Khadga Bikram Angbuhang
GoldenGate International College
Principle:
Anthrone
reaction is the basis of a rapid and convenient methods for the determination
of hexose, aldopentoses either in free form or in when present as
polysaccharide. The reaction is not suitable if protein containing large amount
of tryptophan is present, since a red color is obtained under these conditions.
In
this assay, carbohydrates are dehydrated by using conc. H2SO4
to form furfural and its derivatives, which in turn condenses with anthrone to
form a bluish green complex with an absorption maximum at 620 nm.
D- glucose + conc. H2SO4 Hydroxymethyl
furfural +3H2O
Hydroxymethyl furfural + Anthrone Bluish green
complex
Materials and reagents:
- Anthrone reagents
- Conc. H2SO4
- Glucose standard 50 μg/ml
- Sample carbohydrates (Carbohydrates)
Procedure:
- Take 3 test tubes and label as Test, Standard and Blank.
- Add 1 ml Glucose sample in test labelled test tube, 1 ml glucose standard to standard test tube and 1 ml D/W (protein free) to blank.
- Add 4 ml of anthrone reagent to each tubes.
- Place all tubes in a boiling water bath for 10 minutes.
- Cool and read the absorbance at 620 nm against the blank.
Absorbance of sample X concentration of standard calculate the concentration as:
Concentration glucose (μg/ml): Absorbance of Standard.
