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Clinical enzymology

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ALANINE AMINO TRANSFERASE (ALT)

  • i. In old literature, it was called as serum glutamate pyruvate transaminase (SGPT). The enzyme needs pyridoxal phosphate as co-enzyme.
  • ii. Normal serum level of ALT for male is 13–35 U/L and for female is 10–30 U/L. Very high values (300 to 1000 U/L) are seen in acute hepatitis, either toxic or viral in origin (infective hepatitis).
  • iii. Both ALT and AST levels are increased in liver disease, but ALT > AST. Rise in ALT levels may be noticed several days before clinical signs such as jaundice are manifested.
  • iv. Moderate elevation of amino transferases often between 100–300 U/L is seen in alcoholic hepatitis, autoimmune hepatitis, Wilson’s disease and non- alcoholic chronic hepatitis .
  • v. Minor elevation less than 100U/L is seen in chronic viral hepatitis (hepatitis C), fatty liver and in non- alcoholic steatohepatitis (NASH). In chronic hepatitis and cirrhosis of liver, serum ALT poorly correlates with the degree of liver cell damage.
  • vii. A normal value need not rule out minor liver diseases. On the other hand, normal persons may have elevated ALT levels. This is seen especially in obese persons. 1% loss of weight will reduce ALT values by 8%.
  • Vii The degree of elevation may reflect the extent of hepatocellular necrosis. In most cases the lowering of the level of transaminases indicates recovery, but a sudden fall from a very high level may indicate poor prognosis.
  • viii. Elevation of ALT is more in cases of hepatic disease compared to AST. But AST may be more than ALT in alcoholic liver disease. In alcoholic liver disease, the actual values show only mild elevation; but a ratio of AST/ALT more than two is quite suggestive.

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ASPARTATE AMINO TRANSFERASE (AST)

  • i. In old literature, it was called as serum glutamate oxaloacetate transaminase (SGOT). AST needs pyridoxal phosphate (vitamin B6 ) as co-enzyme.
  • ii. Normal serum level of AST is 8–20 U/L. The level is elevated in myocardial infarction.
  • iii. It is signifcantly elevated in liver diseases. A marked increase in AST may be seen in primary hepatomas.
  • iv. In alcoholic hepatitis AST may be higher than ALT since mitochondrial enzyme is also liberated.

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ALKALINE PHOSPHATASE (ALP)

  • i. ALP is a nonspecific enzyme which hydrolyzes aliphatic, aromatic or heterocyclic compounds. The pH optimum for the enzyme reaction is between 9 and 10. It is activated by magnesium and manganese. Zinc is a constituent ion of ALP.
  • Ii. It is produced by osteoblasts of bone, and is associated with the calcification process (see Chapter 35). It is localized in cell membranes (ecto-enzyme), and is associated with transport mecha nisms in liver, kidney and intestinal mucosa.
  • iii. Normal serum value of ALP is 40–125 U/L. In children, the upper level of normal value may be more, because of the increased osteoblastic activity in children.
  • iv. Moderate (2–3 times) increase in ALP level is seen in hepatic diseases such as infective hepatitis, alcoholic hepatitis or hepatocellular carcinoma (see Chapter 26).
  • v. Very high levels of ALP (10–12 times of upper limit) may be noticed in extrahepatic obstruction (obstructive jaundice) caused by gallstones or by pressure on bile duct by carcinoma of head of pancreas. Intrahepatic cholestasis may be caused by viral hepatitis or by drugs (chlorpromazine). ALP is produced by epithelial cells of biliary canaliculi and obstruction of biliary passage with consequent irritation of epithelial cells leads to secretion of ALP into serum.
  • vi. Drastically high levels of ALP (10–25 times of upper limit) are also seen in bone diseases where osteoblastic activity is enhanced such as Paget's disease (osteitis deformans), rickets, osteomalacia, osteoblastoma, metastatic carcinoma of bone and hyperparathyroidism
  • .vii. There are 6 iso-enzymes for ALP. The one, which is inhibited by phenylalanine is of placental origin. It is found in blood in normal pregnancy. An iso enzyme closely resembling the placental form is characteristically seen in circulation in about 15% cases of carcinoma of lung, liver and gut and named as Regan iso-enzyme or carcinoplacental iso-enzyme

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Gamma Glutamyl Transferase (GGT)

  • 1. The old name was gamma glutamyl transpeptidase. It can transfer gamma glutamyl residues to substrate. In the body it is used in the synthesis of glutathione. GGT has 11 iso-enzymes. It is seen in liver, kidney, pancreas, intestinal cells and prostate gland.
  • 2. Normal serum value of GGT is 10 –30 U/L. It is moderately increased in infective hepatitis and prostate cancers.
  • 3. GGT is clinically important because of its sensitivity to detect alcohol abuse. GGT is increased in alcoholics even when other liver function tests are within normal limits. GGT level is rapidly decreased within a few days when the person stops to take alcohol. Increase in GGT level is generally proportional to the amount of alcohol intake.

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Sorbitol dehydrogenase (SDH)

  • Sorbitol dehydrogenase (SDH) is a liver-specific enzyme in animals, particularly used in veterinary LFTs (especially in horses and cattle) to detect acute liver damage.
  • ↑ SDH = Acute hepatocellular injury (e.g., due to toxins, infection, hypoxia)
  • Normal SDH = No significant hepatocyte damage or recovery phase Increased SDH activity in serum indicates:
  • Acute hepatocellular (liver cell) damage
  • Toxic liver injury
  • Hypoxic liver damage
  • Infectious hepatitis
  • Normal levei 0-8 U/L

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1. CK (CREATINE KINASE)

Principle:

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Method : Increasing Kinetic

Biological reference range :- 25-120 U/L

Clinical Significance:

  • CPK ( Creatine phosphokinase ) activity is increase in
    • Brain (CK-BB = 1%)
      • Cerebro-vascular stroke
    • heart muscle (CK-MB = 5-10%)
      • Myocardial infarction
      • Acute coroary syndrome
    • skeletal muscle. (CK-MM= 85%)
      • Crush injury
      • Myopathy
      • Polymyositis

​

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2. CK-MB

Principle:

  • measurement of CK activity in the presence of an antibody to CK-M monomer.
  • This antibody completely inhibits the activity of CK-MM & half of the activity of CK-MB while not affecting the B subunit activity of CK-MB &CK-BB.

Method :

  • Increasing Kinetic UV method

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Biological reference range :-0 - 25 IU/L

Significance:

  • CK-MB present only in cardiac tissue
  • So…Specific for diagnosis of cardiac disease

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CK-2 & CK-3 in normal subject &

After 24 hours of Myocardial Infarction

Creatine Kinase isoenzymes in blood

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3. LDH (lactate dehydrogenase)

Principle:

  • Lactate dehydrogenase catalyses the conversion of pyruvate to lactate.

​

Lactate dehydrogenase

Lactate + NAD+ ----------------🡪 Pyruvate + NADH + H+

​

​

The rate of NADH+ formation is measured by 340nm filter.

It is direactly propotional to serum LDH activity.

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Method :

  • IFCC method (Lactate to Pyruvate method )
  • Increasing kinetic

Biological reference range :- 70-240 IU/L

Clinical significance :

  • RBC
    • Malaria
    • Sickle cell anemia
    • hemolytic disorder
  • Liver
    • Viral Hepatitis
    • Liver malgnancy
    • Alcoholic liver disease
  • Cardiac tissue
    • Myocardial infarction
  • Skeletal muscle
    • Muscular distrophy
    • Crush injury

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LDH Iso-enzyme

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Species

Normal Serum SDH Activity

Unit

Notes

Human

Very low or undetectable (< 2 U/L)

U/L

SDH is not routinely used in human LFTs — ALT, AST, and ALP are preferred. Detectable levels may indicate acute hepatic necrosis.

Horse

0 – 8 U/L (some labs: up to 12 U/L)

U/L

Highly liver-specific; rises sharply within hours after hepatocellular injury.

Cattle

0 – 6 U/L

U/L

Sensitive indicator of acute liver injury (e.g., toxic or hypoxic damage).

Sheep / Goat

0 – 5 U/L

U/L

Similar diagnostic value to that in cattle.

Dog

0 – 3 U/L

U/L

Usually very low; ALT is preferred in dogs.

Cat

0 – 3 U/L

U/L

Same as dog; ALT and AST are more reliable.

Pig

0 – 7 U/L

U/L

Occasionally used for detecting liver injury due to toxins.

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  • Measurement of pancreatic enzymes: Amylase or alpha-1,4-glucosidase is the major enzyme which digests starch.
  • The serum amylase contains the P (pancreatic) and S (salivary) iso-enzymes. These two can be distinguished by the inhibition test.
  • A protein inhibitor, present in alcoholic extracts of wheat will selectively inhibit the S isoenzyme.
  • Normal amylase level in serum is 50–120 units.
  • The level rises within 5 hours of the onset of acute pancreatitis and the level reaches a peak within 12 hours. But the level need not parallel the severity of the disease. Within 2–4 days of the attack, the level returns to normal.
  • As the serum amylase level starts falling, urinary amylase level rises. If the sample is collected too early, the serum amylase levels may not show the expected rise. If the sample is collected too late, again serum amylase may be low due to necrosis of the pancreatic tissue.

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  • Amylase level in blood is mildly increased in cases of cholecystitis, peptic ulcer, diseases of mesentery and obstruction of intestine.
  • A small percentage of patients with acute pancreatitis fails to show any rise.
  • No significant change or only mild elevation is noticed in chronic pancreatitis.

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Serum lipase

  • is the major lipolytic enzyme which hydrolyzes glycerol esters of long chain fatty acids.
  • The level in blood is highly elevated in acute pancreatitis and this persists for 7 – 14 days. Thus lipase remains elevated longer than amylase. Moreover, lipase is not increased in salivary diseases. Therefore, lipase estimation has advantage over amylase.