COMPETENCY NO.BI11.3 & BI11.

4
PHYSICAL CHARACTERISTICS & CHEMICAL CONSTITUENTS OF NORMAL URINE
Urine is normally composed of:
1) 99% water
2) 1% solids which are:
a) Inorganic salts: Inorganic sulphates, phosphates and chlorides of Na/K,
salts of ammonia
b) Nitrogenous organic compounds like urea, uric acid, creatinine, hippuric
acid
c) Organic Sulphur containing compounds like ethereal sulphates.
➢ Physical characteristics:
1) Volume: Normal volume of urine excreted is 800-1800mL/24hrs.
2) Colour: Normal urine is pale yellow due to pigment urochrome.
3) Appearance: Freshly voided urine is clear but it turns turbid on standing due
to precipitation of phosphates and urates.
4) Odour: Normally aromatic but turns ammonical on standing due to the
presence of urea splitting organisms liberating ammonia.
5) Specific gravity: It is measured using an instrument called urinometer. It is
calibrated at 150C or 200C.So temperature correction has to be applied for
room temperature as follows:
a) For every 30C rise in room temperature, add 0.001to the observed
specific gravity.
b) For every 30C fall in room temperature, subtract 0.001 from the observed
specific gravity.
c) Normal specific gravity is 1.015 to 1.025
 Procedure for measuring Specific gravity of Urine:
a. Take cylindrical flask & fill it up to 100 ml marking with urine.
b. Then put urinometer in this flask so that it does not touch the walls of
flask.
c. This urinometer contains marking of 0,1.010,1.020,1.030 and so on.
d. Lower meniscus is read as observed specific gravity and temperature
correction is to be done and then calculate final corrected specific
gravity.
6) Total solids: Last 2 digits of corrected specific gravity are multiplied by 2.66
(Long’s coefficient). The unit for total solids is Gm/L.
7) pH: It is slightly acidic (approximately 6.00), when fresh, and becomes
alkaline on standing because urea is converted to ammonia. Very often urine
excreted after ingestion of vegetarian diet is alkaline and that excreted after
intake of non-vegetarian diet is acidic.
➢ Chemical constituents:
1) Inorganic constituents:
Cations like Na+ , K+, Ca2+ and Mg2+ are excreted as salts in association with
anions like SO42-, PO4-3 etc.
a) Chlorides: Chlorides are excreted mainly in the form of sodium
chloride.
b) Sulphates: Sulphur is ingested through Sulphur containing amino acids
like cystine, cysteine, methionine and also through chondroitin
sulphates. Sulphur is metabolized and excreted in two forms: ethereal
sulphates and inorganic sulphates.
c) Phosphates: Phosphates are derived chiefly from the metabolism of
phosphorus containing foodstuffs and tissue components like
phosphoproteins, nucleoproteins, nucleotides and phospholipids. The
quantity excreted is extremely variable as it depends on the nature of
the diet. Phosphorus is excreted in urine largely as inorganic
phosphate and to small extent in organic form.
d) Calcium: Under normal dietary intake, urinary excretion is 15-40% of
total calcium and remaining is excreted in faeces. An average of 200-
300 mg of calcium is excreted daily by an adult.
 e) Ammonia: Under normal dietary conditions, an adult excretes 0.3-1.2
grams of ammonia nitrogen daily and is derived from dietary amino
acids.
2)Organic constituents:
a) Ethereal sulphates: They consist of sodium and potassium salts of
sulphuric acid esters of phenols like indoxyl, skatoxyl, phenol and cresol.
These are detoxication compounds of phenols and are found in liver.
b) Urea: It is the principal end product of protein (amino acid) metabolism.
About 30 grams of urea is excreted per day. The amount excreted
depends on protein intake.
c) Uric acid: It is the end product of purine nucleotide catabolism. The
quantity of uric acid excreted in urine varies from 0.5 to 1 gram/24 hours.
d) Creatinine: It is an anhydride of creatine. Creatine is formed from
transamination of the amino acids, glycine, arginine and methionine in
the liver. Creatine conversion to phosphocreatine is catalysed by creatine
kinase; spontaneous formation of creatinine occurs during the reaction.
The amount of creatinine excreted in urine is 0.6-1.2gm/24 hours. It is
purely endogenous and does not depend on the dietary intake of proteins
but in turn depends on the muscle mass.
e) Hippuric acid: It is a detoxication product of benzoic acid with glycine. The
benzoic acid is present in many fruits, vegetables and also used as food
preservative.
➢ Physical characteristics:
S.No. Tests Observations
1 Volume
2 Colour
3 Appearance
4 Odour
5 pH
6 Specific gravity
Room temperature =
Calibration temperature =
Temperature difference =
Observed specific gravity =
Corrected Specific Gravity = Observed Specific gravity +
(0.001 x Temp.diff.)
3
 7 Total solids(gm/L)
= Last two digits of sp.Gr.x 2.66 (Long′ s Coefficient )

➢ Chemical tests-
A. Organic constituents

1. Test for urea:

Test Observations Principle
Specific urease test:

3 ml urine + a drop of The solution will Urea is hydrolysed by urease

phenolphthalein + a pinch of urease
powder. Shake the contents of the
tube and allow to stand for 10
minutes.
2. Test for creatinine:
Test
Jaffe’s Test:
3ml saturated picric acid solution + 3ml 5% NaOH solution. Mix and divide into two equal parts.
1st part + 2ml urine
develop intense pink enzyme to form ammonium
colour after 10 minutes. carbonate which makes the
solution alkaline. Hence,
phenolphthalein turns pink.
Observations Principle
Orange red color Creatinine reacts with picric acid in
alkaline medium to form orange red
colored creatinine picrate.

2nd part + 2 ml distilled water Yellow colour Serves as control

3. Tests for uric acid

Make urine alkaline by adding 1ml of 2% sodium carbonate to 3 ml urine in a test tube and use alkaline
urine for the following tests.

Tests Observation Principle

a. Schiff’s test:

Add 2-3 drops of AgNO3 on filter In alkaline medium, uric acid

paper. Add 5-6 drops of urine reduces AgNO3 to metallic silver.
Greyish/Black spots develop
(alkaline). Warm gently if necessary.

b. Benedict’s uric acid test:

3ml urine (alkaline) + 0.5 ml of Uric acid reduces phosphotungstic
Benedict’s uric acid reagent and acid, present in Benedict’s uric acid
Blue color
mix the content by shaking the reagent in alkaline medium to blue
tube. colored tungstous salt.
4.Test for Hippuric acid:
Test Observation Principle
3 ml urine + 1 ml of 5 % NaOH + Cream coloured Formation of cream coloured
dilute FeCl3 solution (1 drop of precipitate ferric hippurate.
FeCl3 solution in 10 ml D/W) drop
by drop.
5. Test for ethereal sulphate:
Test Observation Principle

5 ml urine + 5 ml Baryta mixture White turbidity Ethereal sulphates are dissociated

(Barium nitrate + Barium hydroxide). on boiling with conc. HCl to liberate
Filter. To the filtrate, add 2ml conc. inorganic sulphates, which form
HCl. Boil for 2 minutes and cool at white precipitate of barium
room temperature. sulphate.

B. Inorganic constituents:

*1. Test for ammonia (Inorganic nitrogenous constituent)

Test Observation Principle

3 ml urine + 1 ml of 5% NaOH.
Heat. When fumes start
appearing, hold the filter paper
premoistened with 3-4 drops of
phenolphthalein close to the
mouth of the test tube.
*Unstable tests
Pink spot appears on the Salts of ammonia are unstable
filter paper which and on heating in alkaline
disappears quickly. medium, decompose to form
vapours of NH3 which being
alkaline turn phenolphthalein
pink.
2. Test for calcium and phosphates:

Test Observation Principle

10 ml urine + 3ml strong
ammonia dropwise. Boil. Cool. Gelatinous white precipitate Precipitate of Ca 3 (PO4 )2
Filter. Discard filtrate. (Scanty)
Add 5ml of 1:5 hot acetic acid on
the precipitate on filter paper.
Collect the solution in a test tube
and divide the solution into 2
parts.
Perform tests for Ca2+ and PO43-
as follows:
a. Test for calcium
1st part + 2ml ammonium oxalate
White turbidity
or potassium oxalate Calcium present gives white
b. Test for PO43- precipitate of calcium oxalate
2nd part + 1ml conc. HNO3 + 5 ml
ammonium molybdate. Boil.
Phosphates present react with
ammonium molybdate to form
Canary yellow precipitate
ammonium phosphomolybdate

3.Test for Chlorides:

Test Observation Principle

3 ml urine + 1 ml conc. HNO 3 + Curdy white precipitate Chlorides are precipitated as AgCl.
1 ml AgNO3 (HNO3 is added to keep phosphates
in solution)

4. Test for Inorganic Sulphates

Test Observation Principle

3 ml urine + 1 ml conc. HCl + 1 Curdy white precipitate Inorganic sulphates are

ml Barium Chloride (2%) precipitated as BaSO4. HCl is added
to keep phosphates in solution.

Assessment Type: Qualitative experiment