ANALYSIS OF ABNORMAL URINE AND WRITING URINE REPORT

Colour:
• The urine colour may assume the colour of the drugs, or of their degradation products
upon the administration of various drugs.
• The Urine colour may be greenish yellow due to the excretion of large amounts of bile
pigments in the urine and may be reddish due to the presence of blood.
• In chyluria, the urine appears milky; and in alkaptonuria, the urine becomes dark on
standing.
Odour:
• Urine in Urinary tract infections has offensive odour.

Volume:
• Oliguria refers to a 24-h urine output of <500 mL, and anuria is the complete absence of
urine formation (<50mL). Polyuria is excretion of more than 3 Litres of urine per day
(>3 L/d).
• Anuria can be caused by total urinary tract obstruction, total renal artery or vein
occlusion, and shock
• Pathological conditions such as diabetes mellitus and diabetes insipidus cause increased
volume of urine.
Appearance
• The turbidity may also be due to the presence W.B.C, RBC or bacteria.
• Turbidity in a fresh sample is suggestive of infection. Excessive foaming of urine when
shaken suggests proteinuria.

Abnormal Constituents of urine:

The abnormal constituents which are routinely analyzed in urine are proteins (albumin,
Bence-jones protein), reducing sugars (usually glucose and in special cases lactose, galactose,
pentose and fructose), ketone bodies (acetone, acetoacetic acid), bile salts, bile pigments and
blood (Hb / erythrocytes).

Proteins:
Presence of detectable amount of protein in urine is known as proteinuria. Presence of
proteins leads to turbid appearance of urine. Albumin is commonest and earliest protein to get
excreted in urine. Most commonly albumin is seen in kidney diseases like
glomerulonephritis, nephritic syndrome. Bence Jones proteinuria is a condition seen in
multiple myeloma where in proteins gets precipitated between 400-600C and the precipitate
dissolves on boiling and reforms on cooling.

Carbohydrates:
Normally carbohydrates are absent in urine. Presence of sugar in urine is called as
Glycosuria. Glucosuria is presence of glucose in urine and occurs in diabetes mellitus and in

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renal diabetes. Other common examples of glycosuria are lactosuria (pregnancy and
lactation), Galactosuria (galactosemia), pentosuria and fructosuria.

Ketone Bodies:
Acetone, acetoacetic acid and β-hydroxy butyric acid are the ketone bodies. Increase in
ketone bodies causes keto acidosis, a clinical condition due to disturbed carbohydrate and
lipid metabolism.
Causes: Diabetic ketoacidosis, prolonged starvation.

Bile Salts:
Bile salts are produced from cholesterol and synthesized in liver. They reach GIT and help in
digestion and absorption of lipids by emulsification.
Bile salts appear in urine in obstructive jaundice.

Bile Pigments:
Bile pigments are derived from Hemoglobin. Bilirubin is conjugated in liver. Conjugated
bilirubin appears in urine in obstructive jaundice. Urobilinogen is normal constituent of urine.
Its level is increased in hemolytic jaundice and it is absent in obstructive jaundice.

Blood:
Presence of blood in urine is known as haematuria. Intact RBCs are excreted in urine. It is
usually observed in Glomerulonephritis, carcinoma, calculi, and injury to renal tract.
Hemoglobinuria is presence of hemoglobin in urine. It is caused by malaria and other
hemolytic disorders.

Principles and Procedures

Benedict’s reagent: Copper sulfate, Sodium carbonate, Sodium citrate
Principle of Benedict’s test: Reduction under alkaline condition:
Benedict’s reagent contains the milder alkali Na2CO3. When reducing sugar is heated with
Benedict’s reagent under alkaline condition, reducing sugar undergo tautomerization to form
1,2 – enediols. These enediols are unstable and give short chain aldehydes which are
powerful reducing agents. They reduce cupric ion in Benedict’s reagent to cuprous ion as
yellow cuprous hydroxide which is not easily soluble. In order to keep the cuprous hydroxide
in solution, a metal chelator like sodium citrate is added in Benedict’s reagent. During the
process of heating cuprous hydroxide is converted to red cuprous oxide.
2CuOH Cu2O + H2O
Clinical importance: Benedict’s test is called semi quantitative test because it gives different
colours with different percentage of sugar present in the solution.
0.5 % green precipitate.
1.0% yellow precipitate.
1.5% orange precipitate.
2.0% and above brick red precipitate.
Hence this test is used routinely in clinical lab to find out the percentage of sugar in the
urine.

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 Hellers Test: Protein reacts with acid to form meta-protein, which appears as a white ring at the
junction.
Heat Coagulation Test: When albumin is heated it undergoes irreversible denaturation to
form coagulum.

Precipitation by Alkaloidal Reagents: Negatively charged picrate, trichloroacetic acid,

sulphosalicylic acid neutralizes the positively charged protein and cause precipitation.

Test for Ketone bodies:

Rothera’s test:
Principle: Acetone and acetoacetic acid react with sodium nitropruside in the presence of
ammonia to produce a purple colour.
* β-hydroxybutyate does not answer Rothera’s test since it lacks the ketone group.

Gerhardt’s Test:
Principle: Ferric chloride reacts with acetoacetic acid to form ferric acetoacetate which is
wine red in colour.
* For Gerhardt’s test fresh urine sample should be used since acetoacetic acid is quickly
decomposed into acetone and CO2.

Test for Bile salts:

Hay’s Sulfur test:
Principle: Bile salts have the property of lowering the surface tension of solution in which
they are dissolved. This results in the sinking of sulphur powder.
* A false positive test may be obtained if the test tube is not cleaned properly and if there are
traces of soap or detergent, which can also lower the surface tension.

Test for Bile Pigments:

Fouchet’s Test:
Principle: BaCl2 reacts with MgSO4 to form BaSO4 precipitate. This binds with Bilirubin
present in the urine. Fouchet’s reagent oxidizes Bilirubin to green coloured biliverdin or blue
coloured bilicyanin products.
*Bile pigments are excreted in urine in obstructive jaundice and in any form of severe
hepatitis involving destruction of liver cells.

Test for presence of Blood

Benzidine Test:
Principle: Peroxidase enzyme present in RBCs decomposes hydrogen peroxide to nascent
oxygen. This liberated nascent oxygen oxidizes benzidine to blue or green coloured products,
which are quite unstable.
*The test is also positive when pus cells are present. However the test is negative when the
urine is first heated before testing and answers positive only if blood is present.

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ANALYSIS OF ABNORMAL URINE

Physical Properties:
Colour :
Odour :
Appearance :
pH :
Specific gravity :

Test Observation Inference

Test for albumin:
Heat and acetic acid test:
Take 10ml of urine, hold the test tube Cloudy white precipitate/ Albumin is
over a flame in a slanting position & coagulum are observed in present in urine.
boil the upper 1/3rdportion of urine. Add heated portion.
2-3 drops of 1% acetic acid.

Sulfosalicylic acid test:

To 3ml urine add 1ml of 20% White precipitate is Protein is present.
sulfosalicylic acid. formed.

Heller’s test:
Take 3ml of urine in a test tube and add White ring is formed at Protein is present
3ml of concentrated HNO3 slowly along the junction of two
the sides of the test tube without liquids
shaking

Test for Reducing sugars:

Benedict’s test:
Take 5ml of Benedict’s reagent + 8 Green/ Yellow/ orange/ Reducing sugar is
drops of urine. Mix well & boil for 2 Brick red precipitate is present.
mins over a small flame. formed.

Test for Ketone bodies:

a)Rothera’s test:
Acetone &
Saturate 5ml of urine with ammonium Purple coloured ring at
acetoacetic acid is
sulphate crystals. Add 2-3 drops of the junction of two
present.
freshly prepared 5% sodium liquids is observed.
nitroprusside solution. Mix well; add
1ml of strong ammonia solution along
the sides of the test tube carefully.

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 b) Gerhardt’s test for acetoacetic
acid:
To 5ml urine add 10% Ferric chloride Wine red colour is Acetoacetic acid
solution drop wise. formed. is present.

Test for Bile Salts:

Hay’s Sulphur test:
Label two test tubes as’ test’ and
‘control’.
Sulfur powder particles Bile salts are
Test: Take 5 ml urine in a Test tube and
slowly sink to bottom present
sprinkle a little sulphur powder over the
surface. Do not mix.
Sulfur particles float on
water
Control: Take 5 ml water in a test tube
and sprinkle a little sulphur powder. Do
not mix.

Test for Bile Pigments

Fouchet’s Test:
To 5 ml of urine, add 5 ml of 10%
Green or blue colour is Bile pigments are
BaCl2 and a pinch of MgSO4. Mix well.
formed. present
BaSO4 is Precipitated. After 5 min,
filter the solution and discard filtrate.
Spread the filter paper on a dry filter
paper and press lightly to remove
moisture. Add few drops of Fouchet’s
reagent on the dry precipitate.

Test for blood:

Benzidine test:
To 2 ml of urine add 3 drops of Blue or green colour
Blood is present
Benzidine solution; add 3 drops of develops and lasts for few
hydrogen peroxide. Mix. seconds.

Report:
The given urine contains pathological constituents like ____________________________

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 URINE REPORT

Name Date
Age Referred by Dr.
Sex Occupation
Physical Characteristics
Sl No Parameters
01 Volume
02 Appearance
03 Colour
04 Odour
04 pH
06 Specific gravity
Urinometer is calibrated at ……..0C Room temperature is ….....0C
Chemical Characteristics
Sl No Abnormal constituents Test Result
Heat coagulation test + / - ve
01 Proteins Heller’s test + / - ve
Sulfosalicylic acid test + / - ve
02 Blood Benzidine test + / - ve
03 Glucose Benedict’s test + / - ve
Rotheras’s test + / - ve
04 Ketone Bodies
Gerhardt’s test + / - ve
05 Bile Salts Hay’s sulphur test + / - ve
06 Bile pigments Fouchet’s test + / - ve

Report: The given urine contains pathological constituents like glucose and ketone bodies.
The given urine contains pathological constituents like protein and blood.
The given urine contains pathological constituents like bile salts and bile pigments.

Interpretation:

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QUESTIONNAIRE:

1. Benedict's test is also known as semi quantitative test, why?

2. What are the substances that answer Biuret test?
3. Name the tests that detect the ketone bodies in urine?
4. Name the Ketone Bodies? What are the conditions, in which Ketone bodies are
excreted in urine?
5. What is Ketosis? How the Ketone bodies are synthesized in the body. Where are they
synthesized? What is their use?
6. What is the condition with both glucose and Ketone bodies in urine?
7. What is Glycosuria? What is the specific test for glucose?
8. What is meant by proteinuria? Which protein is usually excreted in urine?
9. Name the causes of proteinuria?
10. How will you test for proteinuria?
11. What is Bence-Jones protein?
12. How will you detect blood in the urine?
13. What is haematuria, what is haemoglobinuria? How do you differentiate both?
14. In which diseases hematuria is manifested?
15. In which disease both haematuria and proteinuria coexist?
16. What are bile salts? Name the test for bile salts?
17. What are bile pigments? What are the tests to detect bile pigments in urine?
18. Which test detects the urobilinogen?
19. What is Jaundice? What are the types of jaundice?
20. In which type of jaundice bile salts and bile pigments are excreted in urine.
21. What is the normal serum bilirubin level?
22. What is conjugated bilirubin and what is unconjugated bilirubin.
23. What is the mechanism of Fouchet's test?
24. What are the precursors for the bile pigments and bile salts?
25. What is the name of the test by which bilirubin is estimated in serum?
26. How does Urobilinogen help in detecting the type of jaundice?

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