Liver Function Tests

The liver function test panel combines markers of hepatocellular injury (alanine aminotransferase, ALT, and aspartate aminotransferase, AST), cholestasis (alkaline phosphatase, ALP, and gamma glutamyl transferase, GGT), bilirubin handling (total and direct bilirubin) and synthetic function (albumin, with total protein and globulin).

Enzymes are measured by kinetic spectrophotometry, bilirubin by diazo methods, albumin by dye binding and total protein by the biuret method. The panel detects liver disease, defines its pattern, and monitors drug toxicity and chronic disease. It measures injury and excretory function. It is not a measure of true liver function, and normal values do not exclude significant disease, including cirrhosis.

Abnormal liver enzymes are found in up to about 10% of the general population, most often from fatty liver disease, alcohol and drugs. Metabolic dysfunction associated steatotic liver disease (previously non-alcoholic fatty liver disease) is now the commonest cause worldwide, and viral hepatitis remains a leading cause in much of Africa and Asia.

Indications. Jaundice, abdominal pain, fatigue, pruritus, dark urine, pale stools, ascites, encephalopathy, suspected viral hepatitis, alcohol use, obesity and diabetes, monitoring of hepatotoxic drugs (isoniazid, rifampicin, pyrazinamide, methotrexate, statins, azoles, antiretrovirals, valproate, amiodarone), paracetamol overdose, abnormal imaging, pregnancy complications, and routine pre-treatment assessment.

Specimen. Serum or lithium heparin plasma. Fasting is preferred, since a fatty meal can raise ALP. Protect the sample from light for bilirubin, and avoid haemolysis.

Adult reference ranges are given. They are method specific, and local ranges take precedence.

Alanine aminotransferase (ALT). Cytosolic enzyme found mostly in hepatocytes, released when cells are injured. Range: men up to about 40 to 50 U/L, women up to about 30 to 35 U/L.

Raised: viral hepatitis (A, B, C, E, and herpes viruses, EBV, CMV), alcohol, fatty liver disease, drug and toxin injury (paracetamol, antituberculous drugs, statins, amoxicillin-clavulanate, methotrexate, valproate, antiretroviral drugs, herbal products), autoimmune hepatitis, ischaemic hepatitis. Other raised causes: haemochromatosis, Wilson disease, alpha-1 antitrypsin deficiency, coeliac disease, thyroid disease, muscle injury.

Lowered: pyridoxine deficiency and advanced kidney disease, without clinical importance.

It is more liver specific than AST. A value over 1000 U/L suggests ischaemic hepatitis, paracetamol or other toxin injury, acute viral hepatitis or autoimmune hepatitis. Values under 5 times the upper limit are mild and are usually fatty liver, chronic viral hepatitis, alcohol or drugs. ALT can be normal in chronic hepatitis B and C and in cirrhosis.

Aspartate aminotransferase (AST). Enzyme found in liver, heart, skeletal muscle, kidney and red cells, with mitochondrial and cytosolic forms. Range: up to about 35 to 40 U/L. Raised: all causes of raised ALT, plus muscle injury (rhabdomyolysis, myositis, exertion, intramuscular injection), myocardial infarction, haemolysis, macro-AST, hypothyroidism. Lowered: no clinical significance.

An AST to ALT ratio over 2 suggests alcoholic hepatitis, usually with AST under 300 U/L and a raised GGT. A ratio over 1 also occurs in cirrhosis. A ratio under 1 is typical of viral hepatitis and fatty liver disease before cirrhosis develops. An AST higher than ALT with a normal GGT and a raised creatine kinase points to a muscle source.

Alkaline phosphatase (ALP). Enzyme from the biliary canalicular membrane, bone, placenta and intestine. Range: about 30 to 130 U/L in adults.

Raised: biliary obstruction (gallstones, pancreatic or biliary cancer, strictures, parasites such as Ascaris and liver flukes), primary biliary cholangitis, primary sclerosing cholangitis, drug induced cholestasis, infiltrative disease (granulomas, metastases, lymphoma, amyloid, sarcoidosis), cholangitis, sepsis, congestive hepatopathy.

Raised non-hepatic causes: bone disease (Paget disease, osteomalacia, rickets, fractures, bone metastases, hyperparathyroidism), pregnancy, growth in children and adolescents, and a high fat meal in some blood groups.

Lowered: hypophosphatasia, zinc or magnesium deficiency, hypothyroidism, pernicious anaemia, coeliac disease, Wilson disease with acute liver failure, EDTA contamination.

A raised ALP with a raised GGT indicates a hepatobiliary source. A raised ALP with a normal GGT suggests bone or another non-hepatic source, and ALP isoenzymes or 5'-nucleotidase confirm it. In Wilson disease with acute liver failure, an ALP to bilirubin ratio under 4 with an AST to ALT ratio over 2.2 is characteristic.

Gamma glutamyl transferase (GGT). Membrane enzyme of bile duct epithelium and hepatocytes, induced by alcohol and drugs. Range: men about 10 to 60 U/L, women about 7 to 40 U/L. Raised: cholestasis, alcohol, enzyme inducing drugs (phenytoin, carbamazepine, barbiturates, rifampicin), fatty liver disease, obesity, diabetes, pancreatitis, heart failure, hyperthyroidism, smoking. Lowered: no clinical significance.

An isolated raised GGT is non-specific and does not on its own indicate liver disease. It is sensitive for alcohol use but not specific, and it falls over several weeks of abstinence.

Total and direct (conjugated) bilirubin. Bilirubin is the breakdown product of haem, conjugated in the liver and excreted in bile. Range: total 3 to 17 micromol/L (0.2 to 1.0 mg/dL), direct under about 5 micromol/L (0.3 mg/dL).

• Raised unconjugated: haemolysis, Gilbert syndrome, Crigler-Najjar syndrome, ineffective erythropoiesis, neonatal jaundice, drugs (rifampicin, atazanavir, indinavir), large haematomas.
• Raised conjugated: hepatocellular disease, cholestasis, sepsis, parenteral nutrition, Dubin-Johnson syndrome, Rotor syndrome.
• Lowered: no clinical significance.

Jaundice becomes visible at a total bilirubin over about 40 micromol/L (2.5 mg/dL). Conjugated bilirubin is water soluble and appears in the urine (dark urine, positive dipstick), while unconjugated bilirubin does not. Gilbert syndrome affects 3 to 7% of people, with total bilirubin usually under 85 micromol/L (5 mg/dL), a conjugated fraction under 20 to 30%, normal enzymes and no haemolysis, rising with fasting or illness.

ALT over 3 times the upper limit with bilirubin over 2 times the upper limit, without a raised ALP, in a patient on a new drug indicates serious drug induced liver injury with a mortality of about 10%.

Albumin. Main plasma protein, made only in the liver, with a half life of about 20 days. Range: 35 to 50 g/L. Raised: dehydration only. Lowered: chronic liver disease, nephrotic syndrome, protein losing enteropathy, malnutrition, inflammation (it is a negative acute phase protein), sepsis, burns, haemodilution, heart failure, pregnancy.

Because of its long half life, albumin does not fall early in acute liver failure, and the INR and factor V are better measures of acute synthetic function. A low albumin in liver disease signals chronic disease, but it is not specific, since inflammation and nutrition lower it.

Total protein and globulin. Total protein is albumin plus globulins. Globulin is calculated as total protein minus albumin. Range: total protein 60 to 80 g/L, globulin 20 to 35 g/L. Raised globulin: polyclonal hypergammaglobulinaemia (autoimmune hepatitis, cirrhosis, chronic infection, HIV, autoimmune disease), monoclonal protein (myeloma), dehydration. Lowered total protein: the causes of low albumin, and immune deficiency.

A raised globulin with a disproportionate rise in IgG supports autoimmune hepatitis. A globulin fraction over about 35 to 40 g/L prompts serum protein electrophoresis.

Pattern recognition by the R ratio. The R ratio is ALT divided by its upper limit of normal, divided by ALP divided by its upper limit of normal.

• Over 5: hepatocellular pattern.
• 2 to 5: mixed pattern.
• Under 2: cholestatic pattern.

Hepatocellular pattern (ALT and AST raised out of proportion to ALP). Viral hepatitis, alcohol, fatty liver disease, drug and toxin injury, autoimmune hepatitis, ischaemic hepatitis, haemochromatosis, Wilson disease, alpha-1 antitrypsin deficiency, acute Budd-Chiari syndrome, herpes hepatitis, acute fatty liver of pregnancy and HELLP syndrome.

Cholestatic pattern (ALP and GGT raised, bilirubin raised, transaminases modestly raised).

• Extrahepatic: gallstones, pancreatic head and biliary cancer, strictures, pancreatitis, parasitic obstruction.
• Intrahepatic: primary biliary cholangitis, primary sclerosing cholangitis, drug induced cholestasis (amoxicillin-clavulanate, flucloxacillin, chlorpromazine, anabolic steroids), infiltration, sepsis, cholestasis of pregnancy, congestive hepatopathy.

Mixed pattern. Drug injury, viral hepatitis in its cholestatic phase, alcohol, infiltrative disease and early biliary obstruction.

Isolated abnormalities.

• Isolated raised bilirubin with normal enzymes: Gilbert syndrome or haemolysis, separated by the conjugated fraction and a haemolysis screen.
• Isolated raised ALP: bone disease, pregnancy, growth, or early hepatic infiltration.
• Isolated raised GGT: alcohol, drugs or fatty liver.
• Low albumin with a prolonged INR that does not correct with vitamin K: synthetic failure.
• Prolonged INR that corrects with vitamin K: deficiency from cholestasis or malabsorption.

Alcohol related pattern. AST to ALT ratio over 2, GGT raised, raised MCV, and a low platelet count.

Congestive and ischaemic patterns. Right heart failure raises bilirubin, ALP and GGT. Ischaemic hepatitis gives a rapid, high rise in ALT and AST (often over 1000 U/L) with a prolonged INR and a rapid fall over days.

Critical values and findings needing immediate action. Triggers vary by laboratory.

• ALT over 1000 U/L, particularly with jaundice, encephalopathy or a prolonged INR.
• Acute liver failure: INR of 1.5 or over with any encephalopathy in a patient without known chronic liver disease, within 26 weeks of illness onset.
• Raised ALT or AST and bilirubin with paracetamol exposure, which needs a paracetamol level and urgent management.
• Jaundice with fever, rigors and right upper quadrant pain (cholangitis).
• ALT over 3 times the upper limit with bilirubin over 2 times the upper limit on a new drug.
• Pregnancy: raised enzymes with hypertension, low platelets, hypoglycaemia or encephalopathy.

Interferences and pitfalls.

• Haemolysis raises AST and, less so, ALT. Strenuous exercise, intramuscular injection and muscle disease raise AST and creatine kinase.
• Bilirubin degrades in light, and delay falsely lowers the result.
• EDTA contamination lowers ALP.
• Macro-AST and macro-ALP are complexes that cause persistent isolated elevation without disease.
• Biotin supplements interfere with some immunoassay based tests, though most enzyme assays are unaffected.
• A fatty meal raises ALP in some people of blood group O and B.
• Normal liver tests do not exclude cirrhosis, chronic hepatitis B or C, haemochromatosis, or early fatty liver disease.
• The upper limit is set at the 95th percentile, so about 5% of healthy people have a mildly abnormal value, and mild elevations are often repeated before workup.
• Herbal and traditional products and supplements are common unrecognised causes of liver injury and need a direct history.

Special groups.

• Neonates: bilirubin is interpreted against age in hours using treatment thresholds, and conjugated bilirubin over 17 micromol/L (1 mg/dL), or over 20% of total, needs urgent investigation for biliary atresia and other cholestatic disease.
• Children: ALP is high during growth (up to several times the adult upper limit) and in adolescence.
• Pregnancy: ALP rises 2 to 4 times in the third trimester from placental production, and albumin falls to about 28 to 37 g/L. ALT, AST, GGT and bilirubin should remain normal. Raised values suggest intrahepatic cholestasis of pregnancy (with serum bile acids over 10 micromol/L), HELLP syndrome or acute fatty liver of pregnancy.
• Older adults: ALP rises and albumin falls with age, and drug induced liver injury is more frequent.
• Kidney disease: ALT and AST are lower in dialysis patients, so mild elevation can be significant.
• Regions: hepatitis B and E, schistosomiasis, liver flukes, aflatoxin exposure and antituberculous drug injury are important regional causes.

Follow up tests. INR, full blood count, hepatitis A, B, C and E serology, HIV testing. Autoimmune markers (ANA, anti-smooth muscle antibody, anti-mitochondrial antibody, immunoglobulins), iron studies and ferritin, caeruloplasmin, alpha-1 antitrypsin, paracetamol level, creatine kinase, serum bile acids, lipid profile and HbA1c. Imaging and tissue: abdominal ultrasound, transient elastography, MR cholangiopancreatography, liver biopsy.