Lab 7 &8 lipids. Lipids are Biomolecules that contain fatty acids or a steroid nucleus. Soluble in...
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Lab 7 &8 lipids
Lab 7 &8 lipids. Lipids are Biomolecules that contain fatty acids or a steroid nucleus. Soluble in organic solvents but not in water. Named for the Greek
Lipids are Biomolecules that contain fatty acids or a steroid
nucleus. Soluble in organic solvents but not in water. Named for
the Greek word lipos, which means fat. Extracted from cells using
organic solvents.
Slide 3
Definition: Lipids are a group of naturally occurring
substances consisting of the higher fatty acids, their naturally
occurring compounds and substances with different structures,
Organic substances relatively insoluble in water but soluble in
organic solvents like chloroform, ether and benzene.
Slide 4
Storage form of energy Structural component of cell membrane
Precursor of many steroid hormones, vitamin D Protection of
internal organs
Slide 5
Slide 6
Types of lipids Lipids with fatty acids Waxes Fats and oils
(trigycerides) Phospholipids Sphingolipids Lipids without fatty
acids Steroids.
Slide 7
Slide 8
1. Simple Lipids : fatty acid with alcohol (GLYCEROL or other
than GLYCEROL) If alcohol is glycerol .Fats (triglyceride) and Oils
Oils: Triglycerides rich in unsaturated fatty acids are generally
liquid at room. Fats: Triglycerides rich in saturated fatty acids
are generally semisolids or solids at room temperature. If alcohol
other than glycerol .. Waxes Result of hydrolysis gives FA with
alcohol and containing additional[prosthetic] groups. 2. Conjugated
Lipids (polar lipids): Result of hydrolysis gives FA with alcohol
and containing additional[prosthetic] groups. A. Phospholipids -
contain a phosphoric acid molecule and a fat molecule. FA + ALCOHOL
+ PHOSPHORIC ACID B. Glycolipid- contain a carbohydrate and a fat
molecule. Glycolipids FA + ALCOHOL[SPINGOSINE] +CARBOHYDRATE WITH
NITROGEN BASE C. Sulfolipids - contain a sulfate radical. D.
lipoprotein ( Lipid with prosthetic group PROTEIN ) Classification
of Lipids
Slide 9
Fatty acids Fatty acids are saturated with all single CC bonds.
are solids at room temperature unsaturated with one or more double
C=C bonds. are liquids at room temperature.
Slide 10
Slide 11
Lipoproteins Combine lipids with proteins and phospholipids.
Are soluble in water because the surface consists of polar
lipids.
Slide 12
Types of Lipoproteins Lipoproteins Differ in density,
composition, and function. Include low-density lipoprotein (LDLs)
and high- density lipoprotein (HDLs).
Slide 13
Transport of Lipoproteins in the Body Copyright 2007 by Pearson
Education, Inc. Publishing as Benjamin Cummings
Slide 14
Lipoproteins consist of core (triacyglycerol and or
cholostylester)and surface monolayer(phospholipid unesterified
cholestrol and specific proteins) VLDL transport triglycerides from
liver to adipose tissue and mucsle LDL transport cholesterol from
liver to tissue HDL transport the cholesterol from tissues back to
liver
Slide 15
High-density lipoprotein (HDL) Because HDL particles remove
excess cholesterol from the body, they are considered protective.
Thats why HDL cholesterol is often referred to as good
cholesterol.
Slide 16
Low-density lipoprotein (LDL) When the cells have all the
cholesterol they need, they reduce the number of receptors, or
gateways, for cholesterol to enter. As a result, cholesterol
contained in the LDL particles (LDL cholesterol) starts to build up
in the blood stream.
Slide 17
Over time, circulating LDL cholesterol undergoes chemical
changes (becomes oxidized) and also reduces in size. These smaller
particles more easily enter the blood vessel wall and start to
build up under the vessel lining. Deposits of LDL cholesterol
particles in vessel walls are called plaques and can lead to
inflammation, bleeding into the area, and calcium buildup
(calcification). Eventually, the buildup of plaques can start to
crowd the space within the blood vessel and obstruct the blood
flow. This process of plaque accumulation is called
atherosclerosis.
Slide 18
When atherosclerosis affects the arteries leading to the heart
(coronary arteries), one may have chest pain and other symptoms of
coronary artery disease. If plaques tear or rupture, a blood clot
may form blocking the flow of blood or breaking free and plugging
an artery downstream. If blood flow to part of the heart stops, a
heart attack will occur. If blood flow to the brain stops, a stroke
occurs. This is why LDL cholesterol is often referred to as bad
cholesterol.
Slide 19
3.Derived Lipids A. Fatty acids B. Glycerol C. Cholesterol and
other steroid (Vit. D) D. Vitamins A, E, K
Slide 20
It include a wide variety of substances with different
structures:
Slide 21
Cholesterol Is the most abundant steroid in the body. is a waxy
substance found in all of the body cells. Has methyl CH 3 - groups,
alkyl chain, and -OH attached to the steroid nucleus.
Slide 22
Functions help and maintain cell membranes. protect and
insulate nerve fibers. involved in the formation of sex hormones,
such as estrogen. production of bile salts, which help digesting
fats.
Slide 23
Cholesterol in the body Is obtained from meats, milk, and eggs.
Is synthesized in the liver. Is needed for cell membranes, brain
and nerve tissue, steroid hormones, and Vitamin D. Clogs arteries
when high levels form plaque. Considered elevated if plasma
cholesterol exceeds 200 mg/dL.
Slide 24
Chemical tests for: 1. Solubility in polar and nonpolar solvent
2. Emulsification 3. Oxidation of unsaturation FA 4. Qualitative
analysis of detection in natural products ( salkowski
reaction)
Slide 25
Solubility in polar and nonpolar solvent (like dissolve like )
The solubility depend on polarity Reagents : L IPIDS : Olive oil
(vegetable oil), butter (animal fat), stearic acid (saturated fatty
acid), oleic acid (unsaturated fatty acid), Solvents: dilute acid
and alkali solutions, acetone,cold alcohol, hot alcohol, benzene,
chloroform, ether and carbon tetrachloride.
Slide 26
Solubility in polar and nonpolar solvent Procedure 1. Add a
2drops of the liquid fat or 0.1 g of the solid fat in labeled test
tubes. 2. To each test tube add a 1 ml of solvent and write down
your observations. 3. Repeat the experiment with a different
solvent and make your observations.
Slide 27
An emulsion is a mixture of two or more liquids that are
normally immiscible (no mixable).mixtureliquidsimmiscible Bile
Salts Synthesized from cholesterol and stored in the gallbladder
When you eat fat, the bile salts act as soap and help emulsify the
large globules of fat.
Slide 28
A. Two immiscible liquids, not yet emulsified B. An emulsion of
Phase II dispersed in Phase I C. The unstable emulsion
progressively separates D. The surfactant (outline around
particles) positions itself on the interfaces between Phase II and
Phase I, stabilizing the emulsionsurfactant Emulsification types:
Temporary and permanently Permanently more free fatty acids and no
more bond found Temporary few fatty acids and few bond broken
Slide 29
Emulsification Reagents Neutral olive oil(partially oxidation,
produce few fatty acids), 0.5 % Na 2 C0 3 (emulsifying agent),
Rancid olive oil(complete oxidation many of free fatty acids)
Prepare neutral olive oil: shaking ordinary oil with 10% solution
of sodium bicarbonate this mixture should then be extracted with
ether and the ether should be removed by evaporation Rancid olive
oil: to prepare add 5 drops of oleic acid to 10 ml of olive oil and
mix well
Slide 30
Rancidity is the development of unpleasant smells in fats and
oils, which are often accompanied by changes in their texture and
appearance.
Slide 31
Procedure 1.Shake up a drop of neutral olive oil with a little
water in a test tube, the fat becomes finally divided forming an
emulsion, upon standing fat separates and rises to the top. 2.To 5
ml water in a test tube add 2 to 3 drops of 0.5 percent Na 2 CO 3.
Introduce a drop of neutral olive oil and shake, the emulsion is
not permanent and is not so transitory. 3.Repeat step 2 using
rancid olive oil. In this case the alkali combines with the free
fatty acids to form soap and the soap being an emulsifying agent,
it emulsifies the fat.
Slide 32
Oxidation of un saturation FA unsaturated fatty acids have one
or more double bonds Into test tube place one drop of oleic oil and
3 ml of Na 2 CO 3 Warm it slightly and drop solution KMnO4
(oxidizing agent).After each drop the violet color disappear The
end of the reaction recognized by ppt brown solid MnO2
Slide 33
Qualitative analysis of cholesterol detection in natural
products salkowski reaction Concentrated sulfuric acid is highly
hygroscopic and it removes two molecules of water from two
molecules of cholesterol. It causes a connection at position 3
forming bi cholestadien. Simulatineously the sulfuric acid
sulfonates the molecule of bi cholestadien at position 7,7 of
aromatic ring and as a final product, red color bi sulfonic acid of
bicholestadien is formed
Slide 34
B) Bisulfonic acid of bi-cholestadiene
Slide 35
Salkowski reaction Reagents: 2% pure cholesterol, extract of
yellow hen egg, lecithin and oil Chloroform(Inhalation: Inhalation
of high concentrations may cause central nervous system effects
characterized by nausea, headache, dizziness, unconsciousness and
coma) Conc sulfuric acid
Slide 36
Procedure Into the dry test tubes the reaction is very
sensitive for even traces of water add 1ml o chloroform solution of
2% of pure cholestrol an extract of yellow hen egg Add carfully
concentrated sulfuric acid keeping the tube under 45 angle dont mix
In the presence of cholesterol, the upper chloroform layer turns
red whereas a lower acidic become yellow green
Slide 37
Isolation of cholesterol from hen egg Hen eggEgg
whiteAlbuminEgg yolk Lipids + glycoproteins
Egg yolk is rich in lipids of which the most two prominent (are
cholesterol and phosphatidyl choline (lectithin Phosphatidylcholine
(once given the trivial name 'lecithin') is usually the most
abundant phospholipid in animals and plants, often amounting to
almost 50% of the total, and as such it is obviously a key building
block of membrane bilayers. In particular, it makes up a very high
proportion of the outer leaflet of the plasma membrane.
Phosphatidylcholine is also the principal phospholipid circulating
in plasma, where it is an integral component of the lipoproteins,
especially the HDL.lipoproteins On the other hand, it is less often
found in bacterial membranes, perhaps 10% of species, but there is
none in the 'model bacteria', Escherichia coli and Bacillus
subtilis.
Slide 40
Experiment Principle: Based on the solubility in different
organic solvent, lipids can be separated from each other.
Cholestrol is readily soluble in acetone, while most complex lipids
are insoluble in this solvent. Reagents and instruments A hard
boiled hen egg Acetone, Ice, Water bath, Erlenmeyer flask, Funnel,
Filter paper, 250 ml beaker
Slide 41
Reagents and instruments A hard boiled hen egg Acetone, Ice,
Water bath, Erlenmeyer flask, Funnel, Filter paper, 250 ml
beaker
Slide 42
Slide 43
Acetone (systematically named propanone) is the organic
compound with the formula (CH 3 ) 2 CO. It is a colorless,
volatile, flammable liquid, and is the simplest ketone. organic
compoundformulaketone
Slide 44
1. Extraction 1. Peal the hard boiled egg and remove the egg
white. 2.In a 250 ml beaker weigh a egg yolk and mash it. 3. To the
mashed egg yolk add a 25 ml of acetone and keep stirring for 5 min.
4. Allow the solid to settle and carefully remove the acetone (the
above liquid phase) and transfer it to a clean 100 ml Erlenmeyer
flask and cover it tightly. 5. To the remained solid phase add a
new 25 ml of acetone and repeat step 4. Now you have collected
twice the 25 ml of acetone extract (it contains the cholesterol).
1. Peal the hard boiled egg and remove the egg white. 2.In a 250 ml
beaker weigh a egg yolk and mash it. 3. To the mashed egg yolk add
a 25 ml of acetone and keep stirring for 5 min. 4. Allow the solid
to settle and carefully remove the acetone (the above liquid phase)
and transfer it to a clean 100 ml Erlenmeyer flask and cover it
tightly. 5. To the remained solid phase add a new 25 ml of acetone
and repeat step 4. Now you have collected twice the 25 ml of
acetone extract (it contains the cholesterol).
Slide 45
2. Isolation 1. Using a funnel and a filter paper, filter the
50 ml of acetone extract into a clean 100 ml beaker. 2.Evaporate
the acetone by placing the beaker on the steam bath in the hood
until about 10 ml of extract is left. 3.Cover the warm acetone
extract and immediately transfer it on ice and let it stand for 20
min. A white precipitate will form (this is the crude cholesterol).
Remove the flask from ice. Get rid of the liquid phase. 1. Using a
funnel and a filter paper, filter the 50 ml of acetone extract into
a clean 100 ml beaker. 2.Evaporate the acetone by placing the
beaker on the steam bath in the hood until about 10 ml of extract
is left. 3.Cover the warm acetone extract and immediately transfer
it on ice and let it stand for 20 min. A white precipitate will
form (this is the crude cholesterol). Remove the flask from ice.
Get rid of the liquid phase.
Slide 46
4.To the cholesterol precipitate add 15 ml of acetone and stir
at room temp. Note that not all precipitate will dissolve (the
contaminating phospholipids remain insoluble). 5. Weigh a clean,
dry watch glass. Filter the dissolved cholesterol solution into the
pre- weighed watch glass. Allow the acetone to evaporate in the
hood. The remained crystals is the cholesterol. 6. Weigh the watch
glass with cholesterol. Calculate the yield of cholesterol. 4.To
the cholesterol precipitate add 15 ml of acetone and stir at room
temp. Note that not all precipitate will dissolve (the
contaminating phospholipids remain insoluble). 5. Weigh a clean,
dry watch glass. Filter the dissolved cholesterol solution into the
pre- weighed watch glass. Allow the acetone to evaporate in the
hood. The remained crystals is the cholesterol. 6. Weigh the watch
glass with cholesterol. Calculate the yield of cholesterol. 3.
Purification
Slide 47
Solubility of cholesterol: Soluble in diethyl ether, acetone.
Very slightly soluble in cold water. Solubility in water:
0.2mg/100ml or 0.2% Slightly soluble in alcohol; more soluble in
hot alcohol. Soluble in chloroform, pyridine, benzene, petroleum
ether, oils, fats, aqueous solutionsof bile salts. Solubility in
ether: 1 g/2.8 ml ether. Solubilitiy in chloroform: 1 g/4.5 ml
chloroform. Solubility in pyridine: 1 g/1.5 mlpyridine. whereas
acetone a carbonyl group with a negatively charged oxygen and a
short chain of hydrocarbons, this makes it a more suitable
substance to act as a dissolvent for an amphiphilic compound.