Determination of urine glucose
using O-Toluidine
376 BCH
Miss Tahani Al-Shehri
Introduction
• Determination of urine glucose is important in
the diagnosis of diabetes mellitus.
Principle
• When heated with o-toluidine and glacial
acetic acid, glucose reacts with o-toluidine to
form N-glycosylamine. This compound has a
blue-green colour and its absorbance can be
measured at 625 nm.
O-toluidine
Method
1. Half fill 500 ml beaker with water and put it
on the hot plate to boil.
2. Take three clean test tubes and label them:
TEST, STANDARD and BLANK.
3. Add in each tube 5 ml o-toluidine (from a
burette).
Method
Cont..
4. Then, very accurately, pipette:
– 0.1 ml urine into the tube labeled TEST.
– 0.1 ml standard glucose solution into the tube
labeled STANDARD.
– 0.1 ml distilled water into the tube labeled BLANK.
5. Shake the tubes and put them in the boiling
water for 10 minutes. Remove the tubes, cool
them
6. Read the optical densities of the TEST and
STANDARD against the BLANK.
Calculations
Calculations

The concentration of glucose in the
standard solution is 100mg/100ml.
The concentration of glucose in urine is given
by:
O.D. TEST
––––––––––––––  100 =
O.D. STANDARD
mg
Glucose 100/ml urine