Enzymatic method for the determination of Sucrose, D-Fructose and D-Glucose. Based on the spectrophotometric measurement of NADPH produced though the reactions, after addition of hexokinase (HK), phosphoglucose isomerase (PGI) and glucose-6-phosphate dehydrogenase (G6PDH). The D-glucose concentration is determined before and after hydrolysis of sucrose by b-fructosidase. The sucrose content is calculated form the difference in D-glucose concentrations before and after hydrolysis by b-fructosidase.
Sucrose, D-fructose and D-glucose occur widely in plant organisms. In foods, they occur mainly in honey, wine and beer, and a range of solid foodstuffs such as bread and pastries, chocolate and candies. In the wine industry, the addition of sucrose is only allowed in few situations, such as champagne production.
This rapid and simple specific enzymatic method is used for the simultaneous determination of sucrose, D-fructose and D-glucose in foodstuffs, pharmaceuticals, cosmetics and biological samples.
Solution 1. Buffer (25 mL, pH 7.6). Stable for 2 years at 4 °C. Solution 2. NADP+ plus ATP and PVP. Stable for 2 years at - 20 °C. Dissolve in 22 mL of distilled water, divide into appropriately sized aliquots and store in PP tubes at -20 °C between use (stable for 2 years) and keep cool during use. Suspension 3. Hexokinase plus glucose-6-P dehydrogenase (4.1 mL). Stable for 2 years at 4 °C. Swirl bottle before use. Suspension 4. Phosphoglucose isomerase (2.25 mL). Stable for 2 years at 4 °C. Swirl bottle before use. Solution 5. D-Fructose/D-glucose standard solution (5 mL, 0.20 mg/mL of each sugar). Stable for 2 years at room temperature. This standard solution can be used when there is some doubt about the method accuracy. Solution 6. Buffer (20 mL, pH 4.6). Stable for 2 years at 4 °C. Suspension 7. β-Fructosidase (2.25 mL). Stable for 2 years at 4 °C. Swirl bottle before use. Add 0.02 ml of Suspension 7 plus 0.180 ml of Solution 6, per assay, to a test tube and homogenise (Solution 6+7). This solution should be prepared for each assay day. Warm Solution 6+7 to 25-30 ºC before use.
Compatible Sample Types:
Wine, beer, fruit juices, milk, dietetic foods, bread, jam, honey, ice-creams, fruit and vegetables, pharmaceuticals, cosmetics and biological samples.
Features & Benefits:
Rapid reactionStable reagents
Range: 20-800 mg/LDetection limit: 1.40 mg/L