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Fouchet's Test 3

This document describes methods for the qualitative determination of bilirubin in urine. It discusses (1) the foam test, Gmelin's test, Lugol's iodine test, and reagent strip tests to detect bilirubin, (2) the ingredients and principles of Fouchet's reagent test for bilirubin, and (3) the procedure for using Fouchet's reagent to test urine samples, which involves adding barium chloride and observing the development of a green color as indicating the presence of bilirubin.

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538 views5 pages

Fouchet's Test 3

This document describes methods for the qualitative determination of bilirubin in urine. It discusses (1) the foam test, Gmelin's test, Lugol's iodine test, and reagent strip tests to detect bilirubin, (2) the ingredients and principles of Fouchet's reagent test for bilirubin, and (3) the procedure for using Fouchet's reagent to test urine samples, which involves adding barium chloride and observing the development of a green color as indicating the presence of bilirubin.

Uploaded by

david mchembe
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© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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ST JOHN’S UNIVERSITY OF TANZANIA

TOPIC: QUALITATIVE DETERMINATION OF BILIRUBIN IN URINE

Introduction

• Bilirubin is an orange-yellow pigment formed in the liver by the breakdown of


haemoglobin and excreted in bile.
• This occurs during the normal breakdown of red blood cells.
• High bilirubin levels cause yellow pigmentation of the skin and/or sclera. This condition
is called jaundice.

 Conditions associated with bilirubinemia/bilirubinuria include liver diseases (liver


cirrhosis, hepatitis,), increased RBC destruction (sickle anemia, severe malaria and other
forms of hemolytic anemia) and diseases involving the gall bladder.
 Clinical findings include; Urine color becomes dark yellow with yellow foam and
Elevated serumconjugated bilirubin.

Methods for detection of bilirubin in urine

1) Foam test
2) Gmelin’s test
3) Lugol iodine test,
4) Fouchet’s test
5) Ictotest tablet test, and
6) Reagent strip test.
1. Foam test: About 5 ml of urine in a test tube is shaken and observed for development of
yellowish foam. Similar result is also obtained with proteins and highly concentrated
urine. In normal urine, foam is white.
2. Gmelin’s test: Take 3 ml of concentrated nitric acid in a test tube and slowly place equal
quantity of urine over it. The tube is shaken gently; play of colors (yellow, red, violet,
blue, and green) indicates positive test (Figure 823.1).
3. Lugol iodine test: Take 4 ml of Lugol iodine solution (Iodine 1 gm, potassium iodide 2
gm, and distilled water to make 100 ml) in a test tube and add 4 drops of urine. Mix by
shaking. Development of green color indicates positive test.
4. Reagent strips or tablets impregnated with diazo reagent: These tests are based on
reaction of bilirubin with diazo reagent; color change is proportional to the concentration
of bilirubin. Tablets (Ictotest) detect 0.05-0.1 mg of bilirubin/dl of urine; reagent strip
tests are less sensitive (0.5 mg/dl).

Fouchet's Reagent
Ingredients
 Trichloroacetic acid 25.00 gm
 Ferric chloride,10% aqueous 10.00 ml

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ST JOHN’S UNIVERSITY OF TANZANIA

 Distilled water 100.00 ml


Principle and Interpretation
Barium chloride precipitates the sulphate radicals present in urine to form precipitate of barium
sulphate. If bile pigments are present in urine, they adhere to these molecules. Ferric chloride
present in Fouchet’s reagent then oxidizes yellow bilirubin, in the presence of trichloroacetic
acid to green biliverdin. Therefore the development of green color due to the formation of
biliverdin indicates the presence of bilirubin (bile) in urine.

Procedure
1. Take three test tubes, clean with tape water.
2. Label the test tubes as follows; PN (for control normal), PC (for control positive) and T
(for test)
3. Take 5 ml of fresh urine in a test tube (normal, positive and patient)
4. Add 2.5 ml of 10% of barium chloride in each test.
5. Mix well, precipitateof sulphates appears to which bilirubin is bound (barium sulphate-
bilirubin complex)
6. Filter to obtain the precipitate on a filter paper.
7. Unfold a filter paper on an absorbent paper.
8. To the precipitate on the filter paper, add 1 drop of Fouchet’s reagent
9. Immediate development of blue-green color around the drop indicates presence of
bilirubin.
10. Yellowish white indicates the absence of bilirubin.
Results
Development of green color due to formation of biliverdin indicates the presence of bile
(bilirubin) in the urine.

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ST JOHN’S UNIVERSITY OF TANZANIA

Figure 823.1 Positive Gmelin’s test for bilirubin showing play of colors

Figure 823.2 Positive Fouchet’s test for bilirubin in urine

NOTE:Bilirubin is converted to non-reactive biliverdin on exposure to light (daylight or


fluorescent light) and on standing at room temperature. Biliverdin cannot be detected by tests
that detect bilirubin. Therefore fresh sample that is kept protected from light is required.

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ST JOHN’S UNIVERSITY OF TANZANIA

Review questions

Qualitative Determination of Bilirubin in Urine

1. Mention the name of test procedure:


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2. List the requirements needed for the test

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3. State the principle of the test

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4. Explain the reporting system of the test results

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5. Explain the reference value for bilirubin in urine.

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ST JOHN’S UNIVERSITY OF TANZANIA

6. Explain the limitation of the test results.

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