|
 Appearance
Normal urine colour varies
from light yellow to deep amber. Urine colour
sometimes may vary depending upon the drink, if any, consumed by the patient.
The colour of urine is sometimes related to a
pigment called "urochrome". The degree of
colour also depends on whether the specimen is
concentrated or dilute.
Normal urine is usually clear. If the pH is alkaline,
turbidity may be observed due to the precipitation of phosphates. Such urine
should be centrifuged before analysis. Turbidity due to the presence of chyle (chylomicrons) cannot be
centrifuged, but requires filtration using a special cellulose filter having
<0.1 m m diameter.
 pH
Use a narrow range pH paper or a pH meter.
In some clinical situations, measurements of approximate
pH within + 0.5 pH units using a narrow-range pH paper or exact pH
using a pH meter may be very helpful.
Procedure
 Using
pH paper
Put a drop of urine on a portion of pH indicator paper.
The colour obtained is compared with a standard
chart. For checking the reliability of the pH paper cross check the pH of
buffer solutions of known pH values having acidic and alkaline pH ranges.
Using a pH meter
Calibrate the pH meter using standard buffers, one
having an acidic pH and the other an alkaline pH, preferably less than 9.0.
Pour about 40-50ml of urine into a 100ml beaker, calibrate the pH meter once
again using the buffer of pH 7.0 and measure the pH of the urine.
Result
Normal urine pH ranges from 4.5 to 8.0.
The urine pH values are reported for example as 6.0 if
pH paper is used or as 6.1 if a pH meter is used.
Interpretation and quality control
Urine pH is usually acidic in normal people, especially
non-vegetarians, and is usually alkaline in vegetarians.
An early morning urine pH <5.5 indicates that renal
tubular acidification mechanism is intact.
As a quality control measure, use certified reference
buffers (commercial source), one in acidic range, say, pH 4.0 and the other
in alkaline range, preferably pH 9.2 to check the reliability of the pH paper
used or to assess the performance of the pH meter.
Always use a pH indicator paper before the date of
expiry. Do not use outdated pH papers. Always close the bottle containing the
pH paper tightly.
The pH meter must be calibrated against a correct
calibration buffer.
Ketone
Bodies - Rothera's test
The three main ketone bodies
are acetone , acetoacetic
acid (diacetic acid) and beta-hydroxybutyric
acid. Testing for ketone bodies should be done on
fresh urine or the specimen kept at 4 0C.
Principle
Acetone and acetoacetic acid
react with sodium nitropruside in the presence of
alkali to produce a purple colour.
Reagents
Rothera's Reagent :
Dry mixture
Pulve210rize 7.5g sodium nitropruside
with 200g ammonium sulphate. Store in a clean amber
bottle at 250-350C. Stable for 6 months.
Ammonia
concentrated, specific gravity 0.91
Procedure
To about 5ml of urine taken in an 18 x 150mm glass tube,
add about one teaspoon of the mixture, mix well, then
add 0.5 to 1.0 ml of concentrated ammonia down to the side of the tube so
that it layers on top of the urine. Observe for any colour
change within 30-60 seconds.
Result
If acetone and diacetic acid
are present, then a purple (permanganate calomel red) colour
will form at the junction of the two layers within 30-60 seconds. The result
can be graded from trace to 3+ based on the intensity of the colour formed, as detailed below.
No
change in colour - Negative
Pinkish ring - +
Red ring - ++
Deep purple ring - +++
Interpretation and quality control
Ketone bodies are intermediary products
of fat metabolism and their presence in the blood and then in the urine are
indications that the metabolism is disordered or incomplete. This is
associated with metabolic acidosis. This occurs in poorly controlled diabetes
mellitus and also in starvation.
Normal urine does not contain methyl ketone.
Weak false positive reactions may occur if the urine contains L-dopa and
phenyl pyruvic acid.
If there is suspicion of a false positive test, heat the
urine in a test tube in a bunsen burner flame for
one minute, allow to cool and repeat the Rothera's test. Heated urine will not give a positive Rothera's due to ketone bodies.
As a quality control measure, the reagent should be
checked frequently using a positive control (1-2 drops of acetone is added to
5ml of urine). The use of distilled water in place of urine for negative
control isrecommended.
Urobilinogen
– Erhlich’s test
Principle
Erhlich's reagent in conc
HCI. reacts with urobilinogen
to form a pink coloured aldehyde
complex in chloroform.
Reagents
Erhlich's reagent
Dissolve 2 g of P-dimethyl aminobenzaldehyde in 100ml of 20% HCI. Store at 25-350C
in an amber coloured bottle. Stable for 3 months.
10g/dl
Barium chloride
Dissolve 10g barium chloride in 100ml of distilled
water. Store at 25-350C. Stable for 6 months.
Saturated ammonium sulphate
Chloroform AR or GR quality
Procedure
To 12 ml of urine taken in a 25ml or 50 ml measuring
cylinder add 3 ml of barium chloride followed by 3 drops of saturated
ammonium sulphate. Mix well. Transfer a portion of
it into a 15 x 120mm glass tube. Centrifuge for 5 minutes at 3500 rpm.
Transfer about 5ml of the supernatant into an 18 x 150 mm glass tube and add
0.5 ml of Erhlich's reagent. Mix well. Then add 3ml
chloroform and shake well. Allow to stand one minute. Observe for any colour change in the chloroform layer (bottom layer).
Result
Urobilinogen
Colourless : Not detected
Faint red colour : Normal
Red or bright red : Positive or highly positive depending
upon the intensity of colour.
Interpretation and quality control
Urobilinogen is normally excreted in trace
and a normal urine will always show a faint red colour in the chloroform layer. It is always a good
practice to run a normal urine as control whenever
an urobilinogen test is done. Excess urobilinogen is seen in urine in haemolytic
jaundice, viral hepatitis and cirrhosis and is absent in obstructive
jaundice.
Bilirubin
- (Harison spot test) Fouchet's
test
Principle
When ferric chloride in acid solution is added to a
precipitate (Ref : Urobilinogen
procedure) of urine containing bilirubin, a green colour is produced as the bilirubin
in the urine is oxidized to biliverdin.
Reagent
Fouchet's reagent.
Dissolve 25g of trichloroacetic
acid in about 50ml of distilled water, then add 1g
ferric chloride, mix to dissolve and then make up to 100ml with distilled
water. Store at 25 – 350C. Stable for 6 months.
Procedure
To 12 ml of urine taken in a 25 ml or 50 ml measuring
cylinder add 3 ml of barium chloride followed by 3 drops of saturated
ammonium sulphate. Mix well. Transfer a portion of
it into a 15 x 120mm glass tube. Centrifuge for 5 minutes at 3500 rpm. Decant
the supernatant and add 1or 2 drops of Fouchet's
reagent to the precipitate. Examine for any colour
change.
Result
No colour change in the precipitate :
Negative
Appearance of a green or blue colour : Positive
Interpretation and quality control
Bilirubin isnot present in normal
urine. For a positive control, a few drops of either a bilirubin
standard or an icteric serum are added to a normal
urine sample and the specimen is analysed for the
presence of bilirubin. Any bilirubin
present in the urine is conjugated and indicates excess in the serum due to cholestasis.
|
