FORMAL REPORTOF EXPERIMENT IN BIOCHEMISTRYLIPID 1 and LIPID 2

Written by group 4:

Ira Novita Sari(K3309049)Sarry Saraswaty(K3309073)Yoga Bririan Jati(K3309089)

Chemistry Education Study ProgramDepartment of Mathematics and Natural Sciences EducationFaculty of Teacher Training and EducationSebelas Maret UniversitySURAKARTA2012FORMAL REPORTOF EXPERIMENT IN BIOCHEMISTRY

I. TITLE: LIPID 1 and LIPID 2

II. PURPOSE: Canisolateseveraltypes oflipidsfromegg yolk Can identifying of lipis

III. BASIC THEORYLipids constitute a broad group of naturally occurring molecules that include fats, waxes, sterols, fat-soluble vitamins (such as vitamins A, D, E, and K), monoglycerides, diglycerides, triglycerides, phospholipids, and others. The main biological functions of lipids include energy storage, as structural components of cell membranes, and as important signaling molecules. Lipids may be broadly defined as hydrophobic or amphiphilic small molecules; the amphiphilic nature of some lipids allows them to form structures such as vesicles, liposomes, or membranes in an aqueous environment. Biological lipids originate entirely or in part from two distinct types of biochemical subunits or "building-blocks": ketoacyl and isoprene groups.[4] Using this approach, lipids may be divided into eight categories: fatty acids, glycerolipids, glycerophospholipids, sphingolipids, saccharolipids, and polyketides (derived from condensation of ketoacyl subunits); and sterol lipids and prenol lipids (derived from condensation of isoprene subunits). Although the term lipid is sometimes used as a synonym for fats, fats are a subgroup of lipids called triglycerides. Lipids also encompass molecules such as fatty acids and their derivatives (including tri-, di-, monoglycerides, and phospholipids), as well as other sterol-containing metabolites such as cholesterol. Although humans and other mammals use various biosynthetic pathways to both break down and synthesize lipids, some essential lipids cannot be made this way and must be obtained from the diet.Categories of lipidsFatty acidsFatty acids, or fatty acid residues when they form part of a lipid, are a diverse group of molecules synthesized by chain-elongation of an acetyl-CoA primer with malonyl-CoA or methylmalonyl-CoA groups in a process called fatty acid synthesis. They are made of a hydrocarbon chain that terminates with a carboxylic acid group; this arrangement confers the molecule with a polar, hydrophilic end, and a nonpolar, hydrophobic end that is insoluble in water. The fatty acid structure is one of the most fundamental categories of biological lipids, and is commonly used as a building-block of more structurally complex lipids. The carbon chain, typically between four and 24 carbons long, may be saturated or unsaturated, and may be attached to functional groups containing oxygen, halogens, nitrogen, and sulfur. Where a double bond exists, there is the possibility of either a cis or a trans geometric isomerism, which significantly affects the molecule's molecular configuration. Cis-double bonds cause the fatty acid chain to bend, an effect that is more pronounced the more double bonds there are in a chain. This in turn plays an important role in the structure and function of cell membranes. GlycerolipidsGlycerolipids are composed mainly of mono-, di-, and tri-substituted glycerols, the most well-known being the fatty acid triesters of glycerol, called triglycerides. The word triacylglycerol is sometimes used synonymously with triglyceride, however this is misleading with respect to these compounds as they contain no hydroxyl group. In these compounds, the three hydroxyl groups of glycerol are each esterified, typically by different fatty acids. Because they function as an energy store, these lipids comprise the bulk of storage fat in animal tissues. The hydrolysis of the ester bonds of triglycerides and the release of glycerol and fatty acids from adipose tissue are the initial steps in metabolising fat.Additional subclasses of glycerolipids are represented by glycosylglycerols, which are characterized by the presence of one or more sugar residues attached to glycerol via a glycosidic linkage. Examples of structures in this category are the digalactosyldiacylglycerols found in plant membranes and seminolipid from mammalian sperm cells.GlycerophospholipidsGlycerophospholipids, usually referred to as phospholipids, are ubiquitous in nature and are key components of the lipid bilayer of cells, as well as being involved in metabolism and cell signaling. Neural tissue (including the brain) contains relatively high amounts of glycerophospholipids, and alterations in their composition has been implicated in various neurological disorders. Glycerophospholipids may be subdivided into distinct classes, based on the nature of the polar headgroup at the sn-3 position of the glycerol backbone in eukaryotes and eubacteria, or the sn-1 position in the case of archaebacteria.

PhosphatidylethanolamineExamples of glycerophospholipids found in biological membranes are phosphatidylcholine (also known as PC, GPCho or lecithin), phosphatidylethanolamine (PE or GPEtn) and phosphatidylserine (PS or GPSer). In addition to serving as a primary component of cellular membranes and binding sites for intra- and intercellular proteins, some glycerophospholipids in eukaryotic cells, such as phosphatidylinositols and phosphatidic acids are either precursors of or, themselves, membrane-derived second messengers. Typically, one or both of these hydroxyl groups are acylated with long-chain fatty acids, but there are also alkyl-linked and 1Z-alkenyl-linked (plasmalogen) glycerophospholipids, as well as dialkylether variants in archaebacteria.SphingolipidsSphingolipids are a complicated family of compounds that share a common structural feature, a sphingoid base backbone that is synthesized de novo from the amino acid serine and a long-chain fatty acyl CoA, then converted into ceramides, phosphosphingolipids, glycosphingolipids and other compounds. The major sphingoid base of mammals is commonly referred to as sphingosine. Ceramides (N-acyl-sphingoid bases) are a major subclass of sphingoid base derivatives with an amide-linked fatty acid. The fatty acids are typically saturated or mono-unsaturated with chain lengths from 16 to 26 carbon atoms.

Sterol lipidsSterol lipids, such as cholesterol and its derivatives, are an important component of membrane lipids, along with the glycerophospholipids and sphingomyelins. The steroids, all derived from the same fused four-ring core structure, have different biological roles as hormones and signaling molecules. The eighteen-carbon (C18) steroids include the estrogen family whereas the C19 steroids comprise the androgens such as testosterone and androsterone. The C21 subclass includes the progestogens as well as the glucocorticoids and mineralocorticoids. The secosteroids, comprising various forms of vitamin D, are characterized by cleavage of the B ring of the core structure. Other examples of sterols are the bile acids and their conjugates, which in mammals are oxidized derivatives of cholesterol and are synthesized in the liver. The plant equivalents are the phytosterols, such as -sitosterol, stigmasterol, and brassicasterol; the latter compound is also used as a biomarker for algal growth. The predominant sterol in fungal cell membranes is ergosterol.Prenol lipidsPrenol lipids are synthesized from the five-carbon-unit precursors isopentenyl diphosphate and dimethylallyl diphosphate that are produced mainly via the mevalonic acid (MVA) pathway. The simple isoprenoids (linear alcohols, diphosphates, etc.) are formed by the successive addition of C5 units, and are classified according to number of these terpene units. Structures containing greater than 40 carbons are known as polyterpenes. Carotenoids are important simple isoprenoids that function as antioxidants and as precursors of vitamin A. Another biologically important class of molecules is exemplified by the quinones and hydroquinones, which contain an isoprenoid tail attached to a quinonoid core of non-isoprenoid origin. Vitamin E and vitamin K, as well as the ubiquinones, are examples of this class. Prokaryotes synthesize polyprenols (called bactoprenols) in which the terminal isoprenoid unit attached to oxygen remains unsaturated, whereas in animal polyprenols (dolichols) the terminal isoprenoid is reduced.SaccharolipidsSaccharolipids describe compounds in which fatty acids are linked directly to a sugar backbone, forming structures that are compatible with membrane bilayers. In the saccharolipids, a monosaccharide substitutes for the glycerol backbone present in glycerolipids and glycerophospholipids. The most familiar saccharolipids are the acylated glucosamine precursors of the LipidA component of the lipopolysaccharides in Gram-negative bacteria. Typical lipidA molecules are disaccharides of glucosamine, which are derivatized with as many as seven fatty-acyl chains. The minimal lipopolysaccharide required for growth in E. coli is Kdo2-Lipid A, a hexa-acylated disaccharide of glucosamine that is glycosylated with two 3-deoxy-D-manno-octulosonic acid (Kdo) residues.[39]PolyketidesPolyketides are synthesized by polymerization of acetyl and propionyl subunits by classic enzymes as well as iterative and multimodular enzymes that share mechanistic features with the fatty acid synthases. They comprise a large number of secondary metabolites and natural products from animal, plant, bacterial, fungal and marine sources, and have great structural diversity. Many polyketides are cyclic molecules whose backbones are often further modified by glycosylation, methylation, hydroxylation, oxidation, and/or other processes. Many commonly used anti-microbial, anti-parasitic, and anti-cancer agents are polyketides or polyketide derivatives, such as erythromycins, tetracyclines, avermectins, and antitumor epothilones.

IV. EQUIPMENTSa. ToolsNoName of ToolAmount

1234567891011121314Measurement glassBeaker glassDrop pipetteGlass stirrerTest tubeErlenmeyerGlass funnelElectric stoveExicatorWatch glassElectric balanceTweezersSentrifugaceFilter paper1221411111111sufficient

b. MaterialsNoName of MaterialAmount

123456789Chicken egg (egg yolk)AlcoholEtherAcetoneCadmium ChlorideKOH alcoholis 10%AquadestGlycerineKHSO33sufficientsufficientsufficientsufficientsufficientsufficientsufficientsufficient

V. PROCEDURE and OBSERVATION DATA

NoTreatmentObservation

1

2

34

5

6

7

8Cracking egg, take egg yolk + 25 ml ether + 50ml alcohol + 50 ml alcohol, stir it, let it during 10 minutes then filter it.Residue + 7 ml ether + 14 ml alcohol, filter it and its filtrate was vapored.

Residue + ether + acetone then filteredDissolving sediment + alcohol + CdCl2 alcoholis, stir it, let it then filter it.Filtrate was vapored,+ KOH alcoholis, stir it, heated + ether and filter it.

Filtrate + ether, vapored until dry

Cholesterol TestResidue + alcohol, centrifuged + alcohol

Centrifuged againLet it about 30 minutes.Acrolein Test0,5 gr fat + 0,5 gr KHSO3 , heatedRepeat with glycerolOccur coagulating, residue has yellow color and yellow filtrate.

Yellowish filtrateOld yellow filtrate, existence residueTurbid yellowish solutionYellow solution, existence clumped,Yellow sediment more paleBecome pasta, turbid yellowMore turbid solutionFormed 2 layers :Up : clear yellowBottom : orangeFiltrate : clear yellowResidue : orange

Yellow solutionFormed sedimentYellow solutionFormed sediment

MixedMore dissolve and bad smellBad smell very sting

VI. ANALYZE DATAAt trial lipids, aiming to isolate several types of lipids from egg yolk lecithin that will be generated. Where, lecithin is a phospholipid that becomes a major component of fractionphosphatides to extract the yellow egg or peanut soy isolates are mechanical , and chemical by using hexane or can be interpreted is a complex substance that is found in eggs and institutions supply the bone marrow. In its application, the lecithin in egg yolks and most often used as an agent emulsifier to mix oil and water , as in mayonnaise . This can happen because lecithin has a head that is hydrophilic and the tail is hydrophobic . Lecithin in eggs is dominated by the content of phosphatidyl kolina high, gliserolfosfolipid , long-chain fatty acids unsaturated, arachidonic acid , and DHA content not found in other sources of lecithin (such as beans). Lecithin is commercially can be obtained with high purity for the additives of food or medical purposes. In addition, lecithin is also known to help the stability of mayonnaise and mayonnaise to make it look more bold . Lipids are compounds which are prevalent in nature. This compound can be obtained by extracting natural materials both plants and animals with polar solvents not just as petroleum ether, benzene, chloroform, and others. Judging from the lipid compound structure composed of long hydrocarbon chains, so that lipids are not soluble in water. Lipid compounds are named based on the physical properties (solubility) than on the chemical structure. In general, lipids are divided into two major categories, namely simple lipid and lipid complexes. Lipids are included in the class are simple compounds that do not have the ester group and can not be hydrolyzed. This group is a steroid. Lipid complex group of compounds composed of ester groups and may have hydrolyzed, the fatty oils and waxes melipti. In this experiment, will be isolated or lipid content of lecithin and cholesterol from chicken egg yolk using alcohol and ether solvents. The first step in this experiment is to take as many as 3 eggs yolks, stir then add the alcohol and ether and then stirred again. Egg yolks into slightly thickened or lumpy. This indicates that the solvents (alcohols and ethers) can exstracted fat or lipid from the yolk. To be more complete extraction, the mixture allowed to stand for about 10 minutes and the resulting yellow laruran. Second, filter the mixture with filter paper that has been moistened with alcohol. The residue is then washed with alcohol and ether, while the filtrate is then evaporated in a water bath until dry. Washing the residue with ether alcohol intended to bind any residual alcohol ether in the residue thus obtained is pure fat deposition. Thick yellow precipitate then dissolved in ether and acetone was added. Dissolution is intended to bind the alcohol and ether remaining. While acetone has the function to precipitate the mixture. The precipitate formed is called lecithin. Third, filter and precipitate (lecithin) was dissolved again in the alcohol and CdCl2, then stirred and left for 10 minutes. After it was filtered and the residue obtained is used to test akreolin. Lecithin can also be called phosphatidyl kolina . Where, phosphatidyl kolina is sulfuric acid and base containing n-choline that serves to facilitate the transport and use of fatty acids by using the enzyme lecithin cholesterol acyl transferase. The structure of lecithin:

Can be derived from egg lecithin. Lecithin has a structure like fat but contains phosphoric acid, polar groups and nonpolar groups. Polar groups contained in the ester, phosphate is hydrophilic (water soluble likely), while non-polar groups contained in the fatty acid ester is lifofilik (likely to dissolve in fat). Lecithin is a waxy solid soft white color and turns brown due to exposure to light. Because it is hydrophilic, it can form a colloid. Lecithin is soluble in all organic solvents, except acetone. Goal is the addition of acetone to precipitate the lecithin. Lecithin (choline phospatidil) with the main components of choline, is an important nutrient that is found widely in various foods and available as supplements. Lecithin contains about 13% by weight of choline. Lecithin also zwiter ion, has a positive charge on N atom of choline and negative charge on the O atom of the phosphate group. Lecithin can be either polar (the choline) and non-polar (the fatty acid) making it very effective as an emulsifier. The filtrate that was still containingacetone evaporated to residual ether-acetone to evaporate. Then added therewith 10% KOH alcoholicwhile stirring then heated for 30 minutes. The results obtained are orange-colored solution. This is because the solution contains lecithin, when heated with alkali to form glycerol fatty acid choline phosphoric acid. The addition of 10% KOH alcoholics aims to transform fatty acids into soap and glycerol. Reaction of fatty acid with a base KOH:

Next, add 50 mL of ether into the solution and then filter it. Of ether was added, forming two layers where the top layer is yellow and bottom layer is orange. Ether filtrate containing cholesterol and then evaporated to dry residue thus obtained in the form of orange granules. The residue is then dried in the exicator which will then be used to test for cholesterol. Cholesterol is a metabolite of sterol-containing fat ( English : Waxy steroids) are found in cell membranes and circulated in the blood plasma . Is a type of lipid which is a molecule of fat or the like it. Cholesterol is a special type of lipid called steroids . Steroids are lipids that have a chemical structure specific. This structure consists of four ring atomsof carbon . Other steroids, including steroid hormones such as cortisol , estrogen , and testosterone . In fact, all steroid hormones are made from chemical changes in the basic structure of cholesterol. Cholesterol is a fat component and is one nutrient that is needed by the body in addition to other nutrients such as carbohydrates, protein, vitamins, and minerals. Cholesterol has a chemical structure as shown in the following figure:

These elements consist of fats in the blood cholesterol, triglycerides, phospholipids and free fatty acids. Only a quarter of the cholesterol contained in the blood are derived directly from the digestive tract is absorbed from food, the rest is the body's own production by liver cells. Fat contained in food will be elaborated into cholesterol, triglycerides, phospholipids and free fatty acids when digested in the gut. The fourth element of this fat will be absorbed from the intestine and enter the blood. Cholesterol, triglycerides, phospholipids and free fatty acids are not soluble in blood. In order to be transported in the bloodstream, cholesterol along with other fats (triglycerides and phospholipids) must bind to proteins to form a soluble compound and is called a lipoprotein. A cholesterol test performed by dissolving the residue in alcohol as much as 5 mL and then disentrifuge. Will get the white granules on the bottom of the tube in small amounts. Discard the solution above for the next step and add alcohol into granules and then disentrifuge again. The addition of alcohol is intended to dissolve cholesterol, because cholesterol is fat soluble in solvents such as ether, chloroform, benzene, and hot alcohol. Then, the mixture was let stand for 30 minutes. The results that obtained are yellowish solution, and the precipitate formed at the bottom of the tube in the form of yellowish granules called cholesterol. Acrolein test The purpose of acrolein test on lecithin and glycerin is to prove the existence of dehydrated glycerin into aldehydes, aksilat, or akreolin, and the presence of glycerol or fat. Glycerin usually be hydrated into an unsaturated aldehyde or akreolin and usually marked with white smoke and the smell. Akreloin test is usually positive in the form of free glycerol, but not for fatty acids. Of experiments lecithin which is yellow at first and then added KHSO4 in the form of fine grains resulting mixture is mixed evenly. This mixture is then put into a test tube and then heated in a water bath with a small fire in the early beginning and then becomes a large fire. This warming resulted from strong odors and yellow. With the yellow color and pungent odor which indicates that the test indicated its positive acrolein. This means that the lecithin contained in the dehydration is aldehit aksilat or akreolin and there are fat. Then repeat the experiment with the same steps, but replace the lecithin with glycerin. And the result is a fishy smell and stinging occur. But the odor generated no more and no fishy smell more pungent than lecithin. But these results indicate a positive test for akreolin glycerin. Function of KHSO4 in this experiment is to draw water that will hydrate glycerol into the aldehyde or akreolin aksilat characterized by white smoke and strong odors. Reactions that occur Lecithin + KHSO4

Glycerol + KHSO4

VII. CONCLUSION1. Lipids are organic compounds that have physical properties such as fat, and can be obtained from animal or plant by extraction with hot alcohol, ether and other fat solvent. 2. Lecithin is one component of egg yolk contains fat and cholesterol. 3. Inside there is a chicken egg yolk cholesterol in the form of a white crystalline solid. 4. Cholesterol is a necessary intermediate in the biosynthesis of steroid hormones. 5. Akroelin test on fatty acids and glycerol are both positive. This is evidenced by the stench of fatty acids and glycerol to be dehydrated into aldehydes, and akroelin aksilat. 6. KHSO4 additional function is to attract water so be hydrated glycerol into unsaturated aldehydes are marked with white smoke and the smell. 7. Function of the addition of: a. alcohol, ether: as a solvent of the remaining egg whites or a protein that may be left behind when the separation of egg white and yolk. b. KOH alcoholics: to provide the alkaline conditions c. CdCl 2: to hydrolyze lecithin d. acetone: to precipitate the lecithin 8. chemical structure of lecithin:

9. chemical structure of cholesterol:

10. fatty acid reaction with KOH

11. Reactions that occur Lecithin + KHSO4

Glycerol + KHSO4

VIII. BIBLIOGRAPHY (2012, March 3). was taken April 30, 2012, from Wikipedia: http://id.wikipedia.org/wiki/Lesitin (2012, April 12). was taken April 30, 2012, from Wikipedia: http://id.wikipedia.org/wiki/Kolesterol Anna, P., & Supriyanti, T. (2006). Dasar-Dasar Biokimia. Jakarta: UI Press. Fajri , E. (2011, June 28). Archives: 2011 (81): Juni (23). was taken April 30, 2012, from elfa Fajri: http://elfafajri.blogspot.com/2011/06/lipida.html Fessenden dan Fessenden. (2000). Dasar-Dasar Biokimia. Jakarta: Erlangga. muhammadpajri . (2012, January 4). Arsip Blog: Januari (18). Dipetik April 30, 2012, from MUHAMMAD PAJRI THP 09' UNJA: http://muhammadpajri1991.blogspot.com/2012/01/lesitin-sebagai-emulsifier.html Sugiharto. (1989). Biokimia. Jakarta: Gramedia. Tim Dosen. (2012). Petunjuk Praktikum Biokimia. Surakarta: Lab. P. MIPA Kimia UNS. Winarno, F. G. (2002). Kimia Pangan dan Gizi. Jakarta: Erlangga.

IX. ATTACHMENT One sheet of temporary report Lipid 1 and Lipid 2

Surakarta, 3rd may 2012AssitantPracticant,

Group 4

13 | PageExperiment in Biochemistry