Iso enzymes and their clinical significance

Isoenzymes are enzymes that catalyze identical chemical reactions but are composed of different amino acid sequences. They are sometimes referred to as isozymes. Isoenzymes are produced by different genes and are not redundant despite their similar functions. They occur in many tissues throughout the body and are important for different developmental and metabolic processes.

Isoenzymes are useful biochemical markers and can be measured in the bloodstream to diagnose medical conditions. Isoenzymes can be differentiated from one another using gel electrophoresis. In gel electrophoresis, isoenzyme fragments are drawn through a thick gel by an electric charge. Each isoenzyme has a distinct charge of its own because of its unique amino acid sequence. This enables gel electrophoresis to separate the fragments into bands for identification. Some clinically important isoenzymes are as follows

1) Creatine Kinase(CK, CPK) is an enzyme found primarily in the heart and skeletal muscles, and to a lesser extent in the brain but not found at all in liver and kidney. Small amounts are also found in lungs, thyroid and adrenal glands. Significant injuryto any of these structures will lead to a measurable increase in CK levels. It is not found in red blood cells and its level is not affected by hemolysis.

Normal Value- serum activity varies from 10-50 IU/L at 30°C.

Elevations are found in:

• Myocardial infarction
• Crushing muscular trauma
• Any cardiac or muscle disease, but not myasthenia gravis or multiple sclerosis
• Brain injury
• Hypothyroidism
• Hypokalemia

After myocardial infarction- serum value is found to increase within hours, reaches a peak level in 24- 30 hours and returns to normal level in 2-4 days (usually in 72 hours). CK is a sensitive indicator in the early stages of myocardial ischemia. No increase in activity is found in heart failure and coronary insufficiency. In acute MI, CPK usually rises faster than SGOT and returns to normal faster than the SGOT. 

CK/CPK Isoenzymes
There are three Isoenzymes. Measuring them is of value in the presence of elevated levels of CK or CPK to determine the source of the elevation. Each iso enzyme is a dimer composed of two protomers‘M’ (for muscles) and ‘B’( for Brain). These isoenzymes can be separated by, Electrophoresis or by Ion exchange Chromatography. The three possible iso enzymes are;

Isoenzyme	Electrophoretic mobility	Tissue of origin	Mean percentage in blood
MM(CK3)	Least	Skeletal muscle

Heart muscle97-100%MB(CK2)IntermediateHeart muscle0-3% BB(CK1)MaximumBrain0%

• Normal levels of CK/CPK are almost entirely MM, from skeletal muscle.
• Elevated levels of CK/CPK resulting from acute myocardial infarction are about half MM and half MB. Myocardial muscle is the only tissue that contains more than five percent of the total CK activity as the CK2 (MB) isoenzyme.
• Following an attack of acute myocardial infarction, this isoenzyme appears within 4 hours following onset of chest pain, reaches a peak of activity at approximately 24 hours and falls rapidly. MB accounts for 4.5- 20 % of the total CK activity in the plasma of the patients with recent myocardial infarction and the total isoenzyme is elevated up to 20-folds above the normal.
• Atypical Isoenzymes- In addition to the above three isoenzymes two atypical iso enzymes of CPK have been reported. They are; Macro CK(CK-Macro) and Mitochondrial CK(CK-Mi).

o  Macro CK(CK-Macro)- It is formed by the aggregation of CK-BB with immunoglobulins usually with IgG but sometimes Ig A.It may also be formed by complexing CK-MM with lipoproteins. No specific disease has been found to be associated with this isoenzyme.

o  Mitochondrial CK(CK-Mi)- It is presentbound to the exterior surface of the inner mitochondrial membrane of muscle,liver and brain. It can exist in dimeric form or as oligomeric aggregates having molecular weight of approximately 35,000. It is only present in serum when there is extensive tissue damage causing breakdown of mitochondrial membrane and cell membrane. Thus its presence in serum indicates severe illness and cellular damage. It is not related with any specific disease states but it has been detected in certain cases of malignant tumors.

2) Aspartate amino Transferase (AST)

It is also called as Serum Glutamate Oxalo acetate Transaminase (SGOT). The level is significantly elevated in Acute MI.

Normal Value- 0-41 IU/L at 37°C

In acute MI- Serum activity rises sharply within the first 12 hours, with a peak level at 24 hours or over and returns to normal within 3-5 days. The rise depends on the extent of infarction. Re- infarction results in secondary rise of SGOT.

Prognostic significance- Levels> 350 IU/L are due to massive Infarction(Fatal), 

> 150 IU/L are associated with high mortality and levels < 50IU/L are associated with low mortality.

Other diseases- The rise in activity is also observed in muscle and hepatic diseases. These can be well differentiated from simultaneous estimations of other enzyme activities like SGPT etc, which do not show and rise in activity in Acute MI.

3) Alanine amino transferase (ALT)- Also called serum Glutamate pyruvate transaminase.

Normal serum level ranges between 0-45 IU/L at37^oC.

Very high values are seen in Acute hepatitis,toxic or viral in origin. Both ALT and AST rise but ALT> AST. Moderate increase may be seen in chronic liver diseases such as Cirrhosis and Malignancy in liver. A sudden fall in AST level in hepatitis signifies bad prognosis.

4) Lactate dehydrogenase (LDH)

Lactate dehydrogenase catalyzes the reversible conversion of pyruvate and lactate. LDH is essential for anaerobic respiration. When oxygen levels are low,LDH converts pyruvate to lactate, providing a source of muscular energy.

Normal level-  55-140IU/L at 30°C. The levels in the upper range are generally seen in children. LDH level is 100 times more inside the RBCs than in plasma, and therefore minor amount of hemolysis results in false positive result.

In Acute MI-The serum activity rises within 12 to 24 hours, attains a peak at 48 hours (2 to 4 days) reaching about 1000 IU/L and then returns gradually to normal from 8 th to 14 th day. The magnitude of rise is proportional to the extent of myocardial infarction. Serum LDH elevation may persist for more than a week after CPK and SGOT levels have returned to normal levels.

Other diseases-The increase in serum activity of LDH is also seen in hemolytic anemia, hepatocellular damage, muscular dystrophies,carcinoma, leukemias, and any condition which causes necrosis of the body cells. Since the total LDH is increased in many diseases, so the study of Isoenzymes of LDH is of more significance.

Iso enzymes of LDH

LDH enzyme is tetramer with 4 subunits. The subunit may be either H(Heart) or M(Muscle) polypeptide chains. These two chains are the product of 2 different genes. Although both of them have the same molecular weight, there are minor amino acid variations.There can be 5 possible combinations; H4, H3M1, H2M2, H1M3. M4, these are 5different types of isoenzymes seen in all individuals.

No. of Isoenzyme	Subunit make up of isoenzyme	Electrophoretic mobility at pH8.6	Activity at 60°for 30 minutes	Tissue origin	Percentage in human serum(Mean)
LDH-1	H4	Fastest	Not destroyed	Heart muscle	30%
LDH-2	H3M1	Faster	Not destroyed.	RBC	35%
LDH-3	H2M2	Fast	Partly destroyed	Brain	20%
LDH-4	H1M3	Slow	Destroyed	Liver	10%
LDH-5	M4	Slowest	Destroyed	Skeletal Muscles	5%

Normally LDH- 2(H3M1) level in blood is greater than LDH-1, but this pattern is reversed in myocardial infarction, this is called ‘flipped pattern’. These iso enzymes are separated by cellulose acetate electrophoresis at pH 8.6.

5) Alkalinephosphatase (ALP )-is an enzyme that removes phosphate groups  from organic or inorganic compounds in the body. It is present in a number of tissues including liver, bone, intestine,and placenta. The activity of ALP found in serum is a composite of isoenzymes from those sites and, in some circumstances, placental or Regan isoenzymes. The optimum pH for enzyme action varies between 9-10. It is a zinc containing metalloenzyme and is localized in the cell membranes (ectoenzyme). It is associated with transport mechanism in the liver, kidney and intestinal mucosa.

Normal serum Level- of ALP ranges between 40-125 IU/L. In children the upper level of normal value may be more, because of increased osteoblastic activity.

Total Alkaline Phosphatase (ALP)

Serum ALP is of interest in the diagnosis of 2 main groups of conditions-hepatobiliary diseases and bone diseases associated with increased osteoblastic activity.  

Mild increase is observed in pregnancy, due to production of placental enzyme.

A moderate rise in ALP activity occurs inhepatic diseases such as infective hepatitis, alcoholic hepatitis or hepatocellular carcinoma. Moderate elevation of ALP may also be seen in other disorders such as Hodgkin’s disease, congestive heart failure, ulcerative colitis, regional enteritis, and intra-abdominal bacterial infections.

ALP elevations tend to be more marked (more than 3-fold) in extra hepatic biliary obstructions (eg, by stone or cancer of the head of the pancreas) than in intrahepatic obstructions, and the more complete the obstruction, the greater the elevation. With obstruction, serumALP activities may reach 10 to 12 times the upper limit of normal, returning to normal upon surgical removal of the obstruction. The ALP response to cholestatic liver disease is similar to the response o fgamma-glutamyltransferase (GGT), but more blunted. If both GGT and ALP are elevated, a liver source of the ALP is likely.  
The response of the liver to any form of biliary tree obstruction is to synthesize more ALP. The main site of new enzyme synthesis is the hepatocytes adjacent to the biliary canaliculi.

ALP also is elevated in disorders of the skeletal system that involve osteoblast hyperactivity and bone remodeling, suchas Paget’s disease rickets and osteomalacia, fractures, and malignant tumors.

Among bone diseases, the highest level o f ALP activity is encountered in Paget’s disease, as a result of the action of the osteoblastic cells as they try to rebuild bone that is being resorbed by the uncontrolled activity of osteoclasts. Values from 10 to 25 times the upper limit of normal are not unusual. 

Only moderate rises are observed in osteomalacia, while levels are generally normal in osteoporosis. In rickets,levels 2 to 4 times normal may be observed. Primary and secondary hyperparathyroidism are associated with slight to moderate elevations of ALP;the existence and degree of elevation reflects the presence and extent of skeletal involvement. 
Very high enzyme levels are present in patients with osteogenic bone cancer. 

A considerable rise in ALP is seen in children following accelerated bone growth.

Patients over age 60 can have a mildly elevated alkaline phosphatase (1–1½ times normal), while individuals with blood types O and B can have an elevation of the serum alkaline phosphatase after eating a fatty meal due to the influx of intestinal alkaline phosphatase into the blood.

ALP Isoenzymes

Electrophoresis is considered the most useful single technique for ALP isoenzyme analysis. By starch gel electrophoresis at pH 8.6 , at least 6 isoenzyme bands can be visualized.

1) Hepatic ALP isoenzyme- moves fastest towards the anode and occupies the same position as  Alpha 2 globulins. It is associated with biliary epithelium and is elevated in cholestatic processes. Various liver diseases (primary or secondary cancer, biliary obstruction) increase the liver isoenzyme.

2) Bone isoenzyme- It closely follows the hepatic enzyme and occupies the beta region. Osteoblastic bone tumors and hyperactivity of osteoblasts involved in bone remodeling (eg, Paget’s disease)increase the bone isoenzyme. Paget’s disease leads to a striking, solitary elevation of bone ALP.

3) Placental isoenzyme follows bone isoenzyme. It is heat stable isoenzyme and increases during last six weeks of pregnancy.

4) The intestinal isoenzyme-is the slowly moving band and follows the placental isoenzyme. It may be increased in patients with cirrhosis and in individuals who are blood group O or B secretors. Increased levels are also sen in patients undergoing hemodialysis.
  
Atypical ALP isoenzymes (Oncogenic markers)-In addition to 4 major isoenzymes, 2 more abnormal fractions are seen associated with tumors. These are Regan and Nagao isoenzymes. They are also called “Carcino placental ALP”, as they resemble placental isoenzymes.
Regan isoenzyme is elevated in various carcinomas of breast, lungs, colon and ovaries. Highest positivity is observed in carcinoma of ovary and uterus.

Rise in Nagao isoenzyme is observed in metastatic carcinoma of pleural surfaces and adenocarcinoma of pancreas and bile duct.

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Isoenzymes

Isoenzymes are enzymes that catalyze identical chemical reactions but are composed of different amino acid sequences. They are sometimes referred to as isozymes. Isoenzymes are produced by different genes and are not redundant despite their similar functions. They occur in many tissues throughout the body and are important for different developmental and metabolic processes.

Isoenzymes are useful biochemical markers and can be measured in the bloodstream to diagnose medical conditions. Isoenzymes can be differentiated from one another using gel electrophoresis. In gel electrophoresis, isoenzyme fragments are drawn through a thick gel by an electric charge. Each isoenzyme has a distinct charge of its own because of its unique amino acid sequence. This enables gel electrophoresis to separate the fragments into bands for identification. Some clinically important isoenzymes are as follows

1) Creatine Kinase(CK, CPK) is an enzyme found primarily in the heart and skeletal muscles, and to a lesser extent in the brain but not found at all in liver and kidney. Small amounts are also found in lungs, thyroid and adrenal glands. Significant injuryto any of these structures will lead to a measurable increase in CK levels. It is not found in red blood cells and its level is not affected by hemolysis.

Normal Value- serum activity varies from 10-50 IU/L at 30°C.

Elevations are found in:

Myocardial infarction
Crushing muscular trauma
Any cardiac or muscle disease, but not myasthenia gravis or multiple sclerosis
Brain injury
Hypothyroidism
Hypokalemia
After myocardial infarction- serum value is found to increase within hours, reaches a peak level in 24- 30 hours and returns to normal level in 2-4 days (usually in 72 hours). CK is a sensitive indicator in the early stages of myocardial ischemia. No increase in activity is found in heart failure and coronary insufficiency. In acute MI, CPK usually rises faster than SGOT and returns to normal faster than the SGOT.

CK/CPK Isoenzymes
There are three Isoenzymes. Measuring them is of value in the presence of elevated levels of CK or CPK to determine the source of the elevation. Each iso enzyme is a dimer composed of two protomers‘M’ (for muscles) and ‘B’( for Brain). These isoenzymes can be separated by, Electrophoresis or by Ion exchange Chromatography. The three possible iso enzymes are;

Isoenzyme Electrophoretic mobility Tissue of origin Mean percentage in blood
MM(CK3) Least Skeletal muscle
Heart muscle97-100%MB(CK2)IntermediateHeart muscle0-3% BB(CK1)MaximumBrain0%

Normal levels of CK/CPK are almost entirely MM, from skeletal muscle.
Elevated levels of CK/CPK resulting from acute myocardial infarction are about half MM and half MB. Myocardial muscle is the only tissue that contains more than five percent of the total CK activity as the CK2 (MB) isoenzyme.
Following an attack of acute myocardial infarction, this isoenzyme appears within 4 hours following onset of chest pain, reaches a peak of activity at approximately 24 hours and falls rapidly. MB accounts for 4.5- 20 % of the total CK activity in the plasma of the patients with recent myocardial infarction and the total isoenzyme is elevated up to 20-folds above the normal.
Atypical Isoenzymes- In addition to the above three isoenzymes two atypical iso enzymes of CPK have been reported. They are; Macro CK(CK-Macro) and Mitochondrial CK(CK-Mi).
o  Macro CK(CK-Macro)- It is formed by the aggregation of CK-BB with immunoglobulins usually with IgG but sometimes Ig A.It may also be formed by complexing CK-MM with lipoproteins. No specific disease has been found to be associated with this isoenzyme.

o  Mitochondrial CK(CK-Mi)- It is presentbound to the exterior surface of the inner mitochondrial membrane of muscle,liver and brain. It can exist in dimeric form or as oligomeric aggregates having molecular weight of approximately 35,000. It is only present in serum when there is extensive tissue damage causing breakdown of mitochondrial membrane and cell membrane. Thus its presence in serum indicates severe illness and cellular damage. It is not related with any specific disease states but it has been detected in certain cases of malignant tumors.

2) Aspartate amino Transferase (AST)

It is also called as Serum Glutamate Oxalo acetate Transaminase (SGOT). The level is significantly elevated in Acute MI.

Normal Value- 0-41 IU/L at 37°C

In acute MI- Serum activity rises sharply within the first 12 hours, with a peak level at 24 hours or over and returns to normal within 3-5 days. The rise depends on the extent of infarction. Re- infarction results in secondary rise of SGOT.

Prognostic significance- Levels> 350 IU/L are due to massive Infarction(Fatal),

> 150 IU/L are associated with high mortality and levels < 50IU/L are associated with low mortality.

Other diseases- The rise in activity is also observed in muscle and hepatic diseases. These can be well differentiated from simultaneous estimations of other enzyme activities like SGPT etc, which do not show and rise in activity in Acute MI.

3) Alanine amino transferase (ALT)- Also called serum Glutamate pyruvate transaminase.

Normal serum level ranges between 0-45 IU/L at37oC.

Very high values are seen in Acute hepatitis,toxic or viral in origin. Both ALT and AST rise but ALT> AST. Moderate increase may be seen in chronic liver diseases such as Cirrhosis and Malignancy in liver. A sudden fall in AST level in hepatitis signifies bad prognosis.

4) Lactate dehydrogenase (LDH)

Lactate dehydrogenase catalyzes the reversible conversion of pyruvate and lactate. LDH is essential for anaerobic respiration. When oxygen levels are low,LDH converts pyruvate to lactate, providing a source of muscular energy.

Normal level-  55-140IU/L at 30°C. The levels in the upper range are generally seen in children. LDH level is 100 times more inside the RBCs than in plasma, and therefore minor amount of hemolysis results in false positive result.

In Acute MI-The serum activity rises within 12 to 24 hours, attains a peak at 48 hours (2 to 4 days) reaching about 1000 IU/L and then returns gradually to normal from 8 th to 14 th day. The magnitude of rise is proportional to the extent of myocardial infarction. Serum LDH elevation may persist for more than a week after CPK and SGOT levels have returned to normal levels.

Other diseases-The increase in serum activity of LDH is also seen in hemolytic anemia, hepatocellular damage, muscular dystrophies,carcinoma, leukemias, and any condition which causes necrosis of the body cells. Since the total LDH is increased in many diseases, so the study of Isoenzymes of LDH is of more significance.

Iso enzymes of LDH

LDH enzyme is tetramer with 4 subunits. The subunit may be either H(Heart) or M(Muscle) polypeptide chains. These two chains are the product of 2 different genes. Although both of them have the same molecular weight, there are minor amino acid variations.There can be 5 possible combinations; H4, H3M1, H2M2, H1M3. M4, these are 5different types of isoenzymes seen in all individuals.

No. of Isoenzyme Subunit make up of isoenzyme Electrophoretic mobility at pH8.6 Activity at 60°for 30 minutes Tissue origin Percentage in human serum(Mean)
LDH-1 H4 Fastest Not destroyed Heart muscle 30%
LDH-2 H3M1 Faster Not destroyed. RBC 35%
LDH-3 H2M2 Fast Partly destroyed Brain 20%
LDH-4 H1M3 Slow Destroyed Liver 10%
LDH-5 M4 Slowest Destroyed Skeletal Muscles 5%
Normally LDH- 2(H3M1) level in blood is greater than LDH-1, but this pattern is reversed in myocardial infarction, this is called ‘flipped pattern’. These iso enzymes are separated by cellulose acetate electrophoresis at pH 8.6.

5) Alkalinephosphatase (ALP )-is an enzyme that removes phosphate groups  from organic or inorganic compounds in the body. It is present in a number of tissues including liver, bone, intestine,and placenta. The activity of ALP found in serum is a composite of isoenzymes from those sites and, in some circumstances, placental or Regan isoenzymes. The optimum pH for enzyme action varies between 9-10. It is a zinc containing metalloenzyme and is localized in the cell membranes (ectoenzyme). It is associated with transport mechanism in the liver, kidney and intestinal mucosa.



Normal serum Level- of ALP ranges between 40-125 IU/L. In children the upper level of normal value may be more, because of increased osteoblastic activity.

Total Alkaline Phosphatase (ALP)

Serum ALP is of interest in the diagnosis of 2 main groups of conditions-hepatobiliary diseases and bone diseases associated with increased osteoblastic activity.

Mild increase is observed in pregnancy, due to production of placental enzyme.

A moderate rise in ALP activity occurs inhepatic diseases such as infective hepatitis, alcoholic hepatitis or hepatocellular carcinoma. Moderate elevation of ALP may also be seen in other disorders such as Hodgkin’s disease, congestive heart failure, ulcerative colitis, regional enteritis, and intra-abdominal bacterial infections.

ALP elevations tend to be more marked (more than 3-fold) in extra hepatic biliary obstructions (eg, by stone or cancer of the head of the pancreas) than in intrahepatic obstructions, and the more complete the obstruction, the greater the elevation. With obstruction, serumALP activities may reach 10 to 12 times the upper limit of normal, returning to normal upon surgical removal of the obstruction. The ALP response to cholestatic liver disease is similar to the response o fgamma-glutamyltransferase (GGT), but more blunted. If both GGT and ALP are elevated, a liver source of the ALP is likely.
The response of the liver to any form of biliary tree obstruction is to synthesize more ALP. The main site of new enzyme synthesis is the hepatocytes adjacent to the biliary canaliculi.

ALP also is elevated in disorders of the skeletal system that involve osteoblast hyperactivity and bone remodeling, suchas Paget’s disease rickets and osteomalacia, fractures, and malignant tumors.

Among bone diseases, the highest level o f ALP activity is encountered in Paget’s disease, as a result of the action of the osteoblastic cells as they try to rebuild bone that is being resorbed by the uncontrolled activity of osteoclasts. Values from 10 to 25 times the upper limit of normal are not unusual.

Only moderate rises are observed in osteomalacia, while levels are generally normal in osteoporosis. In rickets,levels 2 to 4 times normal may be observed. Primary and secondary hyperparathyroidism are associated with slight to moderate elevations of ALP;the existence and degree of elevation reflects the presence and extent of skeletal involvement.
Very high enzyme levels are present in patients with osteogenic bone cancer.

A considerable rise in ALP is seen in children following accelerated bone growth.

Patients over age 60 can have a mildly elevated alkaline phosphatase (1–1½ times normal), while individuals with blood types O and B can have an elevation of the serum alkaline phosphatase after eating a fatty meal due to the influx of intestinal alkaline phosphatase into the blood.

ALP Isoenzymes

Electrophoresis is considered the most useful single technique for ALP isoenzyme analysis. By starch gel electrophoresis at pH 8.6 , at least 6 isoenzyme bands can be visualized.

1) Hepatic ALP isoenzyme- moves fastest towards the anode and occupies the same position as  Alpha 2 globulins. It is associated with biliary epithelium and is elevated in cholestatic processes. Various liver diseases (primary or secondary cancer, biliary obstruction) increase the liver isoenzyme.

2) Bone isoenzyme- It closely follows the hepatic enzyme and occupies the beta region. Osteoblastic bone tumors and hyperactivity of osteoblasts involved in bone remodeling (eg, Paget’s disease)increase the bone isoenzyme. Paget’s disease leads to a striking, solitary elevation of bone ALP.

3) Placental isoenzyme follows bone isoenzyme. It is heat stable isoenzyme and increases during last six weeks of pregnancy.

4) The intestinal isoenzyme-is the slowly moving band and follows the placental isoenzyme. It may be increased in patients with cirrhosis and in individuals who are blood group O or B secretors. Increased levels are also sen in patients undergoing hemodialysis.
 
Atypical ALP isoenzymes (Oncogenic markers)-In addition to 4 major isoenzymes, 2 more abnormal fractions are seen associated with tumors. These are Regan and Nagao isoenzymes. They are also called “Carcino placental ALP”, as they resemble placental isoenzymes.
Regan isoenzyme is elevated in various carcinomas of breast, lungs, colon and ovaries. Highest positivity is observed in carcinoma of ovary and uterus.

Rise in Nagao isoenzyme is observed in metastatic carcinoma of pleural surfaces and adenocarcinoma of pancreas and bile duct.



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Enzyme Inhibition

Type of Inhibition	Effect on Maximum Reaction Velocity(Vmax)	Effect on Km(Affinity of Enzyme for its substrate)	Reversible/ Irreversible	Examples
Competitive Inhibition  (Inhibitor- a structural Analogue). There is a competition between  substrate and the inhibitor for the active site	Unchanged	Increased	Reversible	1) HMG Co A Reductase- inhibited by Statins- used as cholesterol lowering drugs.  2) Epoxide Reductase – inhibited by Dicoumarol- used as an anticoagulant. 3) Dihydrofolate Reductase-inhibited by Methotrexate- used as anticancer drug. 4) Pteroyl Synthase- inhibited by PABA (Para- amino –benzoic –acid) used as antibiotic. 5) Angiotensin convertase enzyme Inhibitor- inhibited by Captopril –used as an antihypertensive drug. 6) Succinate dehydrogenase- inhibited by Malonate acts as a poison. 7) Lactate dehydrogenase- inhibited by Oxamate- acts as a poison.
Non competitive(Inhibitor binds at a site other than the active site)	Decreased	Unchanged	Can be reversible or irreversible	1) Enolase is inhibited by Fluoride used for sample collection for glucose estimation.  2) PDH complex, Alpha ketoglutarate dehydrogenase complex, glyceraldehyde-3-P dehydrogenase(-SH group containing enzymes) are inhibited by Arsenate, Acts as a slow poison 3) Cytochrome oxidase- Inhibited by cyanide acts as a poison. 4)Cyclooxygenase is inhibited by Aspirin-used as an anti inflammatory, analgesic and antipyretic  drug (Inhibits formation of prostaglandins)
Suicidal inhibition,   Also calledMechanism based  inhibition  Inhibitor gets activated by host enzyme to inhibit the subsequent enzyme	Decreased	Increased	Irreversible	1)Inhibition of Xanthine oxidase by Allopurinol, used for the treatment of gout.  2) Inhibition of Aconitase by fluoroacetate- used as a rat poison 3) Inhibition of Transpeptidase by Penicillin- used as an antibiotic 4) Inhibition of Ornithine decarboxylase by Di fluoro methyl ornithine(DFMO) 5) Mono Amine Oxidase is inhibited by Deprenyl used to treat Parkinson disease and depression.
Allosteric Inhibition (Binding of the inhibitor alters either the affinity of the enzyme for its substrate or the reaction velocity is decreased	V max is decreased in V type enzymes	Km is increased in K type enzymes	Reversible	Most of them are physiological inhibitors.   1)ATP and Citrate are allosteric inhibitors of PFK-1 2) Fr 2,6 bisphosphate is an allosteric inhibitor of Fr1,6 bisphosphatase enzyme.
Feed back inhibition, also called Product inhibition The product of the reaction pathway inhibits the key  regulatory enzyme.	Decreased	constant	Reversible depending upon the need of the product	1) Inhibition of HMG Co A Reductase by Mevalonate(Immediate product) and cholesterol(Final product)  2) Inhibition of Aspartate transcarbamoylase by UTP and CTP, the products of this pathway.
Un competitive inhibition Inhibitor binds to the enzyme substrate complex	Decreased	Decreased	Ir- reversible	Inhibition of  placental alkaline phosphatase by Phenyl Alanine

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Classification of enzymes

Classification of enzymes- More than 2000 different enzymes are currently known. The commonly used names for most enzymes describe the type of reaction catalyzed, followed by the suffix -ase. For example, dehydrogenases remove hydrogen atoms, proteases hydrolyze proteins, and isomerases catalyze rearrangements in configuration. Modifiers may precede the name to indicate the substrate (xanthine oxidase), the source of the enzyme (pancreatic ribonuclease), its regulation (hormone-sensitive lipase) etc.

To address ambiguities, the International Union of Biochemists (IUB) developed an unambiguous system of enzyme nomenclature in which each enzyme has a unique name and code number that identify the type of reaction catalyzed and the substrates involved. Enzymes are grouped into six classes:

1) The oxidoreductases (class 1) catalyze the transfer of reducing equivalents(Hydrogen and electrons)from one redox system to another.

2) The transferases (class 2) catalyze the transfer of other groups from one molecule to another. Oxidoreductases and transferases  generally require coenzymes

3) The hydrolases (class 3) hydrolases cause cleavage of bond using water

4) Lyases (class 4, often also referred to as“synthases”) catalyze reactions involving either the cleavage or formation of chemical bonds, with double bonds either arising or disappearing.(See figure- reversible reaction is shown). Cleavage of bond does not require water.

5) The isomerases (class 5) move groups within a molecule, without changing the gross composition of the substrate.

6) The ligation reactions catalyzed by ligases (“synthetases,” class 6) are energy-dependent and are therefore always coupled to the hydrolysis of nucleoside triphosphates(See figure)

Each enzyme is entered in the Enzyme Catalogue with a four-digit Enzyme Commission number (EC number). The first digit indicates membership of one of the six major classes. The next two indicate subclasses and subsubclasses. The last digit indicates where the enzyme belongs in the subsubclass.

For example, The IUB name of hexokinase is ATP:D-hexose 6-phosphotransferase E.C. 2.7.1.1. This name identifies hexokinase as a member of class 2 (transferases), subclass 7 (transfer of a phosphoryl group), sub-subclass 1 (alcohol is the phosphoryl acceptor), and “hexose-6″ indicates that the alcohol phosphorylated is on carbon six of a hexose. However, it is still called as hexokinase.

Figure- Showing the classification of enzymes with examples of each class of enzymes

 

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Subjective Questions- Haem synthesis and degradation

Q.1- Describe the fate of the hemoglobin molecules when senescent erythrocytes are broken down in the reticuloendothelial system. Explain what is meant by pre-hepatic, hepatic and post-hepatic jaundice. Give causes, laboratory findings and treatment of each of the above conditions.

Q.2- Explain the biochemical basis of Porphyria. Classify and  discuss two clinical condition in which different metabolites of porphyrins accumulate  to cause cutaneous hypersensitivity.

Q.3- What is the cause of neonatal jaundice? Explain the basis of blue light phototherapy of this condition.

Q.4- Explain how biliary obstruction could impair blood coagulation.

Q.5-Discuss the different clinical states of hyperbilirubinemia (congenital or acquired) caused due to impaired activity of UDP Glucuronyl transferase enzyme.

Q.6- Describe the biochemical basis and outline the laboratory tests for (a) obstructive jaundice; (b) acute intermittent Porphyria.

Q.7-(a) Explain the principle of measurement of total and free bilirubin by the van den Bergh reaction (b) Explain the diagnostic value in determining serum bilirubin.

Q.8- Explain how lead poisoning affects haem synthesis.

Q.9- From your knowledge of porphyrin metabolism, explain the justification for the current introduction of unleaded petrol.

Q.10- How is the biosynthesis of haem normally regulated?

Q.11- Explain why measurements of both bilirubin and urobilin are relevant to the diagnosis of obstructive jaundice?

Q.12- In a person with a defect in bilirubin metabolism, what can you deduce from the following laboratory results: (a) Bilirubin in urine and decreased urinary urobilinogen. (b) Absence of bilirubin in urine and increased urinary urobilinogen.

Q.13- In patients with a deficiency of coproporphyrinogen oxidase, what abnormalities in porphyrin metabolites would you expect to see?

Q.14- A rational approach to the diagnosis and treatment of jaundice rests on a good knowledge of the biochemistry of bilirubin metabolism. Discuss.

Q.15- What do you understand by the terms “direct” and “indirect”" Van Den Bergh reactions?

Q.16- Discuss briefly the diagnostic value of measuring stercobilin and or urobilin.

Q.17- Explain the use of biochemical tests to confirm the diagnosis of hepatocellular jaundice.

Q.18- Give the reactions catalysed by (a) haem oxygenase (b) ALA synthase (c) Ferrochelatase

Q.19- What is the biochemical basis of cutaneous hypersensitivity in Congenital Erythropoetic Porphyria?

Q.20- What is the biochemical basis of giving Hematin or Glucose infusion to treat acute attack of Porphyria?

Q.21- Discuss the basis of breast milk jaundice.

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Multiple Choice Questions- Haem synthesis and Degradation

Q.1- Which out of the followings is not a haemo protein?

a) Tryptophan pyrrolase

b) Tyrosinase

c) Myoglobin

d) Cytochrome P450

Q.2- Which out of the following enzymes catalyses a rate limiting step in the pathway of haem biosynthesis?

a) ALA synthase

b) ALA dehydratase

c) PBG deaminase

d) Coproporphrinogen oxidase

Q.3- High levels of lead can affect heme metabolism by combining with SH groups of which out the following enzymes?

a) ALA synthase

b) ALA dehydratase

c) PBG deaminase

d) Coproporphrinogen oxidase

Q.4- Pyridoxal phosphate is necessary in the pathway of Haem biosynthesis, which out of the following enzymes requires Pyridoxal –P as a coenzyme?

a) ALA synthase

b) ALA dehydratase

c) PBG deaminase

d) Ferrochelatase

Q.5- In general, the porphyrias are inherited in an autosomal dominant manner, with the exception of

a) Acute intermittent porphyria

b) Porphyria Cutanea Tarda

c) Variegate Porphyria

d) Congenital Erythropoietic porphyria

Q.6- Choose the incorrect statement out of the followings-

a) Synthesis of ALA occurs in the mitochondria

b) Uroporphyrinogen formed is almost exclusively the III isomer

c) A porphyrin with symmetric substitution of side chains is classified as a type III porphyrin

d) Coproporphyrinogen oxidase is able to act only on type III isomers

Q.7- In which of the following porphyrias, cutaneous hypersensitivity is not observed?

a) Variegate porphyria

b) Acute intermittent porphyria

c) Congenital Erythropoietic porphyria

d) Hereditary Coproporphyria

Q.8-Which of the following statements describes the basis of giving I/V Hemin infusion?

a) Haem/ Hemin acts as a negative regulator of the synthesis of ALA synthase

b) Heme affects translation of the enzyme

c) Heme affects transfer of enzyme from the cytosol to the mitochondrion

d) All of the above

Q.9- Porphyrins are deposited in teeth and in bones, as a result, the teeth are reddish-brown and fluoresce on exposure to long-wave ultraviolet light, so called ‘Erythtrodontia ‘, is a sign of which porphyria ?

a) Variegate porphyria

b) Acute intermittent porphyria

c) Congenital Erythropoietic porphyria

d) Hereditary Coproporphyria

Q.10-A 24- year- old patient was brought to medical OPD with acute abdominal pain, depression and extreme weakness.  Urine analysis revealed the presence of ALA and PBG (Delta amino Levulinic acid and Porphobilinogen).   The patient was diagnosed with acute intermittent porphyria, which of the following enzyme deficiencies is expected in this patient?

a) Uroporphyrinogen III cosynthase

b) Uroporphyrinogen decarboxylase

c) Porphobilinogen decarboxylase

d) None of the above.

Q.11-An 8 year old boy was brought to a dermatologist as he had developed vesicles and bullae on his face and arms that appeared after a week long  football practice in sun. His father had a similar condition. A diagnosis of Porphyria cutanea tarda was confirmed by finding elevated levels of porphyrins in his serum. His disease is due to a deficiency of which of the following enzymes?

a) ALA dehydratase

b) Ferrochelatase

c) PBG deaminase

d) Uroporphyrinogen decarboxylase.

Q.12- A 23 –year-old young woman, who recently began taking birth control pills, presents to emergency room with cramping abdominal pain, anxiety, hallucinations and paranoid behavior. A surgical evaluation, including Ultrasound and computed tomography (CT) scan have failed to demonstrate any abdominal process. Examination reveals vesicles and bullae on the skin of arms and face. Urine analysis reveals the presence of porphyrins (ALA, PBG, Uro and Coproporphyrins). What is the possible diagnosis for this patient?

a)  Variegate porphyria

b) Acute intermittent porphyria

c) Congenital Erythropoietic porphyria

d) Hereditary Coproporphyria

Q.13- A patient presents with dull right sided abdominal pain, fever from the 7 days, loss of appetite, pale stool and jaundice. Blood biochemistry reveals, mixed hyperbilirubinemia, high SGPT but near normal alkaline phosphatase levels. What is the cause of jaundice?

a) Viral hepatitis

b) Post hepatic jaundice

c) Hemolytic jaundice

d) None of the above

Q.14- Impaired Glucuronyl transferase activity is observed in all of the followings except-

a) Breast milk jaundice

b) Physiological jaundice of the new born

c) Crigler Najjar syndrome

d) Dubin Johnson syndrome

Q.15- Which out of the following conditions is not associated with excessive bilirubin formation from hemolysis –

a) Sickle cell anemia

b) Thalassemia

c) Malaria

d) Rotor syndrome

Q.16-What is expected out of Van den Bergh reaction in hepatic jaundice?

a) Direct positive

b) Indirect positive

c) Biphasic

d) None of the above.

Q.17- Which serum enzyme elevation is most diagnostic in obstructive jaundice?

a) ALT(Alanine amino transferase)

b) AST (Aspartate amino transferase)

c) LDH (Lactate dehydrogenase)

d) ALP (Alkaline phosphatase).

Q.18- Acholuric jaundice is –

a) Hemolytic jaundice

b) Hepatic jaundice

c) Post hepatic jaundice

d) None of the above

Q.19- Urine analysis of a patient reveals the presence of Bilirubin and urobilinogen, which serum enzyme is expected to be elevated  much higher than normal?

a) ALT (Alanine amino transferase)

b) AST (Aspartate amino transferase)

c) 5’ Nucleotidase

d) ALP (Alkaline phosphatase).

Q.20-A 65- year –old patient presents with weight loss, loss of appetite, dull dragging pain in the right hypochondrium and jaundice from the last 1 month.  Stool is reported to be clay colored from the same duration.   Blood biochemistry reveals Conjugated hyperbilirubinemia. Urine shows the presence of bilirubin. The patient has been diagnosed with carcinoma of head of the pancreas. Which serum enzyme is expected to be much higher than normal for this patient?

a) ALT (Alanine amino transferase)

b) AST (Aspartate amino transferase)

c) LDH (Lactate dehydrogenase)

d) ALP (Alkaline phosphatase).

Answers-

Q-1-b

Q.2- a

Q-3-b

Q-4-a

Q-5-d

Q-6-c

Q-7-c

Q-8-d

Q-9-c,

Q-10-c

Q11-d

Q12-a

Q-13-a

Q14-d

Q-15-d

Q.16-c

Q-17-d

Q-18-a

Q-19-a

Q-20-d


------------------------------------------ Best Wishes: Dr.Ehab Aboueladab, Tel:01007834123 Email:ehab10f@gmail.com,ehababoueladab@yahoo.com ------------------------------------------

Obesity

Case details The patient is obese and is also suffering from “Metabolic syndrome”. She has a history of obesity dating to early child hood and also has a  positive family history. Her symptoms are suggestive of metabolic syndrome, a common complication of Obesity. Some of her features can be discussed as -

1) Obesity -She has an apple (android) pattern of fat distribution. Her waist to hip ratio is 41/39=1.05. Apple shape is defined as a waist to hip ratio of more than 0.8 in women, and more than 1.0 in men. She has therefore apple pattern of fat distribution which is common in males. Compared with other women of same bodyweight who have gynoid fat pattern, the presence of increased visceral or intraabdominal adipose tissue places her at greater risk for diabetes, hypertension,dyslipidemia and coronary heart disease. (The gynoid, “pear- shaped” or lowerbody obesity is defined as a waist to hip ratio of less than 0.8 for women and less than 1.0 for men. The pear shape is relatively benign health wise and is commonly found in females).

2) BMI (Body Mass Index)

BMI=Weight (kg)/height (m2).

For this patient

188Pounds=85.5 kg (Approximately)

5 feet 1 inch height=1.55meters (154.94cm)

 =85.5/(1.55) 2

= 35.6 kg/ m2

World Health Organization (WHO) criteria based on BMI

Under this convention for adults,

Grade 1 overweight (commonly and simply called overweight) is a BMI of 25-29.9 kg/m2.

Grade 2 overweight (commonly called obesity) is a BMI of 30-39.9 kg/m2.

Grade 3 overweight (commonly called severe or morbid obesity) is a BMI greater than or equal to 40 kg/m2.

From the result calculated for the given patient, it is indicated that the patient is obese (Grade 2 overweight).

3)  Metabolic syndrome The patient is also suffering from ‘Insulin resistance syndrome’.She has hypertension, dyslipidemia, Hyperinsulinemia and impaired glucose tolerance.

Metabolic syndrome also referred to as Syndrome X or insulin resistance syndrome consists of a number of metabolic risk factors that increase the risk for atherosclerotic cardiovascular disease (CVD) and other cardiovascular complications such as cardiac arrhythmias, heart failure, and thrombotic events.

a) Criteria for diagnosis According to the National CholesterolEducation Program (NCEP) Adult Treatment Panel III (ATP III) report, there are six major components of metabolic syndrome relating to the development of CVD:

1) abdominal obesity

2) atherogenic dyslipidemia

3) elevated blood pressure

4) insulin resistance (with or without the presence of glucose intolerance)

5)proinflammatory state, and

6) prothrombotic state.

Note- A prothrombotic state is characterized by abnormalities,specifically elevations, in procoagulant factors, anti fibrinolytic factors,platelet alterations, and endothelial dysfunction. A proinflammatory state is characterized by elevations of circulating inflammatory molecules such as C-reactive protein (CRP), tumor necrosis factor-alpha, plasma resistin, interleukin (IL)-6, and IL-18. CRP is a general marker of inflammation that has been linked to CVD in patients with metabolic syndrome.

b)Associated diseases- Cardiovascular diseases and type 2 diabetes mellitus can be present in association with metabolic syndrome. The relative risk for new-onset CVD in patients with the metabolic syndrome, in the absence of diabetes, averages between 1.5- and threefold. Overall, the risk for type 2 diabetes in patients with the metabolic syndrome is increased three to fivefold. Patients with metabolic syndrome are also at increased risk for peripheral vascular disease. In addition to the features specifically associated with metabolic syndrome,insulin resistance is accompanied by other metabolic alterations. These include increases in uric acid,microalbuminuria, nonalcoholic fatty liver disease (NAFLD) and/or polycystic ovarian disease (PCOS), and obstructive sleep apnea (OSA).

A person suspected of having this syndrome should have a through history taken especially with regard to family history and presence of other cardiovascular risk factors.

c) An examination should include:-

 a) Recording the body weight

 b) Calculating the BMI

 c) Measurement of the waist circumference in inches

 d) Calculating the hip-waist ratio

 e) Measurement of the subcutaneous fat at 4 sites-biceps, triceps, sub scapular and supra-iliac

 f) Blood pressure measurement.

d) Laboratory Tests

1) Fasting lipids and glucose estimationsare needed to determine if the metabolic syndrome is present.

2) The measurement of additional biomarkers associated with insulin resistance must be individualized. Such tests might include apo B, high-sensitivity CRP,fibrinogen, uric acid, urinary microalbumin, and liver function tests.

3) A sleep study should be performed if symptoms of OSA are present.

4) If  PCOS is suspected based on clinical features and an ovulation, testosterone, luteinizing hormone, and follicle-stimulating hormone should be measured.

 e) Management of metabolic syndrome - is highly dependent on the control of all of the contributing factors. This includes both underlying risk factors as well as metabolic risk factors. Lifestyle modifications should be implemented immediately for all patients diagnosed with metabolic syndrome. Lifestyle modifications include weight reduction, increased physical activity and nutritional therapy. Additional risk assessments should be performed in patients to assure appropriate goals of therapy throughout the course of the syndrome.

The key to preventing metabolic syndrome, however,remains diet and exercise. Any person with a strong family history of metabolic syndrome or type 2 diabetes should be especially careful to maintain a healthy lifestyle.


------------------------------------------ Best Wishes: Dr.Ehab Aboueladab, Tel:01007834123 Email:ehab10f@gmail.com,ehababoueladab@yahoo.com ------------------------------------------

Starvation

A 35 –year-old woman became severely depressed after the sudden death of her husband. Two months later,she was brought to emergency room by her friend because of  extreme weakness and lethargy. She appeared thin and pale. Questioning revealed that she had not eaten for several weeks. 

Analysis of a plasma sample indicated elevated levels of Alanine, Acetoacetate, β hydroxy butyrate, and blood urea nitrogen(BUN). However her plasma glucose concentration was low (55mg/dL) . She was hospitalized, given intravenous feeding,  antidepressant medications and subsequently shifted to an 1800 Cal (7500kJ) diet. Her recovery was uneventful. 

How was the patient obtaining energy during the time when she was not eating?

How could patient  maintain her plasma glucose within normal limits even though she was not eating?

What  is the significance of  elevated  plasma Alanine level?

Why is BUN elevated?

What is indicated by the fact that the plasma Acetoacetate and β- hydroxy butyrate levels are elevated?

It is a case of starvation. The high blood Alanine level signifies the catabolic state. Alanine in excess is released during starvation from muscle to serve as a substrate for glucose production in liver. Acetoacetate and β hydroxy butyrate are ketone bodies which are used as alternative fuel during conditions of glucose deprivation . High BUN signifies protein degradation; the carbon skeletons of amino acids are utilized for glucose production while amino groups are converted to urea.

Starvation

Prolonged fasting may result from an inability to obtain food, from the desire to lose weight rapidly, or in clinical situations in which an individual can not eat because of trauma,surgery, neoplasms, burns etc or even in depression (As in the given case) . In the absence of food the plasma levels of glucose, amino acids and triacylglycerols fall, triggering a decline in insulin secretion and an increase in glucagon release. The decreased insulin to glucagon ratio, and the decreased availability of circulating substrates, make this period of nutritional deprivation a catabolic state, characterized by degradation of glycogen, triacylglycerol and protein. This sets in to motion an exchange of substrates between liver,adipose tissue, muscle and brain that is guided by two priorities (i) the need to maintain glucose level to sustain the energy metabolism of brain ,red blood cells and other glucose requiring cells and (ii) to supply energy to other tissues by mobilizing fatty acids from adipose tissues and converting them to ketone bodies to supply energy to other cells of the body.

Fuel Stores

A typical well-nourished 70-kg man has fuel reserves totaling about 161,000 kcal (670,000 kJ).  The energy need for a 24-hour period ranges from about 1600 kcal (6700 kJ) to 6000 kcal (25,000 kJ), depending on the extent of activity. Thus, stored fuels suffice to meet caloric needs in starvation for 1 to 3 months. However, the carbohydrate reserves are exhausted in only a day.

Energy supply during starvation

During starvation the energy needs are  fulfilled by three types of fuels, glucose, fatty acids and ketone bodies.

a) Glucose supply during starvation (Gluconeogenesis)

Energy needs of brain and RBCs

Even under  conditions of starvation, the blood-glucose level has be maintained above 2.2 mM (40 mg/dl). The first priority of metabolism in starvation is to provide sufficient glucose to the brain and other tissues(such as red blood cells) that are absolutely dependent on this fuel.However, precursors of glucose are not abundant. Most energy is stored in the fatty acyl moieties of triacylglycerols. Fatty acids cannot be converted into glucose, because acetyl CoA cannot be transformed into pyruvate. The glycerol moiety of triacylglycerol can be converted into glucose, but only a limited amount is available. The only other potential source of glucose is amino acids derived from the breakdown of proteins. However, proteins are not stored,and so any breakdown will necessitate a loss of function.

Thus, the second priority of metabolism in starvation is to preserve protein,which is accomplished by shifting the fuel being used from glucose to fatty acids and ketone bodies by cells  other than brain cells and the cells lacking mitochondria.

It is a biological compromise to provide glucose to these cells as a priority. During prolonged starvation , when the gluconeogenic precursors are not available, proteins are however  broken down to use carbon skeleton of  glucogenic amino acids for glucose production.

b) Fatty acid oxidation

Energy need of liver

The low blood-sugar level leads to decreased secretion of insulin and increased secretion of glucagon. Glucagon stimulates the mobilization of triacylglycerols in adipose tissue and gluconeogenesis in the liver. The liverobtains energy for its own needs by oxidizing fatty acids released from adipose tissue. The concentrations of acetyl CoA and citrate consequently increase, which switch off glycolysis. Thus glucose utilization is stopped in liver cells to preserve glucose for priority cells

Energy need of muscles

The uptake of glucose by muscle is markedly diminished because of the low insulin level, whereas fatty acids enter freely.Consequently, muscle shifts almost entirely from glucose to fatty acids for fuel.The beta-oxidation of fatty acids by muscle halts the conversion of pyruvate into acetyl CoA, because acetyl CoA stimulates the phosphorylation of the pyruvate dehydrogenase complex, which renders it inactive. Most of the pyruvate is transaminated to alanine, at the expense of amino acids arising from breakdown of “labile” protein reserves synthesized in the fedstate. The alanine, lactate and much of the keto-acids resulting from this transamination are exported from muscle, and taken up by the liver, where the alanine is transaminated to yield pyruvate. Pyruvate is a major substrate for gluconeogenesis in the liver.(Figure-1)

Figure- 1-showing Glucose Alanine and Cori’s cycle

In adipose tissue the decrease in insulin and increase in glucagon results in activation of intracellular hormone-sensitive lipase.This leads to release from adipose tissue of increased amounts of glycerol(which is a substrate for gluconeogenesis in the liver) and free fatty acids,which are used by liver, heart, and skeletal muscle as their preferred metabolic fuel, therefore sparing glucose.

Loss of muscle mass

During starvation, degraded proteins are not replenished and serve as carbon sources for glucose synthesis. Initial sources of protein are those that turn over rapidly, such as proteins of the intestinal epithelium and the secretions of the pancreas. Proteolysis of muscle protein provides some of three-carbon precursors of glucose. The nitrogen partof the amino acids is converted to urea (BUN)

c) Ketosis

Energy need of peripheral tissues

After about 3 days of starvation, the liver forms large amounts of acetoacetate and beta- hydroxybutyrate. Their synthesis from acetyl CoA increases markedly because the citric acid cycle is unable to oxidize all the acetyl units generated by the degradation of fatty acids. Gluconeogenesis depletes the supply of oxaloacetate, which is essential for the entry of acetyl CoA into the citric acid cycle. (Figure-2) Consequently, the liver produces large quantities of ketone bodies, which are released into the blood. At this time, the brain begins to consume appreciable amounts of acetoacetate in place of glucose. After 3 days of starvation, about a third of the energy needs of the brain are met by ketone bodies. The heart also uses ketone bodies as fuel. After several weeks of starvation, ketone bodies become the major fuel of the brain.

Figure- 2-fatty acid oxidation and ketosis during starvation.

In essence, ketone bodies are equivalents of fatty acids that can pass through the blood-brain barrier. Only 40 g of glucose is then needed per day for the brain, compared with about 120 g in the first day of starvation. The effective conversion of fatty acids into ketone bodies by the liver and their use by the brain markedly diminishes the need for glucose. Hence, less muscle is degraded than in the first days of starvation. The breakdown of 20 g of muscle daily compared with 75 g early in starvation is most important for survival.

A person’s survival time is mainly determined by the size of the triacylglycerol depot.

What happens after depletion of the triacylglycerol stores? The only source of fuel that remains is proteins. Protein degradation accelerates, and death inevitably results from a loss of heart, liver, or kidney function.

------------------------------------------ Best Wishes: Dr.Ehab Aboueladab, Tel:01007834123 Email:ehab10f@gmail.com,ehababoueladab@yahoo.com ------------------------------------------