lipids metabolism biochemistry..........

Education purpose

1. Metabolism of lipids. Metabolism of fatty acids. β-oxidation of fatty acids

2.   Plan of the lesson: 1. Classification of lipids. Classification of fatty acids. 2. Didestion and absorption of lipids. 3. Metabolism of fatty acids. 4. β-oxidation fatty acid: a ) . Fatty acid activation. b ). Transport of acyl -CoA into mitochondria. c ). β- oxidation proper 4 . Oxidation of palmitoyl CoA 5 . Energetic of β- oxidation . 6. Metabolism of ketone bodies.  

3. Functions of Lipids 1 . Storage form of energy (triglycerides) 2. Structural components of biomembranes ( phospholipids and cholesterol) 3. Metabolic regulators (steroid hormones and prostaglandins ) 4. Act as surfactants, detergents and emulsifying gents (amphipathic lipids) 5. Act as electric insulators in neurons 6. Provide insulation against changes in external temperature (subcutaneous fat) 7. Give shape and contour to the body 8. Protect internal organs by providing a cushioning effect (pads of fat) 9. Help in absorption of fat soluble vitamins ( A, D, E and K) 10. Improve taste and palatability of food.

4. SATURATED FATTY ACIDS 1. They have the general formula CH3- ( CH2)n-COOH . For example , Acetic acid - CH3—COOH Butyric acid - CH3(CH2)2—COOH Palmitic acid - CH3 —(CH2)14—COOH Stearic acid - CH3 —(CH2)16—COOH 2. The C16 ( palmitic acid) and C18 (stearic acid ) are most abundant in body fat. 3. Each animal species will have characteristic pattern of fatty acid composition. Thus human body fat contains 50% oleic acid, 25% palmitic acid 10% linoleic and 5% stearic acid . The carbon atoms of fatty acids are numbered as C1, C2, etc starting from the COOH group . Or , starting from the methyl end, the carbon atoms may be numbered as omega (ω)-1,2,3 , etc. 6 5 4 3 2 1 CH 3 — CH 2 — CH 2 — CH 2 — CH 2 — COOH ω1 ω2 ω3 ω4 ω5

5. UNSATURATED FATTY ACIDS Unsaturated fatty acids exhibit geometrical isomerism at the double bonds. All the naturally occurring fatty acids have the cisconfiguration . However , in the body during metabolism trans fatty acids are formed

6. SATURATED FATTY ACIDS They have the general formula CH3-(CH2)n-COOH. For example, Acetic acid - CH3—COOH Butyric acid - CH3(CH2)2—COOH Palmitic acid - CH3—(CH2)14—COOH Stearic acid - CH3—(CH2)16—COOH 2. The C 16 (palmitic acid) and C 18 (stearic acid) are most abundant in body fat. 3. Each animal species will have characteristic pattern of fatty acid composition. Thus human body fat contains 50% oleic acid, 25% palmitic acid 10% linoleic and 5% stearic acid. The carbon atoms of fatty acids are numbered as C 1 , C 2 , etc starting from the COOH group. Or, starting from the methyl end, the carbon atoms may be numbered as omega (ω)-1,2,3, etc. 6 5 4 3 2 1 CH 3 — CH 2 — CH 2 — CH 2 — CH 2 — COOH ω1 ω2 ω3 ω4 ω5

7. NEUTRAL FATS Neutral fats are also called as triacylglycerols (TAG ) or triglycerides (TG). These are esters of the trihydric alcohol , glycerol with fatty acids Storage of Energy as Fat The triacylglycerols are the storage form of lipids in the adipose tissue . In a 70 kg normal. Triacylglycerol (TAG) (triglyceride) person, body stores contain about 11 kg of triacyl glycerol , which is roughly equivalent to 100,000 kCal . If the same calories were stored as hydrated glycogen, the total weight of this alone would have been 65 kg. When stored as TAG , water molecules are repelled and space requirement is minimal. Excess fat in the body leads to obesity .

8. Properties of Fatty Acids 1. Hydrogenation Unsaturated fatty acids may be converted to the corresponding saturated fatty acids by hydrogenation of the double bond. (+) 2H (+)2H (+)2H Linolenic —→ Linoleic — → Oleic —→ Stearic Hydrogenation of oils can lead to solidification and saturation, e.g. Vanaspathi .   2. Halogenation When treated with halogens under mild conditions, the unsaturated fatty acids can take up two halogen atoms, at each double bond to form the halogenatedderivative of the fatty acid. For example, Oleic acid + I2 → Di- iodo oleic acid The number of halogen atoms taken up will depend on the number of double bonds and is an index of the degree of unsaturation.

9. 3. Melting Point The short and medium chain fatty acids are liquids, whereas long chain fatty acids are solids at 25 o C. The solubility in water decreases, while melting and boiling points increase, with increase in chain length. The unsaturated fatty acids have lower melting point compared to saturated fatty acids with the same chain length. For example, stearic acid (C18 fatty acid, no double bond) has the melting point 69 o C, oleic acid (C18, 1 double bond) has 13 o C; linoleic acid (C18, 2 double bonds) has –5 o C and linolenic (C18, 3 double bonds) has –10 o C.   4. Salt Formation Saturated and unsaturated fatty acids form salts with alkali. CH 3 —COOH + NaOH → CH 3 — COONa + H 2 O Sodium and potassium salts of long chain fatty acids are called soaps . Calcium and magnesium soaps are insoluble. Calcium soaps are used in grease. Alkyl sulfate (R—CH 2 —O—SO 2 — ONa ) and alkyl sulfonate (R—CH 2 —SO 2 —O—Na) are not precipitated by hard water and are used as detergents.

10. Complete hydrolysis of triglyceride. In the intestines, generally fats are only partially hydrolysed Digestion of Triglycerides 1. Pancreatic lipase can easily hydrolyse the fatty acids esterified to the 1st and 3rd carbon atoms of glycerol forming 2-monoacylglycerol and two molecules of fatty acid. 2. Then an isomerase shifts the ester bond from position 2 to 1. The bond in the 1st position is then hydrolysed by the lipase to form free glycerol and fatty acid. 3. The major end products of the digestion of TAG are 2-MAG (78%), 1-MAG (6%), glycerol and fatty acids (14%). Thus digestion of TAG is partial (incomplete). 4. Cholesterol ester may be hydrolysed to free cholesterol and fatty acid. The action of phospholipase A 2 produces lysophospholipid and a fatty acid.

11. Beta Oxidation Steps The next 4 reactions are sequentially repeated for complete oxidation of fatty acids. After one round of four metabolic steps, one acetyl CoA unit is split off and acyl CoA with 2 carbon atoms less is generated . This would undergo the same series of reactions again until the fatty acid is completely oxidised . Step 1: FAD Linked Dehydrogenase: The fatty acyl CoA is dehydrogenated to a trans enoyl CoA with the FAD accepting the hydrogen atoms. FADH2 when oxidised in electron transport chain will produce 1.5 ATP molecules. Step 2: Hydration: This is catalysed by an enoyl CoA hydratase . This step forms a beta- hydroxy fatty acyl CoA. The L isomer alone is formed during the hydration of the trans double bond . Step 3: NAD+ Dependent Dehydrogenase: The beta- hydroxy fatty acyl CoA is again oxidised to form beta- keto fatty acyl CoA. This dehydrogenase acts only on L isomer. The NADH when oxidised in electron transport chain will generate 2.5 ATPs . Step 4: Cleavage: The beta- keto fatty acyl CoA now undergoes thiolytic cleavage, splitting off a molecule of acetyl CoA and leaving behind a fatty acid with 2 carbon atoms less.

12. Energetics of Beta Oxidation (ATP Yield) Palmitic acid (16 C) needs 7 cycles of beta oxidation. So, it gives rise to 8 molecules of acetyl CoA. Every molecule of acetyl CoA when oxidised in the TCA cycle gives 10 molecules of ATP. Each molecule of FADH2 produces 1.5 molecules of ATP and each NADH generates 2.5 molecules of ATP, when oxidised in the electron transport chain. Hence the energy yield from one molecule of palmitate may be calculated as: 8 acetyl CoA × 10 = 80 ATP 7 FADH2 × 1.5 = 10.5 ATP 7 NADH × 2.5 = 17.5 ATP Gross total = 108 ATP Net yield = 108–2 = 106 ATP (In the initial activation reaction, the equivalents of 2 high energy bonds are utilised ). The efficiency of beta oxidation is about 33%. The differences in oxidation of unsaturated fatty acids are shown in Chapter 13. Important Note: In the previous editions of this textbook , calculations were made assuming that NADH produces 3 ATPs and FADH generates 2 ATPs. This will amount to a net generation of 129 ATP per palmitate molecule. Recent experiments show that these old values are overestimates , and net generation is only 106 ATPs.

13. Summary of beta oxidation of palmiticacid (16 C). It undergoes 7 cycles, which give rise to 8 molecules of acetyl CoA

14. METABOLISM OF KETONE BODIES Carbohydrates are essential for the metabolism of fat or fat is burned under the fire of carbohydrates . The acetyl CoA formed from fatty acids can enter and get oxidized in TCA cycle only when carbohydrates are available . During starvation and diabetes mellitus , the acetyl CoA takes the alternate fate of formation of ketone bodies . A . Ketogenesis Acetoacetate is the primary ketone body while beta- hydroxy butyrate and acetone are secondary ketone bodies. They are synthesised exclusively by the liver mitochondria . Step 1. Condensation Two molecules of acetyl CoA are condensed to form acetoacetyl CoA . Step 2. Production of HMG CoA One more acetyl CoA is added to acetoacetyl CoA to form HMG CoA (beta hydroxy beta methyl glutaryl CoA). The enzyme is HMG CoA synthase . Mitochondrial HMG CoA is used for ketogenesis , while cytosolic fraction is used for cholesterol synthesis.

15. Step 3. Lysis Then HMG CoA is lysed to form acetoacetate . Acetoacetate may also be formed by the degradation of carbon skeleton of ketogenic amino acids like leucine , lysine, phenylalanine and tyrosine . HMG CoA lyase is present only in liver . Step 4. Reduction Beta- hydroxy butyrate is formed by reduction of acetoacetate . Ratio between acetoacetate and beta hydroxy butyrate is decided by the cellular NAD:NADH ratio. Step 5. Spontaneous decarboxylation Acetone is formed.

16. Ketone body formation ( ketogenesis )

17. B. Ketolysis The ketone bodies are formed in the liver; but they are utilised by extrahepatic tissues . The heart muscle and renal cortex prefer the ketone bodies to glucose as fuel. Tissues like skeletal muscle and brain can also utilise the ketone bodies as alternate sources of energy, if glucose is not available . Acetoacetate is activated to acetoacetyl CoA by thiophorase enzyme . Thiophorase Acetoacetate —————- Acetoacetyl CoA + Succinyl CoA + Succinate

18. Formation, utilization and excretion of ketone bodies

19. KETOSIS Normally the rate of synthesis of ketone bodies by the liver is such that they can be easily metabolised by the extrahepatic tissues. Hence the blood level of ketone bodies is less than 1 mg/dl and only traces are excreted in urine ( not detectable by usual tests ). But when the rate of synthesis exceeds the ability of extrahepatic tissues to utilise them, there will be accumulation of ketone bodies in blood . 3. This leads to ketonemia , excretion in urine ( ketonuria ) and smell of acetone in breath. All these three together constitute the condition known as ketosis . 1 . Diabetes Mellitus: Uncontrolled diabetes mellitus is the most common cause for ketosis . Even though glucose is in plenty, the deficiency of insulin causes accelerated lipolysis and more fatty acids are released into circulation. Oxidation of these fatty acids increases the acetyl CoA pool . Enhanced gluconeogenesis restricts the oxidation of acetyl CoA by TCA cycle, since availability of oxaloacetate is less.

20. 2. Starvation: In starvation, the dietary supply of glucose is decreased. Available oxaloacetate israte of lipolysis is to provide alternate source of fuel. The excess acetyl CoA is converted to ketone bodies. The high glucagon level favors ketogenesis . The brain derives 60-75% of energy from ketone bodies under conditions of prolonged starvation. Hyperemesis (vomiting ) in early pregnancy may also lead to starvationlike condition and may lead to ketosis . Explanation for Ketogenesis 1. During starvation and diabetes mellitus, the blood level of glucagon is increased. Glucagon ( see Chapter 24) inhibits glycolysis, activates gluconeogenesis , activates lipolysis, decreases malonyl CoA level and stimulates ketogenesis . High glucagon–insulin ratio is potentially ketogenic . 2. Insulin (see Chapter 24) has the opposite effect ; it favors glycolysis, inhibits gluconeogenesis , depresses lipolysis, increases malonyl CoA level and decreases ketogenesis . The ketone body formation is regulated at the following 3 levels:

21. Level 1: Lipolysis Free fatty acids are the precursors of ketone bodies . So factors regulating the mobilization of fatty acid from adipose tissue will also control ketogenesis . Insulin inhibits lipolysis, while glucagon favors lipolysis. Level 2: Entry of Fatty Acid to Mitochondria The mobilized fatty acid then enters mitochondria for beta oxidation. Carnitine acyl transferase I (CATI ) regulates this entry. Malonyl CoA is the major regulator of CAT-I activity. In diabetes and starvation, glucagon is increased, which decreases malonyl CoA and so beta oxidation is stimulated. Level 3: Oxidation of Acetyl CoA 1. When the above two steps are increased, more acetyl CoA is produced. Normally, acetyl CoA is completely oxidized in the citric acid cycle. In diabetes and starvation, glucagon insulin ratio is increased, and key gluconeogenic enzymes are activated. 2 . When oxaloacetate is diverted for gluconeogenesis ; citric acid cycle cannot function optimally . Thus, on the one hand, acetyl CoA. Formation, utilization and excretion of ketone bodies is generated in excess, on the other hand, its utilization is reduced. This excess acetyl CoA is channeled into ketogenic pathway 3. In both diabetes mellitus and starvation, the oxaloacetate is channeled to gluconeogenesis ; so the availability of oxaloacetate is decreased . Hence acetyl CoA cannot be fully oxidized in the TCA cycle.

22. Summary of ketosis