Estimation of total carbohydrates by

anthrone method
Experiment No: 1 Date: 12.08.2015

Aim: To estimate the amount of carbohydrate present

in the given sample by using Anthrone method
620 nm by using a spectrophotometer
Materials required

1. Anthrone reagent (2%): 2g of anthrone in 100 ml of

Conc. Sulfuric acid
2. Std. solution of glucose (100mcg/ml): 2 mg glucose
in 20 ml of d/w
3. Unknown sample
4. Boiling water bath
5. Colorimeter/ spectrophotomer
6. Graph sheets
7. Other lab consumables
1. Prepare anthrone reagent by dissolving 2g of
anthrone in 100 ml conc. H2SO4 and also
prepare a stock solution of glucose of a
concentration of 100 g/ml

2. Set up a range of serial dilutions of glucose of

10-100 g/ml in a volume of 1 ml each as
given in the Table. Similarly set up a blank
with 1 ml distilled water

3. Transfer 1 ml of serial dilutions of the

unknown sugar sample into separate test
tubes.
Add 4 ml of the anthrone reagent to each tube, mix,
and cover these with glass marbles. Incubate them
in a boiling water bath for 10 min.

Cool the tubes to room temperature and measure

the absorbance of the analyte at 620 nm using
blank.
Draw a standard curve by using the absorbance values
of the standard glucose tubes, that is by taking the
sugar concentration on the X-axis and the
corresponding absorbance value at 620 nm on the Y-
axis.

Calculate the amount of glucose in the unknown

sample from std.curve and record your observations