1. Part 1 Liver enzyme(ALT-AST-ALP-GGT)
Liver function test
Part 1
Liver enzyme(ALT-AST-ALP-GGT)

3. Liver is the largest Organ of the body weighing about 1.5 Kg.
Liver is called kitchen of our body.

4. Major Metabolic Functions of the Liver
Synthetic Function
Plasma proteins (albumin, globulins), cholesterol, triglycerides and lipoproteins
Detoxification and excretion
Ammonia to urea (urea cycle), bilirubin, cholesterol, drug metabolites
Storage Function
Vitamins A, D, E, K and B12
Production of bile salts
Helps in digestion

5. Liver Function Tests (LFTs)
Noninvasive methods for screening of liver dysfunction
Help in identifying general types of disorder
Assess severity and allow prediction of outcome
Disease and treatment follow up

6. Liver Function Tests (LFTs)
Broadly classified as:
Tests to detect hepatic injury:
Mild or severe; acute or chronic
Nature of liver injury (hepatocellular or cholestasis)
Tests to assess hepatic function

7. Classification of LFTs
Group I: Markers of liver dysfunction
Serum bilirubin: total and conjugated
Urine: bile salts and urobilinogen
Total protein, serum albumin and albumin/globulin ratio
Prothrombin Time

8. Classification of LFTs
Group II: Markers of hepatocellular injury
Alanine aminotransferase (ALT)
Aspartate aminotransferase (AST)

9. Classification of LFTs
Group III: Markers of cholestasis
Alkaline phosphatase (ALP)
g-glutamyltransferase (GGT)

10. Liver Enzymes

11. Aminotranferases
Aminotransferases or transaminases are a group of enzymes that catalyze the interconversion of amino acids and ketoacids (oxoacids) by transfer of amino group
The two aminotransferases of greatest clinical significance are:
Aspartate aminotransferase (AST), formerly termed glutamate oxaloacetate transaminase (GOT).
Alanine aminotransferase (ALT), formerly termed glutamate pyruvate transaminase (GPT).
Pyridoxal phosphate (P-5-P) is coenzyme

12. Aspartate aminotransferase (AST)
AST involved in the transfer of an amino group between aspartate and -ketoacids.
Mohammed Laqqan

13. Clinical Significance
Aspartate aminotransferase (AST) is an enzyme found primarily in the heart, liver, and muscle.
Additional AST is released into the circulation after injury or death of cells.
This test is one of several that are performed when there has been damage to the heart muscle, as in myocardial infarction, and in assessing liver damage.
Infants levels approximately twice the adult level, these decline to adult levels by approximately 6 months of age.
Mohammed Laqqan

14. Specimen Collection Specimen: Serum, heparin plasma or EDTA plasma
Hemolysis should be avoided because it can dramatically increase serum AST concentrations
(RBCs contain 15 X the AST activity in serum)
Mohammed Laqqan

15. Assay for Enzyme activity
Measurement by Karmen method
A coupled reaction involving:
pyridoxal-5-phosphate (P-5-P)
and malate dehydrogenase (MDH)
at 37oC:
Decrease in absorbance at 340 nm is determined by continuous monitoring.
AST
Aspartate + -Ketoglutarate Oxaloacetate + Glutamate
Oxaloacetate + NADH + H Malate + NAD MD
Mohammed Laqqan

16. Normal range: 8 – 20 U/L
Post AMI
Rises 6 – 8 hours
Peaks at 24 hours
Returns to normal by day 5
AST levels are highest in acute hepatocellular disorders "viral hepatitis, cirrhosis.

17. Alanine Aminotransferase (ALT)
A transferase with enzymatic activity similar to AST
Converts alanine + α-ketoglutarate to pyruvate and glutamate
Mohammed Laqqan

18. Clinical Significance
It is found in the kidneys, heart, and skeletal muscle tissue but primarily in liver tissue.
The test is used mainly in the diagnosis of liver disease and to monitor the effects of hepatotoxic drugs.
Often higher than AST with liver damage and tend to remain elevated longer(More liver-specific than AST)
Remains normal in AMI
Mohammed Laqqan

19. Specimen Collection Specimen: Serum, heparin plasma or EDTA plasma
Hemolysis should be avoided because it can increase serum ALT concentrations
(ALT in RBCs is roughly 5 X that of serum)
Mohammed Laqqan

20. Assay for enzyme activity
The most common method in use today for measurement of ALT activity utilizes a coupled enzymatic procedure for monitoring disappearance of NADH.
In this approach lactate dehydrogenase (LDH) and its required cofactors are added and catalyze the conversion of pyruvate to lactate
This causes simultaneous oxidation of reduced nicotinamide adenine dinucleotide (NADH).
The disappearance of NADH is followed spectrophotometrically (at 340 nm).
Alanine + -Ketoglutarate Pyruvate + Glutamate
Pyruvate + NADH + H Lactate + NAD LD
ALT

21. Male: 13-35 Female: 10-30 Normal range (U/L):
High serum levels in acute hepatitis ( U/L)
Moderate elevation in alcoholic hepatitis ( U/L)
Minor elevation in cirrhosis, hepatitis C and non-alcoholic steatohepatitis (NASH) (50-100U/L)

22. Levels of AST & ALT
AST is assessed along ALT in monitoring liver damage.
These two values normally exist in an approximately 1:1 ratio.
As a rough guide:
AST>ALT in:
alcoholic hepatitis and cirrhosis,
metastatic cancer of the liver
non-biliary cirrhosis,
while ALT>AST in:
viral and drug hepatitis,
chronic hepatitis C
and hepatic obstruction.
Mohammed Laqqan

23. Alkaline Phosphatase (ALP)
Phosphatases transfer a phosphate moiety from one group to a second, forming an alcohol and a second phosphate compound.
The optimal reaction pH for ALP is between 9 and 10 and varies with the buffer and substrate.
ALP requires Mg2+ as an activator
Mohammed Laqqan

24. Isoenzymes Liver isoenzyme (fastest) Bone isoenzyme
ALP exists as a number of isoenzymes
Major are those found in Liver, bone, placenta, and then intestinal fraction
Electrophoresis for isoenzyme analysis
Liver isoenzyme (fastest)
Bone isoenzyme
Placental isoenzyme
Intestinal isoenzyme (slowest)
Immunochemical methods now available
Mohammed Laqqan

25. Clinical Significance
Alkaline phosphatase (ALP) is an enzyme found in the liver, bone, placenta, intestine, and kidneys
Primarily in the cells lining the biliary tract and in the osteoblasts involved in the formation of new bone.
ALP is normally excreted from the liver in the bile.
Increased ALP levels are found most commonly during:
periods of bone growth (as in children),
in various types of liver disease,
and in biliary obstruction.
Serum ALP activity primarily reflects changes in bone and liver function, even though higher ALP activities can be found in other organs.
Mohammed Laqqan

26. Specimen Collection
Blood should be drawn after a fast of at least 8 hours.
Serum or heparinized plasma.
Slight hemolysis is tolerable, but gross hemolysis should be avoided.
These increases may be caused by:
the release of ALP from complexes with lipoproteins,
It is best to analyze ALP specimens the same day they are drawn.
ALP is inhibited by metal-complexing anticoagulants; EDTA, oxalate, and citrate inhibit the enzyme by complexing Mg2+ and should not be used.
Mohammed Laqqan

27. Assay for Enzyme activity
almost all assays for ALP employ p-nitrophenyl phosphate as the substrate.
Bowers and Combs method based on absorption of p-nitrophenol at 405 nm
At an alkaline pH,
p-nitrophenyl phosphate is colorless;
the product p-nitrophenol is intensely yellow

28. Normal range:
40 – 125 U/L

29. γ-Glutamyltransferase (GGT)
Transfers γ-glutamyl residue from γ-glutamyl peptides (usually glutathione) to amino acids, H2O and other small peptides
glutathione serves as the γ-glutamyl donor Glutathione + Amino Acid
glutamyl-peptide + L-cysteinylglycine
High concentrations in liver tissue
Also in pancreas and kidney
GGT

30. Diagnostic Significance
Increased plasma GGT is associated with
Hepatobiliary disease
Highest seen in biliary obstruction
Alcoholic cirrhosis
Used with ALP to differentiate between liver and bone diseases

31. Assay for enzyme activity
γ- glutamyl residue of γ glutamyl-p-nitroanilide is transferred to glycylglycine, releasing p-nitroaniline
L-γ-glutamyl-p-nitroanilide + glycylglycine
p-nitroaniline + y-glutamyl glycylglycine
The rate of liberation of p-nitroaniline is directly related to the GGT activity in the sample and is quantitated by measuring the increase in absorbance at 405 nm.
GGT
Reference Range: male, 6-45 U/L (37°C); female, 5-30 U/L (37°C)