Calorimetric Determination of Glucose by the 3,5-dinitrosalicylic acid Method Several reagents have been employed which assay sugars by using their reducing properties. This method tests for the presence of free carbonyl group (C=O), the so-called Reducing Sugars. This involves the oxidation of the aldehyde functional group present in glucose and the ketone functional group in fructose. oxidation CHO COOH

Calorimetric Determination of Glucose by the 3,5-dinitrosalicylic acid Method The chemistry of the reaction is complicated since standard curves do not always go through the origin and different sugars give different color yields. The method is therefore not suitable for the determination of a complex mixture of reducing sugar.

Materials 1. Standard Glucose Solution: 0.1g anhydrous glucose is dissolved in distilled water and then raised the volume to 100 ml with distilled water. 2. Dinitro salicylic acid reagent: a. Solution "a" is prepared by dissolving 300g of sodium potassium tartarate in about 500 ml distilled water. b. Solution "b" is prepared by dissolving 10 g of 3,5-DNS in 200 ml of 2N NaOH solution. c. The DNS reagent is prepared by mixing solutions a & b and raising the final volume to 1 litre with distilled water.

Read the Optical Density at 540 nm 60 30 15 45 5 10 20 25 35 40 50 55 Glucose (ml) 0 0.2 0.4 0.6 0.8 1 H2O (ml) 7 7 7 7 7 7 H2O (ml) 1 0.8 0.6 0.4 0.2 0 Shake Well COOL Read the Optical Density at 540 nm DNS (ml) 2 2 2 2 2 2

Results Samar A. Damiati