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  1. This test is done in the serum of the patient.
  2. The random sample can be taken and no need for a fasting sample.
  3. From infants, the blood may be collected from the heel.
  4. The sample is stable at 4 °C for 3 days and protects it from the light.


  1. Avoid hemolysis.
  2. Avoid shaking of the tube that may lead to inaccurate results.
  3. Do not expose the tube to light. Exposure to light like the sun or even artificial light may decrease the value.
  4. If there is a delay in the test then keep the sample away from the light and refrigerate it.

Purpose of the test (Indications)

  1. For the diagnosis of jaundice.
  2. To differentiate different types of jaundice.
  3. For the follow-up of a patient with treatment.
  4. To assess the progress of the disease.
  5. This test is done to evaluate the liver functions.
  6. This is done in a patient with hemolytic anemia in adults.
  7. It is also done to evaluate the hemolytic anemia in the newborn.


  1. Raised Bilirubin is the indicator of liver dysfunction.
  2. Bilirubin is the basic end product of Hemoglobin metabolism.
    1. Total daily production of the bilirubin is 250 to 300 mg/day. 
    2. Hemoglobin from RBC is released and form Heme and Globin.
    3. Heme is metabolized to Biliverdin and this is transformed into bilirubin.

    1. The increased amount of bilirubin causes the yellow color of the skin and conjunctiva (Jaundice). 
      1. Because bilirubin is an orange-yellow pigment derived from the metabolism of heme.
    2. Further metabolism of bilirubin takes place in the intestine.
    3. Most of it is metabolized in the intestine and excreted in feces.
      1. Bilirubin not absorbed in the intestine, hydrolyzed by the β- glucuronidase enzyme which comes from:
        1. Liver.
        2. Intestinal epithelial cells.
        3. Bacteria.
      2. This unconjugated bilirubin is reduced by the intestinal microbial flora to urobilinogen (tetrapyrrole compound which is colorless).
      3. Analysis of the bilirubin from the sera of various patients of liver diseases show the following pattern:
        1. Unconjugated bilirubin = 27%
        2. Monoconjugated bilirubin = 24%.
        3. Diconjugated bilirubin = 13%.
        4. Protein-bound bilirubin = 37%.
      4. The bilirubin which is not conjugated, attach to albumin (carrying protein )is called Indirect bilirubin.
      5. Heme from the RBC gives rise to protoporphyrins.
        1. Now protoporphyrins by microsomal heme oxygenase form Biliverdin IXα.
        2. Biliverdin reductase enzyme transforms it into bilirubin IXα.


  1. Total Bilirubin = Direct bilirubin + Indirect bilirubin.
    1. Indirect bilirubin  is calculated as follows: 
      1. Indirect bilirubin = Total bilirubin – direct bilirubin.
  2. Urobilinogen differs from one another in the degree of hydrogenation of the vinyl side chain, as well as in the two end pyrrole rings.
    1. Urobilinogen containing 6, 8, and 12 more hydrogen ions atoms respectively are called as:
      1. Stercobilinogen.
      2. Mesobilinogen.
      3. Urobilinogen.
    2. Up to 20% urobilinogen produced is reabsorbed and enters the enterohepatic circulation.
    3. Most of the urobilinogen is reabsorbed is taken up by the liver and excreted in the bile.
    4. Small fraction  2 to 5% goes into general circulation and appears in the urine.
    5. The bilirubin conjugate with the glucuronic acid. This process takes place in the liver and gives rise to conjugated or direct bilirubin. Now conjugated bilirubin is no more bound to protein.


  1. Bile is formed in the liver and it consists of Bile salts, phospholipids, cholesterol, bicarbonate, bilirubin and water.
    1. Primary bile acids are Cholic acid and chenodeoxycholic acid. 
    2. Secondary bile acids are Deoxycholic acid and Lithocholic acid. These are formed by the intestinal bacteria.
    3. Both primary and secondary bile acids form the bile acid pool.
      1. Liver assists for the intestinal digestion by secreting 700 to 1200 ml of bile /day.
      2. Bile is bitter taste, alkaline, and yellowish-green in color.
      3. Bile consists of:
        1. Bile salt (conjugated bile acids).
        2. Bilirubin.
        3. Cholesterol.
        4. Electrolytes.
        5. Water.
      4. Bilirubin is one component of bile which is transported from the liver and stored in the gallbladder and from there delivered to the intestine.
      5. In the bile:
        1. Bilirubin diglucuronide is ∼90%.
        2. Bilirubin monoglucuronide is ∼10%.
      6. Functions of bile:
        1. Intestinal fat emulsification.
        2. Absorption of the fat.
          1. Bile has two fractions:
          2. Bile acid-dependant fraction which consists of bile acids, cholesterol, lecithin (phospholipids), and bilirubin. This fraction is secreted by the hepatocytes.
          3. Bile acid-independent fraction which is secreted by the hepatocytes and epithelial cells of bile canaliculi is bicarbonate-rich aqueous fluid which gives bile alkaline pH. 
  1. Jaundice appears when the bilirubin level is above 2.5 mg/dl.
    1. In the newborn when the liver can not conjugate bilirubin and if the level increases then this indirect bilirubin can cross the blood-brain barrier and leads to toxic injury to the brain and called Kernicterus.
    2. If the bilirubin level exceeds 15 mg/dl then immediately start the treatment to avoid the brain damage.
    3. While physiologic jaundice appears after 3 to 4 days and subsidies itself.

Normal adult values

  1. Total bilirubin = 0.3 to 1.0 mg/dL or 5.1 to 17.0 mmol/L
  2. Direct bilirubin  (conjugated bilirubin) = 0.1 to 0.3 mg/dL or 1.0 to 5.1 mmol/L
  3. Indirect bilirubin (unconjugated bilirubin) = (total bilirubin minus direct bilirubin level) =  0.2 to 0.7 mg/dL or 3.4 to 11.9 mmol/L
  4. Total bilirubin in:
    1. Umbilical cord blood = less than 2 mg/ dl.
    2. Infants
      1. 0 to 1 days = less than 6 mg/ dl
      2. 0 to 2 days = less than 8 mg/ dl
      3. 3 to 5 days = less than 12 mg/dl
      4. after 5 days = less than .2 to 1.0 mg/ dl
    3. Urine is negative for bilirubin.
      1. For conversion to SI unit multiply by 17.1 and report as  µmol/L (mmol/L)

Another source Total bilirubin level

Age Premature  mg/dL Full-term   mg/dL Adult   mg/dL
Cord blood <2 mg <2.0  
0 to 1 day <8.0 1.4 to 8.7  
1 to 2 days <12.0 3.4 to 11.5  
3 to 5 days <16.0 1.5 to 12.0  
5 days to 60 years     0.3 to 1.2
60 to 90 year     0.2 to 1.1
>90 year     0.2 to 0.9

Physiologic jaundice of the newborn:

  1. This is seen in the newborn where the liver is immature and lacks a sufficient amount of conjugating enzymes. This will lead to an increased amount of unconjugated bilirubin.
  2. This unconjugated bilirubin can cross the blood-brain barrier and give rise to encephalopathy (Kernicterus).
  3. Bilirubin above 15 mg /dl in the newborn needs immediate treatment.
    1. This is treated by the exchange of blood transfusion or light photo-therapy.
  4. Bilirubin levels that may require treatment in a full-term, healthy baby:

    1. 24 hours or younger infant need treatment when the bilirubin level is More than 10 mg/dL or more than 170 mmol/L.
    2. 25 to 48 hours infant need treatment when bilirubin is more than 15 mg/dL or more than 255 mmol/L.
    3. 49 to 72 hours of infant needs treatment when bilirubin is more than 18 mg/dL or more than 305 mmol/L
    4. Older than 72 hours of infant needs treatment when the bilirubin level is more than 20 mg/dL or more than 340 mmol/L 

Raised Bilirubin level is seen in:

  1. Some infections, such as an infected gallbladder, or cholecystitis.
  2. Some inherited diseases, such as Gilbert’s syndrome. 
    1. Although jaundice may occur in some people with Gilbert’s syndrome, the condition is not harmful.
  3. Diseases that cause liver damage, such as hepatitis, cirrhosis, or mononucleosis.
  4. Diseases that cause blockage of the bile ducts, such as gallstones or cancer of the pancreas.
  5. Rapid destruction of red blood cells in the blood, such as from sickle cell disease or an allergic reaction to blood received during a transfusion (called a transfusion reaction).
  6. Medicines that may increase bilirubin levels. This includes many antibiotics, some types of birth control pills, indomethacin (Indocin), phenytoin (Dilantin), diazepam (Valium), and flurazepam (Dalmane).

Decreased Bilirubin level is seen in:

  1. Medicines that may decrease bilirubin levels. This includes vitamin C (ascorbic acid), phenobarbital, and theophylline.

Causes of direct hyperbilirubinemia:

Causes of Indirect hyperbilirubinemia:

Types of jaundice may be of following types.

  1. Pre-hepatic jaundice. The etiology is before the liver like increased hemolysis of RBC.
  2. Hepatic jaundice. Now the causes are in the liver like hepatitis.
  3. Post-hepatic jaundice. Where the cause is after the liver like gallstone, cancers and these are the obstructive type of jaundice.

Table showing the Different type of Jaundice and their causes

Type of Jaundice Causes Pathophysiology



Extravascular hemolysis Heme is converted to unconjugated bilirubin

Right heart failure and Cirrhosis

There is a defect in the delivery of unconjugated bilirubin to liver

Hypothyroidism and Crigler-Najjar syndrome

There is a defect in the conjugation of bilirubin in the hepatocytes

Drugs like Rifampicin  and Gilbert syndrome

There is a defect in the uptake of unconjugated bilirubin into hepatocytes



Mechanical obstruction by tumors, stones or strictures, and primary biliary cirrhosis

There is a defect in the flow of conjugated bilirubin through canaliculi and bile ducts. Called cholestatic jaundice.

Drugs like estrogen and cyclosporine

Dubin-Johnson syndrome


There is a defect in the transmembrane secretion of conjugated bilirubin into the canaliculus. This is also called hepatocellular jaundice 

  Sepsis Like above

Possible References Used

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