ABNORMAL CONSTITUENTS
OF URINE
IDENTIFICATION OF ABNORMAL CONSTITUENTS OF
URINE
•Physical examination
•COLOUR:
Red : Blood
Orange: Due to intake of riboflavin,
rifampicin
Black: Alkaptonuria
Green : Bile salts, Bile pigments.
•ODOUR:
Fruity: Glucose & ketone bodies
Garlic odour: After intake of spicy diet
Mousy odour : Phenylketonuria
•VOLUME:
• Polyuria: >2L/day
Seen in Diabetes mellitus, Diabetes Insipidus
•Oliguria : <500ml/day
Seen in kidney failure, circulatory collapse,
mismatched blood transfusion.
•Anuria: <50ml/day
shock, Acute tubular necrosis, Incompatible blood
transfusion
• Appearance :Turbid-in case of presence of proteins.
• Seen in urinary tract infections ,diabetic nephropathy
• Reaction to litmus: Blue-Red : Acidic
Red-Blue : Basic
• Urine is acidic after intake of non vegetarian diet & basic after intake of
vegetarian diet
• Specific gravity : Normal:1.020-1.030
Increased in Diabetes mellitus
Decreased in diabetes insipidus.
1. TEST FOR REDUCING SUGARS (GLUCOSE) IN URINE: BENEDICT’S TEST
•Significance: Identification of reducing sugars (glucose,
lactose, fructose)
•Principle: Reducing sugars in urine under hot alkaline
conditions tautomerise & forms enediols which are powerful
reducing agents & reduce cupric to cuprous ions. The cupric
hydroxide formed during this reaction is kept in solution by
chelators like citrate.
•Clinical significance: Positive in condition like diabetes
•Note: Thymol, formaldehyde, chloroform, lactic acid,
vit c & dextrin give false positive test with Benedict's
reagent.
1. BENEDICT’S TEST:
TEST
OBSERVATION
INFERENCE
Take 5ml of Benedict's Brick red precipitate is Reducing sugars in urine
reagent add 0.5ml(8drops) observed
under alkaline conditions
of urine. Hold the test tube
tautomerise
firmly & boil the content
enediols which reduces
for 2 mins over a stead
cupric ions to cuprous ions
flame.
(red)
(Heat intermittently)
to
form
2. TEST FOR KETONE BODIES IN URINE: ROTHERA’S TEST
• Significance: To detect presence of ketone bodies
• Principle : Acetone, Acetoacetic acid forms a purple
coloured complex with sodium nitroprusside in presence of
ammonia. The permanganate color is pressambly due to
formation of ferropentacyanide with isonitro compound of
ketone.
• Clinical interpretation: Normal level <1mg/day
• Increase ketone bodies excretion is seen in starvation
• Diabetic Keto Acidosis (DKA)
• Diet rich in fat but restricted in proteins ,carbohydrates
• Fever, severe anemia.
• Note : cannot be regarded as –ve until mixture had stood for
10 min without developing this colour.
2. ROTHERA’S TEST:
TEST
OBSERVATION
Saturate 2ml of urine with Permanganate
INFERENCE
pink The permanganate colour of
solid ammonium sulphate. colour ring is observed at solution is due to formation
Add 2 drops of freshly junction
prepared
nitropruside
5%
of
sodium ammonia layer
and
mix
gently. Layer 2 ml of
ammonia over urine.
urine
& of ferropentacyanide with
isonitro
ketone.
compound
of
3. TEST FOR BLOOD IN URINE: BENZIDINE TEST
• Significance: Identification of blood
pigments like haemoglobin & its
derivatives
• Principle: Heme of Hb decomposes
H2O2 into nascent O2 which oxidises
benzidine to bluish green colour
product .
• Clinical
interpretation
(Hematuria): Injury to urinary tract,
urinary calculi
• Benign/malignant tumour of kidney
&urinary tract.
• Ruptured venous plexus of enlarged
prostate.
3. BENZIDINE TEST:
TEST
OBSERVATION
INFERANCE
In a dry test tube dissolve Deep blue colour is (PSEUDOPEROXIDASE)
a pinch of benzidine in seen
Heme of Hb decomposes
1ml of glacial acetic acid
H2O2 into nascent oxygen
& add 1ml of H2O2to it.
which
Then add 5-10 drops of
benzidine to bluish colour
urine to it.
Control : Use distilled
water
oxidises
the
4. TESTS FOR PROTEINS IN URINE: HEAT & ACETIC ACID
TEST/ HEAT COAGULATION TEST
Significance : Identification of heat coaguble proteins.
Principle: when proteins in urine is heated, it is denatured when coaguble
proteins are heated, a series of changes occurs involving disruption of
quaternary, tertiary, secondary structure & making together of uncoded
polypeptide chains.
• Sulphosalicyclic acid test:
Significance: Identification of proteins in urine
Principle: Proteins are amphoteric, they behave as acids in alkaline
medium & bases in acidic medium. In presence of alkaloid reagents like
sulphosalicylic acid they act as bases & react with acid to form insoluble
salts of protein sulphosalicylate.
Clinical interpretation (Proteinuria): Pregnancy, nephrosis/nephritis,
Diabetes nephropathy, Bence Jones Proteinuria, Multiple myeloma,
Tuberculosis, Neoplasm, congestive cardiac failure.
4. HEAT & ACETIC ACID TEST/ HEAT COAGULATION TEST
TEST
OBSERVATION
INFERENCE
Heat and acetic acid test: White
coagulum
is When a protein in urine is
Fill ¾ of test tube with observed at upper layer
heated, it looses its
clear urine add 1 % acetic
precipitate
&
forms
acid. Mix well and heat
coagulum
upper 1/3 rd of test tube
over small flame.
Sulphosalicyclic
acid Turbidity is seen
test: Take 2ml of urine
sample add few drops of
20% sulphosalicyclic acid.
The proteins in urine in the
presence
of
alkaloid
reagent acts as bases to
form an insoluble protein
sulphosalicylate.
5. TEST FOR BILESALTS IN URINE: HAY’S TEST
• Significance: Identification of bile
salts in urine
• Principle: Bile salts lowers the
surface tension of urine &allows the
sulphur particles to sink
• Clinical Interpretation: Obstructive
jaundice
5. HAY’S TEST
TEST
Take 5ml of given
OBSERVATION
INFERENCE
T tube-sulphur powder Bile salts lower the
urine & sprinkle a little sinks to bottom
surface tension of
sulphur powder over
C tube-sulphur powder urine & allow sulphur
the urine. note if
floats
sulphur particles sink
Control: repeat with
H2O as control.
particles to sink
6. TEST FOR BILE PIGMENTS IN URINE: FOUCHET’S TEST
• Significance: Identification of bile pigments
in urine
• Principle: BaCl2 reacts with MgSO4 to form
BaSO4. The bilirubin present in urine adheres
to BaSO4
• Fouchet’s reagent : (FeCl3 in trichloracetic
acid) acts as an oxidising agent & oxidises
bilirubin (yellow) to biliverdin which is green
in colour.
• Note: Urine collected should be protected
from day light & fluorescent light as it is
rapidly oxidized by UV light to biliverdin
which is not detected by reagents used in the
test.
6. FOUCHET’S TEST
TEST
OBSERVATION
INFERENCE
To 5ml of urine, add 5ml of Greenish – blue green colour Bacl2 reacts with Mgso4 to
10%Bacl2, add 2-3 drops
of seen on filter paper
form Baso4.The bilirubin in
saturated Mgso4& mix well. Mix
urine adheres to precipitate & it
the contents &let the test tube
is detected by oxidation of
stand for precipitate to settle
bilirubin to biliverdin.
down. Decant the supernatant &
filter the rest. spread the filter
paper & add drops of Fouchet's
reagent to one part of precipitate
on filter paper.
ABNORMAL URINE ANALYSIS SAMPLE-1
I. Physical Examination:
1.Volume: More than 2500 ml/day
2.Colour: Pale yellow
3.Odour: Fruity
4.Appearance : Clear and Transparent
5.pH: Decreases (Acidic)
6.Specific Gravity: 1.030 at 15 °C
Correction Factor at 27 °C
= RT - 15
3 x0.001
= 0.004
Specific gravity at 27 °C =1.030+0.004
=1.034
II. Chemical Examination:
1.TEST FOR REDUCING SUGARS (GLUCOSE) IN URINE: BENEDICT’S TEST
TEST
OBSERVATION
INFERENCE
Take 5ml of Benedict's Brick red precipitate is Reducing sugars in urine
reagent add 0.5ml(8drops) observed
under alkaline conditions
of urine. Hold the test
tautomerise
tube firmly & boil the
enediols which reduces
content for 2 mins over a
cupric ions to cuprous
stead flame.
ions (red). Presence of
(Heat intermittently)
reducing sugar
to
form
2. TEST FOR KETONE BODIES IN URINE: ROTHERA’S TEST
TEST
OBSERVATION
INFERENCE
Saturate 2ml of urine with Permanganate pink colour The permanganate colour of
solid ammonium sulphate. ring is observed at junction solution is due to formation
Add 2 drops of freshly of urine & ammonia layer
of ferropentacyanide with
prepared
isonitro compound of
nitropruside
5%
and
sodium
mix
gently. Layer 2 ml of
ammonia over urine.
ketone.
Presence of Ketone bodies.
3. TEST FOR BLOOD IN URINE: BENZIDINE TEST
TEST
OBSERVATION
INFERENCE
In a dry test tube dissolve a Deep blue colour is not Blood is absent
pinch of benzidine in 1 ml seen
of glacial acetic acid & add
1 ml of H2O2 to it. Then
add 5-10 drops of urine it.
Controlwater.
Use
distilled
4. TESTS FOR PROTEINS IN URINE: HEAT & ACETIC ACID TEST/
HEAT COAGULATION TEST
TEST
OBSERVATION
INFERENCE
Heat and acetic acid test: White coagulum is not No Presence of Protein
Fill ¾ of test tube with clear observed at upper layer
urine add 1 % acetic acid.
Mix well and heat upper 1/3
rd of test tube over small
flame.
Sulphosalicyclic acid test: Turbidity is not seen
No Presence of Protein
Take 2ml of urine sample
add few drops of 20%
sulphosalicyclic acid.
5. TEST FOR BILESALTS IN URINE: HAY’S TEST
TEST
OBSERVATION
INFERENCE
Take 5ml of given urine & T tube - sulphur powder Absence of bile salts
sprinkle a little sulphur floats
powder over the urine. note C tube - sulphur powder
if sulphur particles sink
Control:
Repeat
H2O2 as control.
floats
with
6. TEST FOR BILE PIGMENTS IN URINE: FOUCHET’S TEST
TEST
OBSERVATION
To 5ml of urine, add 5ml of
No Greenish – blue green
10%Bacl2, add 2-3 drops of
colour seen on filter paper
saturated Mgso4& mix well. Mix the
contents &let the test tube stand for
precipitate to settle down. Decant
the supernatant & filter the rest
.spread the filter paper & add drops
of Fouchet's reagent to one part of
precipitate on filter paper.
INFERENCE
Absence of bile pigments
Compound
Test
Result
1. Carbohydrates
1. Benedict's test
2. Rothera’s test
+
+
2. Blood
3. Proteins
1. Benzidine test
1. Heat & Acetic acid test
(Heat coagulation test)
2. Sulphosalicyclic Acid test
-
1. Hays’s test
1. Fouchet’s test
-
4. Bile Salts
5. Bile Pigments
-
Diagnosis: As the given urine sample contains reducing sugars and ketone bodies
patient may be suffering form Diabetic Ketoacidosis.
• Report: The given urine sample contains reducing sugars, ketone
bodies , negative results for Blood, Proteins, Bile salts and Bile
Pigments.
• Reducing sugar: Excretion of glucose in urine is known as glycosuria.
• This is detected by Benedict's test.
• In normal subjects a very small amount glucose is excreted usually less
than 0.5% and can not be detected by benedict’s test.
• Excretion of abnormal amount of glucose in the urine may be divided into
two types, hyperglycemic and renal.
• Hyperglycemic glycosuria is due to increased blood glucose level beyond
renal threshold. Renal glycosuria is due to decreased renal absorption of
glucose by renal tubules due to renal diseases.
• Causes for hyperglycemic glycosuria:
• Alimentary glycosuria
• Nervous or emotional glycosuria
• Endocrinal Causes: Diabetes mellitus
• Causes for renal glycosuria: It may be hereditary due to Fanconi
Syndrome, Wilsons’s disease, hereditary Tyrosinemia etc. or may be
acquired due to renal diseases or may be due to heavy metal
poisoning. Lowering of renal threshold is physiologically noticed in
pregnancy.
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