Urine for Urobilinogen and Ehrlich Reagent
Urine for Urobilinogen
Sample of urine for urobilinogen
- The test is done in the urine.
- A random sample can be taken.
- 2 hours fresh urine sample can be used, kept cool, and protect from the light.
- The sample is stable at 2 to 8 °C for 24 hours when urine is kept in the dark.
- Don’t expose urine to light or store it at room temperature.
- 2 to 3 hours postprandial after the noon meal is the best sample.
Purpose of the test (Indications)
- To assess liver function.
- This is the earliest sign of liver disease.
- This is positive in hemolytic anemia.
Precautions for estimation of urobilinogen in urine:
- False-positive test seen in:
- Drugs containing azo dyes:
- p-aminobenzoic acid.
- Highly colored pigments and their metabolites.
- False-negative test seen in:
- Oxidation of urobilinogen to urobilin.
- Formalin preservative.
Pathophysiology of Urobilinogen
- Hemoglobin breaks into bilirubin enter the intestine, and thereby the action of bacteria is transformed into urobilinogen.
- Urobilinogen in the intestine is excreted as urobilin, which gives a yellow color to urine.
- Urobilinogen in the intestine is reduced to brown color stercobilin, which gives color to stool.
- Urobilinogen is absorbed into the portal circulation, reaches the liver, is metabolized, and is excreted into the bile.
- Urobilinogen traces escape removal by the liver carried to the kidney excreted in the urine.
- Urobilinogen is colorless.
- The intestinal bacteria oxidize urobilinogen to Urobilin, which is a brown pigment.
- Its absence indicates obstructive jaundice.
Normal urobilinogen in urine
- Random sample = < 1 mg/dL.
- 24 hours sample = 0.5 to 4 mg/dL.
- Urine = 0.5 to 4.0 Ehrlich units/day.
- Feces = 75 to 400 Ehrlich units/day.
- Urine urobilinogen = 0.1 to 1.0 Ehrlich units /2 hours.
- One Ehrlich unit = approximately 1 mg of urobilinogen.
Procedure for urobilinogen in urine
- Add fresh urine sample to Ehrlich’s reagent.
- Ehrlich’s reagent:
- Urobilinogen + p-dimethylamino benzaldehyde (Erhlich’s reagent) = Form red color.
- Ascorbic acid is added as a reducing agent to keep urobilinogen in a reduced state.
- The addition of saturated sodium acetate stops the reaction.
- Preparation of the reagent Ehrlich’ reagent:
- p-Dimthylaminobenzaldehyde (4-dimethylaminobenzaldehyde) = 4 grams.
- Hydrochloric acid, concentrated = 40 mL.
- Distle water = 160 mL.
- Take the 4-dimethylaminobenzaldehyde dissolve in distle water.
- Add the conc HCL and mix well.
- This is stable at room temperature in a colored bottle, especially in dark brown. Protect from the daylight.
- The procedure of Ehrlich reaction:
- Take two test tubes.
- Label one as a test (T) and the other for comparison (B).
- Add 5 ml of freshly voided urine to each test tube.
- Add 0.5 mL of Ehrilich reagent to test (T).
- Add 0.5 mL of conc HCL to the B test tube and mix.
- Leave both test tubes at room temperature for 5 minutes.
- Note for the red color in the test (T) tube.
- For feces, the same method can be used except:
- Stool Aquos extract is treated with alkaline ferrous hydroxide to reduce urobilin to urobilinogen.
- Now add Ehrlich’s reagent.
Increased Urobilinogen is seen in:
- Cirrhosis due to:
- Alcoholic cirrhosis.
- Viral diseases.
- Biliary obstruction.
- Biliary tract infection, cholangitis.
- Hepatotoxic drugs e.g.aminosalicylic acid, Phenothiazine, and penicillin,
- Due to increased RBCs destruction:
- Hemolytic anemia.
- Pernicious anemia.
- Acute hepatitis.
- Pulmonary infarction.
- Excessive ecchymosis.
Decreased Urobilinogen is seen in:
- Biliary obstruction.
- Biliary tract infection (severe).
- Pancreatic cancer (head of the pancreas)
- Drugs, e.g., Aspirin, Chloramphenicol, Insulin, phenytoin, Streptomycin, tetracycline, Methyldopa, and sulphonamide.
Table showing the presence of urine bilirubin and urobilinogen in various conditions:
|Test||Normal person||Hemolytic anemia||Liver disease||biliary obstruction|
|urine bilirubin||negative||negative||positive/Negative||positive (+++)|
|urine urobilinogen||negative||positive (+++)||positive (++)||absent (Low)|
Normal urine picture:
|Physical features||Chemical features||Microscopic findings|