enters the cycle.

The oxaloacetate can be the precursor of the glucose synthesis too.

12.8. ábra - Figure 49: Entering of carbon skeleton of amino acids the citric acid cycl

4.4. 12.4.4. Protein turnoverProteins are subject to continuous biosynthesis and degradation. The process is called protein turnover. Many of the amino acids released during protein turnover are reutilized in the synthesis of new proteins.



METABOLISMIn animals, protein intake can exceed the need for protein synthesis; the excess nitrogen is largely degraded. Depending on the lifestyle and the development of an organization, the nitrogen coming from the amino acids and other nitrogenous compounds can be excreted in different forms. Mammals excrete nitrogen in the form of urea, reptiles and birds excrete as uric acid and fish excrete nitrogen as ammonia or as trimethylamine-oxide.

4.5. 12.4.5. Nitrogen excretion4.5.1. 12.4.5.1. Nitrogen excretion in mammals, synthesis of urea (carbamide)

Those organisms can excrete most of their nitrogen as urea, whose liver contain arginase enzyme.

Urea is generated from two nitrogen atoms originated from two amino acids and from carbon dioxide in the liver of mammals. The urea formation requires energy.

The first amino-group is incorporated from free ammonia. The ammonia is generated from glutamic acid in the mitochondria. This process is catalyzed by glutamic acid dehydrogenase.

The first step in the formation of urea is the synthesis of carbamoyl phosphate from carbon dioxide and ammonia by using two molecules of ATP. This process takes place in the mitochondrial matrix and is catalyzed by the enzyme carbamoyl phosphate synthetase. The process is practically irreversible.

2 ATP + NH3 + CO2 + H2O → karbamoyl-phosphate + 2 ADP + Pi

This energy-rich group enters the cycle. In the second step, carbamoyl phosphate reacts with ornithine to form citrulline. Ornithine can enter the mitochondrial matrix with the help of a specific carrier and there it is connected to the carbamoyl group while inorganic phosphate breaks off. This reaction is catalyzed by carbamoyl transferase.

Citrulline gets to the cytosol from the mitochondria. The cycle goes on there. Ornithine and citrulline are non-proteinogenic amino acids. They are present only in small amounts in mammals.

The second nitrogen of urea comes from aspartate, which reacts with citrulline to form argininosuccinate by using ATP. This reaction is catalysed by argininosuccinate synthetase.

citrullin + aspartate + ATP → argininosuccinate + AMP + PPi

Aspartic acid ulinked to the carbonyl carbon atom of citrulline. The necessary energy for the reaction is provided by ATP. This reaction is coupled with the hydrolysis of ATP to AMP.

In the next step, cleavage of argininosuccinate produces arginine and fumarate. This reaction is catalyzed by argininosuccinate lyase. The fumaric acid returns to the mitochondria with the help of a carrier. Finally, hydrolysis of arginine yields urea and ornithine. This reaction is catalysed by arginase.

arginine + H2O → ornithine + urea

Ornithine can get back into the mitochondria with the help of a transport system. The cycle is thus completed (Fig. 50). Ornithine with taking up another carbamoyl phosphate is transformed to citrulline. Urea is transported in the bloodstream to the kidneys then is excreted in the urine.

Synthesis of 1 mol urea:

2 NH3 + CO2 + 3 ATP + 3 H2O → urea + 2 ADP + AMP + 4 Pi

The excretion of urea is very energy-intensive process, but the ammonia, which is a cytotoxin, has to be removed.

There is a relationship between the urea cycle and the citric acid cycle. The fumarate produced from argininosuccinate in the urea cycle can enter the citric acid cycle.

The aspartic acid originated from oxaloacetic acid as the starting compound of the citric acid cycle ulinks to the citrulline in the urea cycle to form argininosuccinate.



METABOLISM12.9. ábra - Figure 50: Synthesis of carbamide

Mammals can excrete nitrogen as ammonium ion, too.

In the liver, ammonia is ulinked to the glutamic acid with the help of glutamate synthase to form glutamine. This reaction requires energy.

glutamic acid + NH3 + ATP → glutamin + ADP + Pi

In the kidney, glutamine is cleaved hydrolitically by glutaminase to form ammonia, which is excreted in the urine.

glutamine + H2O → glutamic acid + NH3

4.5.2. 12.4.5.2. Nitrogen excretion of birds and reptiles. Synthesis of uric acid

Birds and reptiles lack the arginase enzyme, so they are not able to synthesize urea, they excrete nitrogen as uric acid.

The nitrogen arising in the process of deamination attaches to glutamate and gets into the liver with the blood stream. The glutamine will be the one of the precursors of the purin-based uric acid in the liver. The synthesis of purine skeleton is the sum of complex processes. Inosinic acid formed during reactions is a nucleotide containing purine skeleton, which is considered as a direct precursor of uric acid.

The synthesis of uric acid is shown in the following equation:

2 glutamic acid + 2 formic acid + CO2 + aspartic acid + glycine + 6 ATP→

→ inosine + 2 glutamic acid + fumaric acid + 5 ADP + AMP + 5Pi + PPi

The organization uses 6 mol ATP to the synthesis of 1 mol uric acid, in which there are 4 mol N. For the excretion of 1 mol ammonia (N) as carbamide or as uric acid 1,5 mol ATP is required.

4.6. 12.4.6. Disturbances of amino acid metabolismThe amino acid metabolism is regulated similarly to the other metabolic processes. The inherited metabolic diseases might be caused by the deficiency or pathological overfunctioning of an enzyme. Enormous number of (~ 80) hereditary anomalies are known by the researchers. One of the best-known amino acid metabolic disorder is the phenylketonuria (PKU). Phenylketonuria is an autosomal recessive trait characterized by a mutation in the gene of the phenylalanine hydroxylase. Those people whose phenylalanine-4-monooxygenase enzyme is missing or works incomplete, are not able to break down the phenylalanine. Due to a lack of this enzyme phenylalanine can not transform into tyrosine, thus catecholamines are not formed in further changes. The catecholamines are necessary for the development of nervous system. The phenylalanine is enriched in tissues, than goes through the transformation processes, which anomalous products (phenyl pyruvate, phenyl-lactate,



METABOLISMphenyl acetate) are excreted in high dose in the urine.

The operating condition of the phenyl-hydrolase enzyme is the tetrahydrobiopterin, which is formed in the process catalyzed by dihidrobiopterin reductase. In the absence of this enzyme the phenylketonuria can also develop.

The weight of brain of patients with phenylketonuria is small, their lifetime is short (max.: 20 years). Treatment of phenylketonuria includes the elimination of phenylalanine from the diet.


13. fejezet - 13. OTHER BIOCHEMICAL PATHWAYS1. 13.1. The biochemical bases of the function of skeletal muscleThe energy required for the functioning of muscles is obtained from biochemical processes. During muscle contraction the glycogen concentration of muscle decreases and the lactate and phosphate contents of venous blood flowing from muscle rises. For contraction, the muscle obtains energy not directly from the decomposition of glycogen, but from ATP.

From the energetics viewpoint, important part processes taking place in the muscle are the following:

• By the hydrolysis of ATP, ADP and Pi are formed. This supplies energy for the contraction.

• The ADP obtains energy from the creatine phosphate for the resynthesis.

KrP + ADP → Kr + ATP

From the breakdown of 1 mol of creatine phosphate at most half mole of ATP can be resynthesized.

• For the resynthesis of creatine phosphate and ATP the energy is supplied by the breakdown -in anaerobic conditions- of glycogen.

• For the resynthesis of glycogen the oxidation of pyruvic acid -aerobic- supplies the energy.

The majority of lactic acid originated from the breakdown of glycogen transform back into glycogen in the Cori cycle. In the muscle beside the oxidation of carbohydrate, fat-burning is also going on.

The muscle contraction is a series of reactions, which take place between actomyosin, ATP and certain ions. During the contraction of the muscle calcium ions are released. During relaxation of muscle chelate-like compounds are released, which bind calcium ions to form complexes.

2. 13. 2. Factors influencing the quantity and quality of the urineThe healthy kidneys are able to dilute or concentrate the urine in a wide range. The role of kidneys is to keep the proper balance of salts, acids and water and to ensure the osmotic potential in the body. The quality and quantity of urine is determined by three factors, the filtration, reabsorption and secretion.

The composition of ultrafiltrate formed in glomeruli is practically identical to the composition of blood plasma, but does not contain a higher molecular weight protein. Its daily volume is 180 litres. Two kinds of processes take place in the tubules, reabsorption and secretion. Reabsorption can be made by active and passive transport. Glucose, phosphate ion, amino acids, bicarbonates, uric acid, sodium-, potassium- and chloride ions are reabsorbed by active transport. Urea is reabsorbed by passive transport (from higher concentration to lower one). Secretion is a result of active cell work.

In case of thirst, the decreasing uptake of high amount of liquids increases the water retraction of kidneys. As a result, the quantity of the blood moving on the blood circulation hardly changes. The concentration of urine is also influenced by hormonal factors. The hormonal effect upon the adiuretin (ADH) is responsible. The adiuretic hormone (ADH), produced by hypothalamus and stored in the posterior lobe of hypophysis, is also responsible for hormonal effect. With increasing water intake, the blood plasma is diluted, the osmotic concentration decreases, thus the production of ADH also decreases. The urine becomes more dilute. In the thirsty body the plasma thickens, the production of ADH increases. In the distal tubules ADH increases the reabsorption of water, thus excreted urine becomes concentrated. The epithelium of tubules of kidneys is impermeable to water


13. OTHER BIOCHEMICAL PATHWAYS

molecules. ADH makes the epithelium more permeable to water.

Kidneys play important role in the regulation of blood volume, extracellular volume and acid-base balance. In the regulation of blood volume, the ADH is involved as previously described. When protein concentration of the plasma decreases, the osmotic pressure of blood also decreases, this is perceived by osmoreceptors and thus enhanced ADH secretion is induced.

The reabsorption of sodium ion is directed by hormone aldosterone. If the sodium concentration decreases in the extracellular space, the osmotic potential decreases. The water flows into the intracellular space. The increase of sodium ion concentration in cells enhances the water flow from intracellular space into extracellular space.

The amount of aldosterone is set by the renin-angiotensin system. Aldosterone in the kidneys increases the reabsorption of sodium ions and enhances the excretion of potassium ions.

Kidneys play role in the regulation of pH and composition of body fluids. The blood pH varies between narrow limits: from 7.35 to 7.45. In maintaining of this value, buffer systems in fluid spaces, the respiratory centre and the kidneys are also involved. Such buffer systems are protein buffers and bicarbonate buffers. The hemoglobin in the context of carbon dioxide transport is able to neutralize a significant amount of acid, thus is able to influence the pH. The phosphate buffer is significant in tubules of kidneys and in the intracellular space. Respiratory centre located in the medulla is able to detect the blood pH, thus in the case of acidification respiration increases, while with increasing of pH respiration decreases it.

Kidneys excrete acidic or alkaline urine depending on the pH value of blood. Urine pH can vary from 4.5 to 8.5. The plant food alkalizes the urine. In proximal tubule cells carbonic acid is formed from carbon dioxide by the enzyme carbonic anhydrase, which immediately dissociate to bicarbonate and hydrogen ion. The hydrogen ion gets into the lumen of the tubule, where forms carbonic acid with bicarbonate, and after that carbonic acid dissociates to carbon dioxide and water.

Water excretes in the urine, carbon dioxide diffuses back to the tubule cells. In the distal tubule cells, ammonia is produced from glutamine with cleaving of amino group by glutaminase. The ammonia diffuses to the lumen and there it is united with hydrogen ion to form ammonium and is excreted in the urine.

The biochemical mechanism of excretion of hydrogen-ion and potassium-ion is common. The excretion of K+

and Na+ through kidneys is related to the concentration the hormone aldosterone. The excretion of Ca2+ and HPO4

2- depends on the hormone production of parathyroid glands.

3. 13. 3. The gastric juice and its separationThe pH of gastric juice is 0.9-1.5. Gastric juice contains three types of proteolytic enzyme, the pepsin (the pH optimum is between 1.5 - 2), the rennin or chymosin (the pH optimum is 5.35) and cathepsin (the pH optimum is between 3.0-5.0). It contains trace amounts of fat-degrading enzyme, lipase and also contains mucin.

The pepsin is released by the chief cells of stomach as an inactive form zymogen, pepsinogen. Hydrochloric acid, which is released from parietal cells in the stomach lining activates pepsinogen to pepsin. Chymosin converts caseinogen (milk protein) into insoluble casein in the presence of calcium ions.

3.1. 13. 3. 1. The mechanism of the hydrochloric acid production of the stomachThe mechanism of hydrochloric acid production by parietal cells is still not fully understood. The parietal cells are able to concentrate the hydrogen ions of the blood. When acid secretion is stimulated the outflow of hydrogen-carbonate into venous blood leaving from a stomach lining results in an elevation. The activity of carbonic anhydrase of the parietal cells increases.

The essence of Davenport theory of gastric acid production is that parietal cells decompose water contained therein to hydrogen and hydroxide ions.

The carbonic anhydrase enzyme catalyses the reaction between carbon dioxide and hydroxide ions originated from water to form bicarbonate ions.



The hydrogen ions are secreted actively by using ATP from the cytoplasm of parietal cells and mixed in the canaliculi.

The energy obtained may be derived from the oxidative degradation of glucose. The bicarbonates get into the blood, but chemically equivalent amount of chloride ions get from the blood plasma to the cell. The migration of chloride ion (so far) is passive, requiring no power process. The central role of carbon anhydrase is supported by the fact that substances, which retard the activity of the enzyme also inhibit the formation of hydrochloric acid (Fig. 51).

13.1. ábra - Figure 51: Steps of gastric acid secretio

In terms of the secretion of gastric juice, two states can be distinguished, the basal phase and the digestive phase. In the basal phase variable amount of hydrochloric acid- and pepsin are being secreted. This is called basal secretion. In the digestive phase, the gastric juice production adapts to the quality of the food. This is ensured by a neuro-humoral regulation.

An example is that if you eat meat, large amount of gastric juice is produced, wherein the amount of hydrochloric acid is high, while the amount of pepsin is medium. If we eat bread, moderate amounts of gastric juice is produced, which contains little hydrochloric acid and a lot of pepsin.

4. 13. 4. The control of metabolic processes4.1. 13. 4. 1. The control of lipid metabolismLipid metabolism is in strong connection with glucose catabolism, it depends on the carbohydrate supply of the body. The liver plays a major regulating role in the process of metabolism. Fats are the only nutrient forms that can avoid the controlling function of the liver, because they can get into the lymphatic system and can get into the adipose tissue directly.

The mobilisation of the stored fat from the adipose tissues is under hormonal regulation. Adrenaline, glucagon and ACTH ulink to the membrane of the fat tissue. This initiates the activation of the adenylate cyclase system. The intracellular lipase is activated by adenylate cyclase-cAMP system. This enzyme catalyzes the hydrolysis of triglycerides stored in fatty tissue. The reaction products get into the site of application via circulation. Fatty acids are transported by serum albumin in the circulation.

A large amount of acetyl groups originated from the β-oxidation of fatty acids decomposes to carbon dioxide and water in the citric acid cycle and in the terminal oxidation. This process is in control of respiration that works in the citric acid cycle and in terminal oxidation.

If there is large amount of ATP and small amount of ADP in the system the resulting energy is not utilized in



anabolic processes by the body. Then the high concentration of ATP inhibits citrate synthetase, isocitrate dehydrogenase and α-ketoglutaric acid dehydrogenase enzymes allosterically. The activity of these enzymes decreases, the citric acid cycle slows down.

If a large amount of food gets into the organism the cell glucose concentration increases and its degradation becomes faster. The quantity of acetyl-CoA generated through pyruvic acid increases. The resulting acetyl-CoA concentration can become so large that it exceeds the absorption capacity of citric acid. This has an inhibitory effect on the pyruvate dehydrogenase enzyme, thus a part of acetyl-CoA gets into the cytoplasm from the mitochondria as citric acid. The fatty acid synthesis requires large amount of acetyl-CoA and NADPH + H +. NADPH + H+ are only formed in a few enzyme catalyzed processes. These are malate, isocitrate dehydrogenase, glucose-6-phosphate dehydrogenase enzyme-catalyzed reactions. The lack of NADPH + H+ can influence the fatty acid synthesis.

The fatty acid synthesis can be influenced by the formation of malonyl-CoA. In the process, the carboxylation of acetyl-CoA is catalyzed by acetyl-CoA carboxylase enzyme. This enzyme with acyl-ACP synthetase that is the central enzyme in the synthesis is allosterically regulated.

These enzymes are activated by citric acid and isocitric acid in large amount, while they are inhibited by acyl-CoA derivatives.

4.2. 13. 4. 2. The function of adenylate cyclase - cAMP systemHormones influence the pathways of biochemical processes catalyzed by enzymes. They exert their effects in two ways.

1. They regulate gene activity. They interact with repressor molecules that inhibit the function of DNA in the nucleus, thus the synthesis of enzyme proteins is started.

2. They activate adenylate cyclase system. In this case, they connect to the cell membrane and induce the synthesis of secondary messenger compounds that activate the enzymes.

In both cases, they act by enzymes; they change the enzyme concentration or activity.

1. During gene activation, steroid hormones initiate enzyme protein synthesis in the nucleus of target organ cells. The non-polar steroid hormones are able to pass through the target cell membrane. In the cytosol they are bound to a steroid-specific receptor.

The formed hormone-receptor complex is able to enter the nucleus through the nucleus membrane. In the nucleus the DNA chains that presented as chromosomes are surrounded by proteins. These repressor proteins prevent the binding of DNA dependent RNA polymerase and thus they also prevent the mRNA synthesis as DNA dependent RNA polymerase initiates the mRNA synthesis.

To construct a particular protein (e.g. an enzyme that catalyzes a biochemical process) the hormone transforms to a hormone receptor complex in the cytosol of the cell, enters the nucleus and there cleaves repressor protein from the DNA molecule containing the code for the protein synthesis (at DNA stage that starts mRNA transcription). The RNA polymerase is then able to bind to the chain. The mRNA synthesis can start. The hormone can only start up the transcription of DNA to mRNA, the RNA polymerase activation in such a form that binds to the receptor. The outcome of hormonal effect is the messenger RNA transcription that is necessary for protein synthesis. Rewriting of information carried by the mRNA into protein takes place in the ribosomes.

2. The hormone attaches irreversibly to the hormone-receptor proteins localized in the membrane of the target cell. As a result, the configuration of the receptor changes, thus it is able to bind the protein type carrier molecules on the inside surface of cell membrane (G protein). As a result of connection, G-protein undergoes conformational changes. Due to the altered conformation, it can connect to enzymes that catalyze synthesis of secondary messengers.

The adenosine 3’, 5’ cyclic-monophosphate (cAMP) is a secondary messenger that is formed by adrenaline hormone. The cAMP is synthesised from ATP by the enzyme adenylate-cyclase with a loss of pyrophosphate.

The cAMP is also called secondary messenger.



The main role of cAMP is the activation of protein kinases. It also influences membrane transport processes.

Protein kinases activate other proteins by chemically adding phosphate groups to them.

By cleaving macroergic phosphate groups, protein kinases attach phosphate groups to proteins that become active.

The cAMP provides connection between hormones that transfer stimulus as chemical information and enzymes that catalyze the metabolic processes directly.

After the termination of the hormonal effects, the unnecessary cAMP molecules hydrolyse to AMP by the enzyme, phosphodiesterase.

Phosphodiesterase enzyme is inhibited by methylated xanthines, caffeine, theobromine thus the effect of the hormones is elongated in the presence of them.

The cAMP influences the permeability of cell membranes and the transport processes.

4.2.1. 13.4. 2. 1. The presentation of adenylate-cyclase system operation through the mobilization of glycogen

Adrenaline is able to mobilize glucose from the stored glycogen quickly.

The effect of this hormone prevails through a mediator, by increasing the activity of the phosphorylase. The liver cells respond to the signal of adrenaline and glucagon, while the muscle cells respond to adrenaline hormone. Adenilate-cyclase connected to the membrane of the cell is activated. One part of ATP transforms to cAMP in the cell as an effect of protein kinase. Activation occurs allosterically.

Protein kinase activates phosphorylase kinase by using ATP. Phosphorylase kinase is sensitive to the concentration of Ca2+ ions as well. This is another controlling factor in the muscle. Muscle contraction is started up by Ca2+ ions released as an effect of nerve impulses.

In addition to contraction, they also affect the operation of the phosphorylase kinase.

The active phosphorylase kinase converts the inactive phosphorylase-B to active phosphorylase-A. Two phosphorylase B molecules (dimer) form a tetramer in the muscle and become active in this way. In the liver, both of them are in dimeric form (Fig. 52).

13.2. ábra - Figure 52: cAMP system

The reduction of activity is carried out by phosphorylase phosphatase; it transforms the active form A into inactive form B. The synthesis of glycogen is controlled by a system that is similar to the previous one, but it



has an opposite effect.

4.2.2. 13. 4. 2. 2. Hormone control of carbohydrate metabolism

Complex carbohydrates are broken down in the process of digestion to monosaccharides that transform to glucose. In the small intestine glucose is absorbed and get into the bloodstream. Glucose in the blood is called blood sugar.

Glucose molecules get into the tissues via bloodstream, where they become the main energy supply carrier of the cells. Some tissues, such as brain tissue do not have oxidizable carbohydrate reserves, thus they are reliant on blood glucose very much.

Carbohydrates from food sources and the body need to determine what kind of processes the glucose steps into.

The conversion speed of carbohydrates in the living body is high although mammals have constant blood sugar level. In humans it ranges from 4.0 to 5.5 mmol / l. If the body wants to utilize blood sugar as energy, it decomposes it in the cells and depending on the circumstances, it transforms to lactic acid or pyruvic acid. Excess glucose is stored as glycogen in the liver and muscles, which become available later. The endocrine system keeps blood glucose level at a constant level. The glucose originated from foods, digested and absorbed from the gut can get into the blood and can cause increased blood sugar level. This compound might also come from the breakdown of glycogen stored in the liver. Sugar generated from the breakdown of amino acids, lactic acid and oxaloacetic acid by gluconeogenesis also may increase the blood sugar level.

The glucose concentration of blood can be reduced by the sugar utilization and oxidation of muscle and other tissues. The blood sugar level is also reduced when glycogen is synthesized in the liver and muscle. The formation and the storage of fat, produced from glucose - which takes place in the liver and in adipose tissues - can also reduce the blood sugar level.

The regulation of the processes mentioned above is performed by several hormones collectively (Fig. 53). Insulin has central role in the regulation, which reduces the blood sugar level. Insulin promotes the uptake of glucose by most cells and also helps the breakdown of it. Insulin enhances the formation and storage of glycogen in the muscles. This hormone increases the synthesis of fats in the liver, inhibits the breakdown of glycogen in the liver to glucose and also inhibits the release of glycogen into the bloodstream.

Glucagon is secreted by the pancreas effects on the process of raising the blood sugar level. This hormone enhances the breakdown of glycogen in the liver and helps the release of glucose into the blood stream. Glucagon enhances the resynthesis of glucose from lactic acid in the liver.

Adrenalin stimulates the increase of blood sugar level, because it enhances the breakdown of glycogen into glucose in the liver. Adrenalin promotes the breakdown of muscle glycogen into lactic acid. The resulting lactic acid may be a precursor of resynthesis of glucose.

Cortisol raises the blood glucose level, because it increases the glucose resynthesis from amino acids in the liver. Cortisol enhances the amount of required amino acids, because this hormone inhibits protein synthesis, increases the breakdown of proteins in the muscles.

Somatotropin (growth hormone) decreases the blood glucose levels, because enhances the synthesis of proteins. It inhibits the conversion of amino acids to glucose.

Gluconeogenesis is controlled by adenohypophysis by its hormonal activity (ACTH, STH) in the body. Thyroxine, LTH (luteotropin) and androgens can also affect the carbohydrate balance.

13.3. ábra - Figure 53: The outline of the neourohormonal control of the carbohydrate metabolism and the blood-sugar level



Hypoglycemia (decreased blood glucose level) is an abnormally diminished content of glucose in the blood. The cause of hypoglycemia might be the damage of a central nervous system. Hypoglycemia can cause unconsciousness, which might be irreversible. The hypoglycemia induces sympato-adrenal symptomes.

Hyperglycemia is a condition in which an excessive amount of glucose circulates in the blood. Because of the excitement of the sympato-adrenal system, the glucose and lactic acid content of blood inrease. This occurs due to mobilization of compounds from depots and due to enhance of gluconeogenesis. The sucking capacity of kidneys decreases, glucose is secreted in the urine. Thus, a biologically important compound is lost.

The glucose transport between organs and its hormonal regulation are shown in 54 Figure.

13.4. ábra - Figure 54: The glucose transport between organs and its hormonal regulation

5. 13. 5. The role of liver in the intermediate metabolismThe liver plays a central role in the intermediate metabolism like gluconeogenesis and glycogen metabolism. Liver is the scene of anabolism, catabolism and storage of glycogen. The liver cells are only able to convert galactose to glucose. Like other cells, liver cells are also able to isomerize fructose into glucose. The lactic acid, glycerol, oxaloacetic acid and other intermediates also are converted into glucose in the liver. The gluconeogenesis or conversion of amino acids to glucose takes place in the liver.

Liver plays a role in the lipid metabolism:



• Ketone bodies are produced here from large amounts of acetyl-CoAs originated from degradation of fatty acids.

• The synthesis of plasma-phosphatides takes place in the liver. Plasma-phosphatides are important during transports of lipids from the liver to the plasma.

• Cholesterol is also formed in the liver.

• Bile acids formation takes place in the liver. Bile acids react with taurine and glycine here.

• The storage of carotene takes place here. Carotene is converted to vitamin A in the liver.

The liver plays crucial roles in nitrogen metabolism, too. It has central role in the metabolism of amino acids and in the maintenance of dynamic balance of proteins. The synthesis of urea and trans-amination, deamination processes of amino acids also take place in the liver. The trans-amination and deamination processes create new amino acids. Without these reactions, the gluconeogenesis does not occur and ketone bodies are not formed.

Without the work of the liver amino acids accumulate and are excreted in the urine. The formation of choline and creatine are also take place here. Some specific plasma proteins, albumins, globulins, prothrombin, fibrinogen and the majority of coagulation factors are also synthesized in the liver. Because of this liver is important organ of blood clotting and defense mechanism of the body. The majority of uric acid also forms in the liver.

Liver has important role in the porphyrin metabolism:

• The breakdown of hemoglobin to bilirubin is localized in the liver. (This process takes place in the spleen and in the bone-marrow, too).

• The excretion of bilirubin esterified with glucuronic acid occurs in the bile into the intestine. The esterification reaction takes place in the liver.

• Liver is also a detoxification organ.


14. fejezet - 14. BIOCHEMICAL PATHWAYS IN THE FOOD INDUSTRYDuring the storage and treatment of food raw materials produced in large quantities by food processing industries for human and animal nutrition, biochemical processes may take place which might be favourable and unfavourable.

The biochemical processes can be or must be influenced, enhanced or reduced by changing of the circumstances.

One of the basic processes is the fermentation, which in the narrow sense refers to anaerobic catabolism of carbohydrates. The end product of this process is lactic acid or alcohol and carbon dioxide.

The fermentation in the food technology is a common pathway, which is necessary in order to obtain the desired quality of product. The processes that take place spontaneously can adversely affect quality, therefore it is essential to avoid them.

1. 14. 1. The application of the fermentation in the food industryThe fermentation is carried out by enzymes produced by microbes, or fermentation is occurred by exposed tissue enzymes. During the fermentation flavours may also develop. This is also true for yeast, lactic acid fermentation. The hetero-fermentative lactic acid bacteria also produce more flavour components. The resulting acetic acid and acetaldehyde are major flavour components of dairy products. Yeast produces mainly ethanol as the end product of the fermentation, but they are also able to convert amino acids by trans-amination and decarboxylation reactions. In these processes alcohols, aldehydes, acids and esters are formed.

The cocoa beans, coffee before roasting, the tobacco and the tea are fermented before further transformation. During tea fermentation tissue enzymes work. The fermentation of yellow tea is shorter than black tea’s one. In the case of green tea, there are not fermentation processes.

The lactic acid fermentation is known as microbiological preservation procedure for centuries.

Acidification of cabbage, cucumber happens with lactic acid fermentation. In this process, lactic acid bacteria produce lactic acid from the sugar content of raw material used to preserve. At the 0,7 to 1% lactic acid content the activity of lactic acid bacteria and other microbes ceases, so the product can be stored for a long time.

At the production of alcoholic beverages (beer, wine, brandy) fermentation processes can take place. At the alcoholic fermentation various yeast strains are used, which convert the fermentable sugars of liquid into alcohol.

The reaction sequence of the process is: hexose → pyruvic acid → acetaldehyde → ethanol.

The formation of pyruvic acid to acetaldehyde is conducted by yeast during anaerob metabolism. This process is catalyzed by pyruvic acid decarboxylase. Then acetaldehyde is reduced to ethanol by the NAD-specific alcohol dehydrogenase enzyme.

The alcohol content of wines and spirits is originated from the sugar content of fruit juice of ripe grapes, fruits.

Beer is brewed from barley. Barley contains starch that has to be broken down to simply sugars before the yeast can make alcohol. Therefore malting step is needed. Malting is the process in which inactivated enzymes become active again and degrade starch into fermentable sugars.

2. 14. 2. The biochemical processes of cereals germination


14. BIOCHEMICAL PATHWAYS IN THE FOOD INDUSTRY

Germination replaces the resting stage that is fixed duration and is due to external or internal circumstances. In seed germination, characteristic and fast reactions take place, plant hormones work intensively.

The enzyme activity increases, so the breakdown and usage of reserved nutrients are started. The changes concern the seed coat, the nourishing tissues and the germ as well. There are active energy and material circles between them.

In the first stage of germination, the seed swells greatly and germ reactivation occurs. In order to start the process appropriate temperature, humidity and oxygen are required. At this stage macromolecules, cell organelles and phytohormones are reactivated. Due to the swelling and water absorption, the dormant enzymes are also activated. The biochemical processes start. The oxidation processes of free sugars, amino acids occur. The protein synthesis starts as well.

In the second stage of the germination, which clearly can not be separated from the previous stage, the mobilization of reserved nutrients occurs. With the help of activated enzymes mobilization and hydrolysis of starch, storage proteins, lipids, phosphates take place. For faster processes a large amount of hydrolase enzyme is needed. At this stage, the system is able to synthesize this enzyme. The stored energy in the form of lipids is converted into monosaccharides.

3. 14. 3. Respiration during storageDuring the storage of foods, the cells respire, so quality may change. The breathing process mainly refers to the breakdown of carbohydrates to carbon dioxide and water. In aspect of food industry, the carbohydrates decomposed during respiration counts losses. There is an important requirement during food processing and - storage to minimize the breathing losses. Many factors affect the intensity of respiration, such as external conditions (temperature, humidity, atmosphere composition), the varieties and agrotechnical factors. The oxygen concentration influences the carbohydrate degradation speed as well beside the direction of the process.

3.1. 14. 3. 1. Respiration of grain during storage Oligo- and monosaccharides with low molecular weights required for respiration are found in grain in small amount, or originated from degradation of reserved polysaccharides. In the dormant state of grain the β-amylases are the most active amylolytic enzymes, they produce maltose. There can also be found other sources (glucose, fructose, maltose, galactose, sucrose, raffinose) in the grain for respiratory processes, and their supply may also come from degradation of several reserved compounds.

The respiration rate of grains is mainly affected by the moisture content, temperature, oxygenation and the lives of the seeds.

The increase of the moisture content of germ enhances the respiration, the function of glutamic acid decarboxylase increases. The quantity of microbes also increases.

With the increasing of the temperature, the velocity of the respiration enhances. Its effect is influenced by the oxygen supply and the concentration of CO2. The anaerobic fermentation may occur under a particular oxygen concentration level.

The production, ripening, harvesting and drying processes of grains might be called “the life of the grain”, collectively.

Overall, we can say that the fuller ripening, the storage on the dry place reduces the velocity of the breathing.

3.2. 14.3.2. The respiration of fruits and vegetablesFruits and vegetables like other foodstuffs of plant origin respire during storage. There might be considerable differences between the species (even 50%) in the respiration intensity. The breathing intensity changes during the growth of the fruit, too. Increased respiration intensity decreases initially then approaching the ripening stage the respiration rises again.

In the case of vegetables such phenomena can not be detected. The respiratory intensity of vegetables grows steadily and in the end of storage is stronger than in the beginning of the picking and storage. Different tissues



have different respiration. The respiratory intensity of dermal tissues is much greater than vascular and ground tissues. This can be explained by improved oxygen supply and greater enzyme activity (oxido-reductase). The increase of temperature both in case of fruits and vegetables can also improve the breathing.

The temperature of storage can influence the biochemical processes in varying degrees, as a result we get the right quality stored product. The optimum storage temperature depends on the species, variety and the maturity.

The potatoes become sweet stored at low temperature, its pleasure value decreases, and the non-enzymatic browning increases. During storage the amount of mono- and oligosaccharides, especially sucrose greatly increases. The formation of mono- and oligosaccharides and change of respiration rate are different depending on the temperature.

The increase of sugar content depends on the balance of the next biochemical processes:

• breakdown of starch into mono- and oligosaccharides,

• resynthesis of starch from mono- and oligosaccharides,

• formation of sucrose from monosaccharides,

• respiratory process.

The ability of the resynthesis decreases with the time of the storage and decreasing of the temperature mostly. Thus increases the concentration of mono- and oligosaccharides.

The oxygen and carbon dioxide tension of containers significantly affects the intensity of respiration. The decrease in oxygen and increase in carbon dioxide reduces the respiration. The decrease of oxygen concentration below the critical level favours the anaerobic breakdown of sugar in the case of fruits and vegetables, which can lead to deterioration in quality.

3.3. 14. 3. 3. The ripening of fruitsThe ripening is the process by which a variety of fruit reaches the proper size and color, develops the taste, flavor and aroma. During ripening the fruit becomes suitable for consumption or industrial processing.

During ripening the ground color of fruit peel becomes yellow from green, the cover color becomes red or blue from yellow.

The hardness of flesh can also change during this process. The ripe fruit is generally flexible, soft. Most of the unripe fruits are sour, bitter, while ripe fruits are sweet or slightly sour. The ripe fruit has a noticeable characteristic flavor as well.

Among the fruits, there are some, which are able to continue ripening after being picked. These fruits depending on their ripening stage will ripe within longer or shorter period. Its condition is that a certain degree of maturity of fruit must have attained before its removal from the plant.

Such fruits are apple, pear, quince, apricot, peaches and strawberries. During the ripening, process the metabolism of the fruit changes. Many enzyme activity increases. The starch is converted into sugars, acids are converted to pyruvic acid then to carbon dioxide and water. The fruit becomes sweeter and slightly sour. The cellulose and protopectin content of fruit decrease and proportionally the amount of dissolved pectin increases. The protein content increases as well.

The chlorophyll begins to decompose, but formation of carotenoids, anthocyanins and flavourings start. These processes are followed by the change of permeability of cells and cell organelles. In case of most fruits during the ripening the respiration and CO2 production increases, that can only slow down at the aging stage.

The fruits can be classified based on their intensity of respiration at the ripening stage. As citrus fruit ripens CO2 production reduces. The bananas and tomatoes are characterized by a gradual increase in CO2 production and the mature state occurs after the maximum CO2 production.

In the case of third group (peach and strawberries) the CO2 production increases, but the mature state of fruit occurs at the maximum CO2 production.



Ripening agent, ethylene produced in the fruit speeds up the ripening process.

In foods during their storage and processing, often brown pigments, flavours are developed. Brown pigments may be developed through either enzymatic (enzymatic browning) or non-enzymatic processes (non-enzymatic browning (NEB)).

The non-enzymatic browning (NEB) means that the interaction between free amino group and reducing sugar, under certain circumstances, causes the browning. In that process aroma compounds and brown pigments, melanoidins are produced. The reaction is promoted if the mole proportion of the sugar and amino-compounds is 3:1, the temperature is high and the pH is slightly acidic, neutral or alkaline (pH 3-11). In order to run the reaction at least 10% water content is required.

Enzymatic browning

In this process, melanins are formed by the certain enzymes of body tissue. The melanins may be also a normal life functioning products: pigments of hair, fur, feathers. Often, due to injuries like peeling or pressing of fruits, vegetables are formed these pigments.

In the damaged cells, due to the damage of endoplasmic network substrates come into contact with enzymes being not in contact before.

The enzymes are called polyphenol oxidases (PPO). These enzymes contain Cu 2+ ions, which are activated by reduction of the copper. Enzymes can bind molecular oxygen and can oxidize monophenols to O-diphenols. In the meantime, the copper is reconverted to Cu2+ and becomes inactive again. With the oxidation of O-diphenol to quinone the enzyme is activated again. The enzyme has a phenol hydroxylase and polyphenol oxidase activity.

The substrates may be tyrosine, catechins, flavones, cinnamic acid, leukoantocianidins, etc. The enzymatic browning can also be beneficial at the fermentation process of tea and cocoa. In adverse cases, the process may be deferred and inhibited. In such cases the inactivation or inhibition of enzymes to be solved. Inactivation most easily can be done by heating. The process can be delayed by the exclusion of the air or by application of reducing agent.

The reducing of browning tendency can be achieved by such variety of plant cultivation, in which the enzyme activity is low or the substrate concentration is low.

4. 14. 4. The biochemistry of meat ripeningThe main function of muscle tissue in the living animal is to ensure the mechanical work. To do this, a large amount of energy is required, so it is important to ensure adequate energy supply in the muscle.

Direct source of energy is the ATP, which is derived from the breakdown of carbohydrates. In the case of large and sustained muscle work in the energy supply the role of lipids are also appreciated. The muscles contain large amounts of creatine phosphate, which is involved in the ATP supply with the help of creatine kinase.

After the animal has been slaughtered, the conditions of the biochemical processes are changed in the muscular tissue. The connection of muscular tissue with the liver breaks up, and there will be no nutrient supply any longer. The muscle does not receive fresh blood, so that the oxygen-supply breaks off. The process has taken place in the changed circumstances is called meat ripening.

The final product, which undergoes chemical changes between changed circumstances, is called meat.

In practical terms, the post-mortem transformations can be divided into three phases. The first stage is the stage before rigor mortis. The meat is soft yet, the quantity of the ATP and the creatine phosphate decrease, the anaerobic processes start.

The second stage is the stage of the rigor mortis. The glycogen in the muscle is converted into lactic acid causing a fall in pH, muscles become stiff or rigid one known as muscle rigor, proteins are denatured.

The third stage is the stage of the post rigor mortis. The meat becomes tender again. In secondary processes, the flavour and organoleptic characteristics improve (Fig. 55).



14.1. ábra - Figure 55: The biochemistry of meat ripening

After slaughtering of animal the oxygen supply of muscle tissues stops. Due to the lack of oxygen the function of mitochondrial electron transport chain is not possible. ATP is formed only in the anaerobic glycolysis, but its quantity is only a fraction of that produced under aerobic conditions. When the ATP decrease reaches a threshold that is no longer sufficient to inhibit actin-myosin connection, rigor mortis occurs.

In the absence of ATP the condition of muscle relaxation can not come to be. Lactic acid is reduced from pyruvic acid -under these circumstances- originated from glycolysis. Lactic acid is not able to get out of the meat, so it is accumulated. The acidic pH is shifted to neutral, the pH reaches 5.3 to 5.5 value. These biochemical processes lead to changes in the consistency and the water absorbing capacity of meat

The rigor mortis disappears after a while because some proteolytic enzymes, cathepsins operate under such conditions.

The membranes are damaged in the state of rigor mortis, the enzymes can get out and can expose their effects. The changes in consistency may be related to the ions redistribution within the protein system.

The meat ripening processes are influenced by those effects (pre-slaughter effects) which may influence the glycogen content of the muscle tissue. Such effect is the stress as well.

Maturation can be accelerated – it is often used in practice – with proteolytic enzyme preparations. In meat industry, it is used for beef because it has very hard consistency.

Earlier there were often used proteolytic enzymes with plant origin such as papain and phycin, but today microbial enzyme preparations are rather used.

After slaughtering the negative effects of cooling can be reduced by electrical stimulation. In this case, the shortening of muscle strength occurs (cold shortening phenomenon). If within one hour after slaughtering the meat is treated from 1.5 to 2 s time period at a voltage of 500-600V, the shortening can be reduced.

5. 14. 5. Changes of colour through the meat processingThe colour of meat indicates the freshness of meat for the customers. Today, the inclusion of additives give meat



prolonged intense red colour. These added compounds (KNO2-KNO3) prevent natural biochemical processes. The colour (intensity and brightness) of meat is based on the quantity of principal muscle pigment—myoglobin. Myoglobin has a protein portion, globin (96%) and a heme portion (4%). Original colour is crimson. The colour change of myoglobin is due to heme can bind different groups (ions). Each heme residue contains one central coordinately bound iron atom that is in the Fe2+ (ferrous) oxidation state. The Fe2+ is being chelated with six ligands. Heme is a square planar molecule containing four pyrrole groups, whose nitrogen form coordinate covalent bonds with four of the iron's six available positions.

One position is used to form a coordinate covalent bond with the side chain of a single histidine amino acid of the protein. The sixth bond is an unstable connection with water, which can be exchanged with other ion groups or ions. In living organisms, where the oxygen partial pressure is high, the sixth bond is a connection with oxygen. The resulting complex is called oxy-myoglobin that has bright red colour. If instead of oxygen, carbon monoxide is bound, it is called carboxy-myoglobin, and when nitrogen monoxide is bound, it is called nitroso-myoglobin. Both forms arise during smoking, pickling. Their colours similar to the oxy-myoglobin are red.

In products exposed to heat protein is denaturated, thus myochromogens are formed. If the partial pressure of oxygen is reduced, the oxidation number of iron is changed in the hem.

The Fe(II) is oxidized to Fe(III), hem is transformed to hemin and myoglobin is transformed to metmyoglobin. In this case, the sixth bond is not attached to oxygen, but is attached to hydroxide ion. The metmyoglobin has grayish red colour. The product denatured by heat is the metmyochromogen. The formation of the metmyoglobin is helped by heat, UV light and by decrease of pH.

Myoglobin products in which the oxidation number of iron is +2 are red. The distribution of myoglobin in meat is not equable. Its concentration and thus the intensity of the red colour depends on the species, sex, age and life style of the animal. The more myoglobin contains meat, the darker red the colour (Fig.56).

14.2. ábra - Figure 56: Changes of colour through the meat processing


15. fejezet - 15. RECOMMENDED REFERENCESWiliam H. Brown, Judith A. McClarin, Introduction to Organic and Biochemistry. 3rd edition Copyright 1981. by Willard Grant Press, 20 Providence Street, Boston, Massachusetts 02116. ISBN: 0-87150-738-2.

Christopher K. Mathews; K. E. van Holde, 1990. Biochemistry, The Benjamin/ Cummings Publishing Company, 390 Bridge Parkway, ISBN: 0-8053-5015-2.

Karla L. Roehrig, Carbohydrate Biochemistry and metabolism. Copyright 1984 by THE AVI PUBLISHING COMPANY, INC. Westport, Connecticut. ISBN: 0-87055-447-6.

James Darnell, Harvey Lodish, David Baltimore, Molecular Call Biology. Copyright 1986 by Scientific American Books, Inc. ISBN: 0-7167-6001-0.


16. fejezet - Questions1. How can the living organism be classified according to their type of metabolism?

2. How can carbohydrates be classified?

3. What does the reductive sugars expression mean?

4. What kind of biological functions do the polysaccharides have?

5. What are the similarities and differences between amilose and amilopectine?

6. What kind of bonds stabilise the spatial structure of proteins?

7. What are the biological functions of proteins?

8. What are the biological functions of phospholipids and neutral fats?

9. What are the similarities and differences between the primary and secondary functions of DNA and RNA?

10. What are the biological functions of nucleoside-triphosphates?

11. How can the vitamins expound their effects?

12. Which type of gland produces hormones and how can the hormones be classified by their chemical structure?

13. What does the fact mean that the hormonal regulation has hierarchical order (through an example)?

14. What are the similarities and the differences between the hormones and the tissue hormones, phytohormones?

15. What kind of parts do the enzymes consist of, what is the function mechanism of the enzymes, and what factors influence the function of the enzymes?

16. What is the essence of photosynthesis, what are the stages of photosynthesis?

17. What does the expression of photo-phosphorylation mean?

18. What are the biochemical processes of the catabolism of glucose and what are the places of these reactions in the cell?

19. Where is it produced and how much energy is recovered from the entire oxidative catabolism of one mole of glucose?

20. What is the importance of pentose phosphate cycle?

21. What kind of fermentation processes do you know?

22. What is the similarity and the difference between the fermentation processes taking place in the silo and the first stomach?

23. What is the essence of gluconeogenesis, where and why does this process take place?

24. Describe thesteps of glycogen mobilization (Cory cycle)!

25. What substances and what kind of enzyme complex are necessary to the biosynthesis of fatty acids?

26. Describe the steps of fatty acid synthesis!

27. How does the catabolism of fat begin and what enzymes catalyze this process?


Questions

28. Describe the steps of catabolism of palmitic acid .

29. How many ATPs are formed in the entire oxidative catabolism of one mole of palmitic acid?

30. What is the process of ketogenesis?

31. Where is it formed and what is the role of cholesterol in the human body?

32. What is the role of glyoxylate cycle?

33. What is the difference between glyoxylate cycle and citric acid cycle?

34. How can plants take up nitrogen, and how can they transform it into organic compounds?

35. Describesome pathways of essential aminosynthesis!

36. What is the transcription process of the protein synthesis?

37. What is the translation process of the protein synthesis?

38. How can you determine the quality of proteins?

39. What kind of protein-digesting enzymes do you know?

40. Where do these enzymes work and what circumstances influence their functions?

41. What are the biogenic amines and what is their significance?

42. Review the catabolism of carbon skeleton of amino acids in the tricarboxylic acid cycle!

43. Review the urea cycle!

44. How can birds and reptiles excrete the nitrogen?

45. What kind of chemical processes do muscles obtain energy for their function from?

46. What kind of factors influence the quantity and quality of the urine?

47. Describe the hypotheticalmechanism of the formation of gastric juice.

48. What are the functions of adenylate cyclase-cAMP system?

49. What kind of hormones regulate the carbohydrate metabolism?

50. What are the roles of liver in the metabolism?

51. What is fermentation in food processing used for?

52. Review the biochemical processes going during ripening of fruits!

53. What does non-enzymatic browning mean?

54. What is enzymatic browning and how does it affect food products?

55. Review the biochemistry of meat ripening!


Questions


Documents

regi.tankonyvtar.hu · Web viewApplied biochemistry. Applied biochemistry. Tárgymutató. 1. INTRODUCTION. 2. THE LIVING SYSTEMS. 3. BIOMOLECULES I. CARBOHYDRATES. 4. BIOMOLECULES