Lipids Types: Simple, Compound and Derived Lipids

The following points highlight the top three types of lipids. The types are: 1. Simple Lipids 2. Compound Lipids 3. Derived Lipids.

Type # 1. Simple Lipids:

A. Fats:

(a) They are esters of fatty acids with glyc­erol.

(b) They are found in nature in large quanti­ties.

(c) They are the best reserve of food material in the human body.

(d) They act as insulator for the loss of body heat.

(e) They act as a padding material for pro­tecting internal organs.

The chemical structure of fat (triglyceride) consists of three different molecules of fatty acids with one molecule of glycerol.

The three different fatty acids (R₁, R₂, R₃) are esterified with the three hydroxyl groups of glycerol:

Physical Properties of Fats:

(a) The fats are insoluble in water, but readily soluble in ether, chloroform, benzene, car­bon tetrachloride.

(b) They are readily soluble in hot alcohol but slightly soluble in cold.

(c) They are themselves good solvents for other fats, fatty acids, etc.

(d) They are tasteless, odorless, colourless and neutral in reaction.

(e) Several neutral fats are readily crystallized, e.g., beef, mutton.

(f) Their melting points are low.

(g) The specific gravity of solid fats is about 0.86. So fat people float in water more read­ily than thin ones.

(h) They spread uniformly over the surface of water; so the spreading effect is to lower surface tension.

Identification of Fats and Oils:

(a) Hydrolysis:

1. Hydrolysis of triacylglycerol takes place by lipases producing fatty acids and glyc­erol.

2. Phospholipases attack the ester linkage of phospholipids.

(b) Saponification:

1. Boiling with an alcoholic solution of strong metallic alkali hydrolyses triglycerides into glycerol and fatty acids —this is called saponification.

2. The products are glycerol and the alkali salts of the fatty acids which are called soaps.

3. Fats, phospholipids, glycolipids and waxes are called saponifiable lipids.

4. Steroids, polyisoprenoids and higher alcohols are grouped as un-saponifiable lipids because they cannot give rise to soap.

(c) Saponification number:

1. The number of milligrams of KOH required to saponify 1 gram of fat or oil.

2. The amount of alkali needed to saponify a given quantity of fat will depend upon the number of-COOH group present. It is inversely proportional to the average mo­lecular weight of the fatty acids in the fat i.e. the fats containing short chain fatty acids will have more -COOH groups per gram than long chain fatty acids—this will take up more alkali and, hence, will have higher saponification number.

Example:

Butter—containing a larger proportion of short chain fatty acids such as butyric and caproic acids, has relatively high saponification number 220 to 230.

(d) Acid number:

1. The number of milligrams of KOH required to neutralize the free fatty acids of 1 gram of fat.

2. Significance: The acid number indicates the degree of rancidity of the given fat.

(e) Iodine number:

1. This is the amount (in grams) of iodine absorbed by 100 grams of fat.

2. This is the measure of the degree of unsaturation of a fat.

3. Significance: If the fat contains higher number of unsaturated fatty acids, it be­comes essential for the protection of heart disease. These unsaturated fatty acids, combined with the cholesterol, are oxi­dized in the liver—producing bile acids, bile salts, vit., D, gonadotrophin hormones. They prevent atherosclerosis.

(f) Acetyl number:

1. The number of milligrams of KOH required to neutralize the acetic acid obtained by saponification of 1 gram of fat after it has been acetylated.

2. This is a measure of the number of hy­droxy acid groups in the fat.

(g) Polenske number:

1. The number of milliliters of 0.1 (N) KOH required to neutralize the insoluble fatty acids from 5 grams of fat.

(h) Reichert-Miessl number:

1. This is same as the Polenske number ex­cept that the soluble fatty acids are meas­ured by titration of the distillate obtained by steam distillation of the saponification mixture.

2. Significance:

It measures the amount of volatile soluble fatty acids.

(i) Halogenation:

1. Chlorine, bromine and iodine atoms may be added to the double bonds of unsatu­rated fatty acids containing fats.

(j) Rancidity:

1. Nearly all natural fats are oxidized when exposed to air, light, moisture, particularly, if warm, it develops an unpleasant odour and taste. The enzyme lipase—in the pres­ence of moisture and warm temperature— bring about hydrolysis rapidly.

2. This happens due to the formation of peroxides at the double bonds of unsatu­rated fatty acids.

3. Vitamin E is an important natural antioxi­dant and prevents development of rancid­ity.

(k) Soaps:

1. Soaps are metallic salts of fatty acids.

2. Soaps are formed by adding alkalis to fatty acids.

3. Soaps of unsaturated fatty acids are softer and more water soluble than those of satu­rated fatty acids.

4. Potassium soap of an acid is more water-soluble and softer than the sodium soap; calcium and magnesium soaps are far less soluble.

B. Waxes:

1. They are esters of fatty acids with higher alcohols other than glycerol.

2. In the human body, the commonest waxes are esters of cholesterol.

3. They are mainly three types:

(a) True waxes are esters of higher fatty acids with acetyl alcohol or other higher straight chain alcohols.

(b) Cholesterol esters are esters of fatty acid with cholesterol.

(c) Vitamin A and vitamin D esters are palmitic or stearic acid esters of vita­min A (Retinol) or vitamin D, respec­tively.

Type # 2. Compound Lipids:

A. Phospholipids (phosphatides):

(i) They are esters of fatty acids with glyc­erol containing an esterified phosphoric acid and a nitrogen base.

(ii) They are present in large amounts in nerve tissue, brain, liver, kidney, pancreas and heart.

Biological functions of phospholipids:

(i) They increase the rate of fatty acid oxida­tion.

(ii) They act as carriers of inorganic ions across the membranes.

(iii) They help blood-clotting.

(iv) They act as prosthetic group to certain en­zymes.

(v) They form the structures of membranes, matrix of cell wall, myelin sheath, microsomes and mitochondria.

Classification:

It is based on the type of alcohol present in the phospholipid.

There are three types:

1. Glycerophosphatides — In this, glycerol is the alcohol group.

Example:

(i) Phosphatidyl ethanolamine (cephalin).

(ii) Phosphatidyl choline (Lecithin).

(iii) Phosphatidyl serine.

(iv) Plasmalogens.

(v) Phosphatidic acid.

2. Phosphoinositides — In this, inositol is the alcohol.

Example:

Phosphatidyl inositol (Lipositol).

3. Phosphosphingosides — In this, sphingosine is an amino alcohol.

Example:

Sphingomyelin.

The phospholipids include the following groups:

1. Phosphatidic acid and phosphatidyl glycerol’s:

Phosphatidic acid is important as an intermediate in the synthesis of triacylglycerol’s and phospholipids.

Cardiolipin:

(a) It is formed from phosphatidyl glyc­erol.

(b) Chemically, it is di-phosphatidyl glyc­erol.

(c) It is found in inner membrane of mi­tochondria and bacterial wall.

2. Lecithin (Phosphatidylcholine):

The lecithin’s contain glycerol and fatty acids, phosphoric acid and choline (nitrogenous base). Lecithin’s generally contain a satu­rated fatty acid at α position and an un­saturated fatty acid at β position. They can exist in α or β forms.

Physical Properties:

(i) Lecithin’s are waxy, white substances but become brown soon when exposed to air

(ii) They are soluble in ordinary fat solvents except acetone.

(iii) They decompose when heated.

(iv) They constitute valuable agents for the emulsifications of fats and oils.

Chemical Properties of Lecithin:

(i) When aqueous solution of lecithin’s are shaken with H₂SO₄, choline is split off, forming phosphatidic acid.

(ii) When lecithin’s are boiled with alkalis or mineral acids, not only choline is split off, but phosphatidic acid is further hydrolyzed to glycerophosphoric acid and 2 molecules of fatty acids:

Lecithin → H₂SO₄ Phosphatidic acid + choline.

Phosphatidic acid → Glycerophosphoric acid + fatty acids (2 mol)

Physiological Functions of Lecithin:

(i) It facilitate the combinations with proteins to from lipoproteins of plasma and cells.

(ii) Acetylcholine formed from choline has an important role in the transmission of nerv­ous impulses across synapses.

(iii) Choline is the most important lipotropic agent as it can prevent formation of fatty liver.

(iv) Lecithin lowers the surface tension of lung alveoli. Dipalmityl lecithin is a major constituent of “lung surfactant” which pre­vents the adherence of the inner surface of the alveoli of the lungs (preventing the collapse of the alveoli) by its surface ten­sion lowering effect. The absence of this in the alveolar membrane of some prema­ture infants causes the respiratory distress syndrome in them.

(v) It lowers the surface tension of water mol­ecule and helps in the emulsification of fat.

Difference of Lecithin and Cephalin:

Cadmium chloride compound of cephalin is soluble but cadmium chloride compound of leci­thin is insoluble.

3. Cephalitis (Phosphatidyl ethanolamine):

They always occur in the tissues in asso­ciation with lecithin’s and are very similar in properties. The only difference is the nitrogenous base.

4. Phosphatidyl Serine:

A cephaline like phospholipid is found in tissues.

5. Phosphatidyl inositol (Lipositol or Phosphoinositides):

(i) It acts as second messenger in Ca⁺⁺ dependent hormone action.

(ii) Some signals must provide commu­nication between the hormone receptor on the plasma membrane and intracellular Ca⁺⁺ reservoirs.

(iii) They are more acidic than the other phospholipids.

6. Lysophospholipids:

(i) These are phosphoacylglycerols con­taining only one acyl radical in a po­sition e.g., Lysolecithin.

(ii) Formation:

(a) By the action of phospholipase.

(b) By interaction of lecithin and cho­lesterol in presence of the enzyme lecithin cholesterol acyl transferase, so lysolecithin and cholesterol ester are formed

7. Plasmalogens:

(i) These are the contents of brain and muscle.

(ii) Structurally, these resemble lecithin’s and cephalins but give a positive re­action when tested for aldehydes with Schiff s reagent (fuchsin-sulphurous acid) after pretreatment of the phos­pholipid with mercuric chloride.

(iii) They possess an ether link in a posi­tion instead of ester link. The alkyl radical is an unsaturated alcohol.

8. Sphingomyelins:

(i) These are found in large quantities in brain and nerve tissue.

(ii) The concentrations of these phospholipids are increased in Niemann-Pick disease in the liver and spleen.

(iii) These contain sphingosine (18 car­bon) (amino alcohol) fatty acid, phos­phoric acid and choline. No glycerol is present.

(iv) In sphingosine molecule -NH₂ group binds a fatty acid by an amide link­age to produce ceramide. When phos­phate group is attached to ceramide it is called ceramide phosphate.

(v) When choline is split off from sphin­gomyelin, ceramide phosphate is left.

Action of Phospholipase:

(a) Phospholipase A₁ attacks the ester bond in position 1 of phospholipid.

(b) Phospholipase A₂ attacks β position and form

Lysolecithin + one mol. fatty acid.

(c) Phospholipase B (lysophospholipase) at­tacks lysolecithin and hydrolyzes ester bond in α position and forms glyceryl phosphoryl choline + 1 mol. fatty acid.

(d) Phospholipase C hydrolyzes phosphate ester bond and produces α, β di-acyl glyc­erol + phosphoryl choline.

(e) Phospholipase D-splits off choline and phosphatidic acid is formed

B. Glycolipids:

These contain an amino alcohol (sphingosine or iso-sphingosine) attached with an amide linkage to a fatty acid and glycosidically to a carbohydrate moiety (sugars, amino sugar, sialic acid).

These are further classified into:

(i) Cerebrosides.

(ii) Gangliosides.

(i) Cerebrosides:

(a) Cerebrosides contain galactose, a high molecular weight fatty acid and sphingosine. Therefore, they may also be classified as sphingolipids.

(b) They are the chief constituent of my­elin sheath.

(c) They may be differentiated by the type of fatty acid in the molecule.

These are:

Kerasin—Containing lignoceric acid [CH, — (CH₂)₂₂ — COOH].

Cerebron—Containing a hydroxylignoceric acid (cerebronic acid).

[CH₃—(CH₂)₂, — CH(OH)—COOH].

Nervon—Containing an unsaturated homologue of lignoceric acid called nervonic acid. [CH, — (CH₂)₇ — CH = CH — (CH₂)₁₃ — COOH].

Oxynervon—Containing hydroxy-nervonic acid [CH₃ — (CH₂)₇ — CH = CH — (CH₂)₁₂— CH(OH) — COOH].

(d) Stearic acid is a major component of the fatty acids of rat brain cerebrosides.

(e) Cerebrosides, specially cerebronic acid, increases in Gaucher’s disease and the Kerasin characterized by glu­cose replacing galactose.

(f) The cerebrosides are in much higher concentration in medullated than in non-medullated nerve fibers.

(ii) Gangliosides:

(a) These are glycolipids occurring in the brain.

(b) Gangliosides contain ceramide (sphingosine + fatty acids), glucose, galactose, N-acetylgalactosamine and sialic acid.

(c) Some gangliosides also contain di-hydro-sphingosine or gangliosine in place of sphingosine.

(d) Most of the gangliosides contain a glucose, two molecules of galactose, one N-acetylgalactosamine and up to three molecules of sialic acid.

Types of Ganglioside:

C. Other compound lipids:

1. Lipoproteins:

(i) Triacylglycerol (45%), phospholipids (35%), cholesterol and cholesteryl esters (15%), free fatty acids (less than 5%) and also protein combine to form a hydrophilic lipoprotein complex.

(ii) Since pure fat is less dense than water, the proportion of lipid to protein in lipoproteins in plasma is by ultracentrifugation.

(iii) The density of lipoproteins increases as the protein content rises and the lipid con­tent falls and the size of the particle be­comes smaller.

(iv) Lipoproteins may be separated on the ba­sis of their electrophoretic properties and may be identified more accurately by means of immuno-electrophoresis.

(v) Four major groups of lipoproteins have been identified which are important physi­ologically and in clinical diagnosis in some metabolic disorders of fat metabo­lism.

These are:

(a) Chylomicrons.

(b) Very low density lipoproteins (VLDL or pre-β-lipoproteins).

(c) Low density lipoproteins (LDL or β-lipoproteins).

(d) High density lipoproteins (HDL or α-lipoproteins).

(vi) Chylomicrons and VLDL: Predominant lipid is triacylglycerol (50%) and choles­terol (23%). The concentrations of these are increased in atherosclerosis and coro­nary thrombosis etc.

LDL:

Predominant lipid is cholesterol (46%) and phospholipids (23%). Increase in atherosclerosis and coro­nary thrombosis, etc.

HDL:

Predominant lipid is phospholipid (27%) and proteins (45%).

(vii) The protein moiety lipoprotein is known as an apoprotein which constitute nearly 60% of some HDL and 1% of chylomi­crons. Many lipoproteins contain more than one type of apoprotein polypeptide.

(viii) The larger lipoproteins (such as chylomicrons and VLDL) consist of a li­pid core of nonpolar triacylglycerol and cholesteryl ester surrounded by more po­lar phospholipid, cholesterol and Apo proteins.

Importance:

(i) To transport and deliver the lipids to tis­sues.

(ii) To maintain structural integrity of cell sur­face and subcellular particles like mito­chondria and microsomes.

(iii) The β-lipoprotein fraction increases in severe diabetes mellitus, atherosclerosis etc. Hence determination of the relative concentrations of α-and β-lipoproteins and pre-β- lipoproteins are of diagnostic im­portance.

2. Amino lipids:

Phosphatidyl ethanolamine and serines are amino lipids and sphingomyelins and gangliosides contain substituted amino groups.

3. Sulpholipids (Sulphatides):

(i) These have been isolated from brain and other animal tissues.

(ii) These are sulphate derivatives of the galactosyl residue in cerebrosides.

Type # 3. Derived Lipids:

A. Fatty acids:

(i) These are obtained by the hydrolysis of fats.

(ii) Fatty acids occurring in natural fats usu­ally contain an even number of carbon atoms because they are synthesized from 2-carbon units and are straight chain de­rivatives.

(iii) The straight chain may be saturated (con­taining no double bonds) or unsaturated (containing one or more double bonds).

(iv) Carbon atoms of fatty acids are numbered from the carboxyl carbon (carbon No.l). The carbon atom adjacent to the carboxyl carbon (Carbon No. 2) is also known as the α -carbon. Carbon atom No. 3 is the β- carbon and the end methyl carbon is known as the γ-carbon.

(v) Various conventions are used for indicat­ing the number and position of the dou­ble bonds, e.g., Δ⁹ indicates a double bond between carbon atoms 9 and 10 of the fatty acid.

Types:

i. Straight chain.

ii. Branched chain.

iii. Substituted (methyl substituted- cerebronic acid)

iv. Cyclic (chaulmoogric acid) used in lep­rosy.

1. Straight chain:

(a) Saturated {odd (less than 10 carbon atoms) & even (greater than 10 carbon atoms)}.

(b) Unsaturated (odd & even).

(Straight chain even number fatty acid is common)

B. Saturated fatty acids:

General formula for saturated fatty acids is C_nH_2n+1 COOH. Other higher fatty acids occur in waxes. A few branched-chain fatty acids have also been isolated from both plant and animal sources.

Prostanoids include Prostaglandins (PG), and thromboxane’s (TX).

General characteristics of prostanoid

(a) All are 20 carbon compounds.

(b) Trans double bond at 13 positions.

(c) -OH group at 15 position.

Prostaglandins (PG):

(a) They virtually exist in every mammalian tissue and act as local hormones.

(b) They have important physiologic and pharmacologic activities.

(c) They are synthesized in vivo by cyclization of the center of the carbon chain of 20-C polyunsaturated fatty acids (e.g., arachidonic acid) to form a cyclopentane ring.

(d) Three different eicosanoic fatty acids give rise to three groups of eicosanoids charac­terized by the number of double bonds in the side chains, e.g., PG₁, PG₂, PG₃. Varia­tions in the substituent groups attached to the rings give rise to different types in each series of prostaglandins, as for exam­ple, “E” type of Prostaglandin has a keto group in position 9, whereas the “F” type has a hydroxyl group in this position.

Prostacyclin’s (PGI):

(a) They are formed in vascular endothelium and continually formed in heart. They are also formed in kidneys.

(b) They are formed from cyclic endo-peroxide PGH₂ by the action of microsomal Prostacyclin synthetase.

(c) They inhibit platelet aggregation and gas­tric secretion from the pyloric mucosa.

(d) They decrease blood pressure and protect coronary arteries.

(e) They increase renal blood flow and stimu­late renin production.

(f) They are inhibited by hyperlipemia, vit. E deficiency and radiation.

Thromboxane’s:

(a) They contract smooth muscles on blood vessels, GI Tract, uterus, bronchioles.

(b) They are discovered in platelets, and have the cyclopentane ring interrupted with an oxygen atom (Oxane ring).

(c) The substituent groups attached to the rings being varied give rise to different types in each series of thromboxane’s la­belled A, B, etc.

(d) They produce vasoconstriction and in­crease blood pressure.

(e) They cause release of serotonin and cal­cium ion (Ca⁺⁺) from platelet granules.

(f) Imidazole’s inhibit their synthesis.

Leukotriene’s:

(a) They are the third group of eicosanoid de­rivatives formed via the lipoxygenase pathway rather than cyclization of the fatty acid chain.

(b) They are first described in leukocytes.

(c) They are characterized by the presence of three conjugated double bonds.

(d) They are stimulators of mucus secretion and are responsible for vasoconstriction of bronchial muscles.

(e) They are inhibited by prolonged use of aspirin.

The group of compounds known as prostaglandins are synthesized from arachidonic acid in the body. They have pharmacologic and biochemical activity.

C. Many Other Fatty Acids:

(i) These have been detected in biologic ma­terial.

Example:

Fish oil contain 5 and 6 un­saturated fatty acids having carbon atoms 22.

(ii) Various other structures with hydroxy groups (ricinoleic acid) or cyclic groups have been found in nature.

Example of cyclic groups is chaulmoogric acid which was used many years ago in the treatment of leprosy.

Essential Fatty Acids:

Burr and Burr (1930) introduced the term “Essen­tial Fatty Acids” (EFA) on the basis that they are essential for the growth and health of young albino rats. These polyunsaturated fatty acids which are not synthesized in the body but are taken from natu­ral sources are called essential fatty acids.

They are (mentioned above):

Linolenic and arachidonic acids are formed from linoleic acids provided linoleic acids are avail­able in the body in sufficient quantities.

Properties:

(i) The essential fatty acids of vegetable oils have low melting points and iodine number.

(ii) They become saturated fatty acids on hydrogenation and the oils become solid fats.

Functions:

a. The essential fatty acids in high concen­tration along with the lipids constitute the structural elements of the tissues.

b. The lipids of gonads also contain a high concentration of polyunsaturated fatty acids which suggest the importance of re­productive function.

c. They effect the prolongation of clotting time and increase the fibrinolytic activity.

d. They retard atherosclerosis being esterified and emulsified with cholesterol and are incorporated into lipoproteins for transport to the liver for further oxidation.

e. They cure skin lesions.

f. The deficiency of these acids in the diet of babies causes eczema.

Isomerism in Unsaturated Fatty Acids:

Variations in the locations of the double bond in unsaturated fatty acid chains produce isomers. Oleic acid has 15 different positional isomers. Geometric isomerism depends on the orienta­tion of radicals around the axis of double bonds. If the radicals which are being considered are on the same side of the bond, the compound is called “cis”, if on opposite side, “trans”. This can be illustrated with maleic acid and fumaric acid.

There are more geometric isomers in case of acids with greater degree of unsaturation. The un­saturated long chain of fatty acids occurring in na­ture are nearly all in the ‘cis’ form and the mol­ecules are “bent” at the position of the double bond. Thus, arachidonic acid is U-shaped.

Refined and Hydrogenated Oils:

Refined oil:

It is prepared in the following man­ner:

(i) Free fatty acids are removed by alkali treat­ment.

(ii) Colouring matter is removed by activated carbon.

(iii) Odour is removed by superheated steam.

Hydrogenated oils:

The refined oils are hydrogenated under optimum temperature and pres­sure with hydrogen in the presence of nickel cata­lyst. Unsaturated fatty acids are converted into satu­rated fatty acids.

Oleic acid Stearic acid.

The liquid oil becomes solid fat and the un­saturated fatty acid content decreases. Vanaspati is hydrogenated refined groundnut oil.

2. Alcohols:

Alcohols found in lipid molecules include glyc­erol, cholesterol and higher alcohols (acetyl alco­hol), usually found in the waxes.

The unsaturated alcohols are important pig­ments. Phytyl alcohol is a constituent of chloro­phyll and lycophyll (C₄₀H₅₆O₂); a polyunsaturated dihydroxy alcohol occurs in tomatoes as a purple pigment.

Steroids:

The steroids are often found in association with fat. They have a similar cyclic nucleus resembling phenanthrene (rings A, B, C) to which a cyclopentane ring (D) is attached. The parent substance is better designated as cyclopentano perhydrophenanthrene. The position on the steroid nucleus are numbered as shown in Fig. 4.17.

Methyl side chains occur typically at posi­tions 10 and 13 (constituting C atoms 19 and 18). A side chain at position 17 is usual (as in choles­terol). If the compound has one or more hydroxyl groups and no carbonyl or carboxyl groups, it is a sterol, and the name terminates in -OL.

Steroids may be divided in the following man­ner:

Sterols—cholesterol, ergosterol, coprosterol. Bile acids—Glycocholic acid and taurocholic acid.

Sex hormones—Testosterone, Estradiol.

Vitamin D—Vit. D₂ and D₃.

Adrenocortical hormones—Corticosterone.

Cardiac glycosides—Stropanthin.

Saponins—Digitonin.

Cholesterol:

It is widely distributed in all cells of the body. It occurs in animal fats but not in plant fats. Its struc­ture is given below. The metabolism of cholesterol is discussed in the chapter of lipid metabolism.

Erogesrol:

(i) It occurs in ergot and yeast.

(ii) It is the precursor of vitamin D.

(iii) It acquires anti-rachitic properties with the opening of ring B when irradiated with ultraviolet light.

Coprosterol:

It occurs in feces as a result of the reduction by bacteria in the intestine of the double bond be­tween C₅ and C₆ of cholesterol.

Important tests:

1. Greese spot test:

A drop of oil placed over a piece of ordinary paper. A translucent spot is visible. This indicates the presence of fat.

2. Emulsification test:

2 ml water is taken in one test tube and 2 ml of diluted bile salt solution in another test tube. Add 3 drops of the given oil to each test tube and shake vigorously. Note the stability of the emul­sification formed.

3. Saponification test:

Take 10 drops of co­conut oil in a test tube. Add 20 drops of 40% NaOH and 2 ml of glycerol to it. Gen­tly boil for about 3 minutes until com­plete saponification occurs. If oil globules are visible, boiling must be continued. Di­vide the solution into 3 parts to carry the following experiments in test tube 1, 2, 3.

To test tube No. 1 add saturated solution of NaCl. Note that the soap separates out and floats to the surface (salting out process).

To test tube No. 2 add a few drops of conc. HCl. An oily layer of the fatty acids rises to the surface.

To test tube No. 3 add a few drops of CaCl₂ solution. The insoluble calcium soap is precipi­tated.

Unsaturation test:

Add 10 drops of Hubble’s iodine reagent to 10 ml of chloroform. The chloro­form assumes a pink colour due to the free iodine. The solution is divided equally into three test tubes as (a), (b) and (c) and three types of oil are added.

Add the oil No. 1 to the test tube (a) drop by drop shaking the tube vigorously after each addi­tion till the pink colour of the solution just disap­pears. The number of oil drops required is noted.

The experiment is repeated by adding oil 2 and 3 to test tubes (b) and (c), respectively. The more the number of drops required to discharge the pink colour, the less is the unsaturation.

Colour Reactions to Detect Sterols:

Liehermann-Rurehard Reaction:

A chloroform so­lution of a sterol when treated with acetic anhy­dride and sulphuric acid gives a green colour. This reaction is the basis of a colorimetric estimation of blood cholesterol.

Salkowski test:

A red to purple colour ap­pears when a chloroform solution of the sterol is treated with an equal volume of concentrated sul­phuric acid.

Clinical Orientation:

i. The high concentration of polyunsaturated fatty acids in the lipids of gonads are important in repro­ductive function.

ii. The essential fatty acid deficiency causes swelling of mitochondrial membrane resulting in the reduc­tion in efficiency of oxidative phosphorylation pro­ducing increased heat.

iii. Docosahexenoic acid formed from dietary linolenic acids enhances the electrical response of the photoreceptors to illumination. Therefore, linolenic acid of the diet is essential for optimal vision.

iv. The deficiency of essential fatty acids causes skin lesions, abnormal pregnancy and lactation in adult females, fatty liver, kidney damage.

v. The genetic deficiency of lecithin cholesterol acyl transferase (LCAT) causes Norum’s Disease.

vi. Sitosterol decreases the intestinal absorption of exogenous and endogenous cholesterol and thereby lowers the blood cholesterol level.

vii. The deficiency of the enzyme sphingomyelinase causes the large accumulations of sphingomyelins in brain, liver and spleen of children resulting in the Niemann-Pick. disease with the symptoms of en­larged abdomen, liver, spleen and mental deterio­ration.

viii. Absence of dipalmityl lecithin (DPL) in premature foetus produces respiratory distress syndrome (Hyaline-membrane disease).

ix. The inherited Gaucher’s Disease in infancy and childhood is caused by the deficiency of the en­zyme glucocerebrosidase involving the large ac­cumulations of glucocerebrosides (usually Kera­sin) in the liver, spleen, bone marrow, and brain with the manifestations of weight loss, failure in growth, and progressive mental retardation.

x. The autosomal recessive Tay-Sach’s Disease (GM₂ Gangliosidosis) results in the accumulation of large amounts of gangliosides in the brain and nervous tissues due to the absence of the enzyme hexosaminidase A with the association of progres­sive development of idiocy and blindness in infants soon after birth.

Clinical Orientation:

i. The inherited disorder Metachromatic Leukodys­trophy (MLD) happens on the sulfatide, formed from galactocerebroside, accumulation in various tis­sues owing to the deficiency of the enzyme sulfatase (Aryl sulfatase) with the symptoms of weakness, ataxia, defects in locomotion, paralysis, difficulties in speech in children before three years of age and psychiatric manifestation including progressive dementia in adults.

ii. Obesity and atherosclerosis are distinctly related to the concentrations of cholesterol and polyun­saturated fatty acids in the body.