Good Example Of Report On Benedicts Test For Reducing Sugars

Reducing sugars are sugar molecules that have a free ketone or aldehyde group. All monosaccharide are reducing sugars. Disaccharides and polysaccharides can be reducing or non reducing. Sucrose is a non reducing sugar disaccharide. The C1 of the glucosyl unit is linked to the C2 of the fructosyl unit by a glycosidic bond to form sucrose. Since the reducing groups of C1 and C2 are involved in the glycosidic bond, sucrose is a non reducing sugar. Maltose on the other hand is a reducing sugar. The C1 of the sugar glucose is linked to C4 of another glucose by alpha (1—4) bond, to form maltose. Since reducing sugar at C2 of one of the glucosyl moiety is free in maltose, it is a reducing sugar. Benedicts test is a simple test to identify reducing sugar:
Principal: Benedict qualitative reagent is an alkaline copper reagent. The alkali converts reducing sugar into their enediol form. This enediol sugar has strong reducing properties. It then converts cupric ions in the reagent to cuprous ions, which reacts with hydroxyl group to form red color cuprous oxide. Thus in the presence of reducing sugar, the initial blue color of the reagent changes to brick red.
Glucose + alkali Enediol.
Enediol + Cu ++ Cu+ (cuprous) + sugar acid.
Cu+ + OH- CuOH Cu2O (Cuprous oxide-red)

Reagents:

1. Benedicts Reagent: Dissolve, 173 gm of Sodium citrate and 90 gm of anhydrous Na2CO3 in 500ml of distilled water. Slightly heat the solution to dissolve the content. Filter this solution and make up the volume to 850ml with distilled water. Dissolve separately 17.3 gm of CuSO4. 5H2O in 150 ml of water. Add this solution slowly, with stirring to the above solution and mix well. Benedicts reagent is ready for use. Sodium carbonate gives the alkaline nature to the reagent. Sodium citrate prevents the precipitation of cupric ions as cupric hydroxide by forming a loosely bound cupric –sodium citrate complex, which on dissociation gives a continuous supply of cupric ions.
2. Glucose (positive control) solution, equivalent to 1.0 % of reducing substance. One gram of glucose is dissolved in 100 ml of 1g/L benzoic acid. This solution is stable at room temperature for several months.
3. Negative control: 1g/L of Benzoic acid or distilled water.

Method:

1. Take 3 test tubes and place it on the test tube holder and mark each as negative control, test and positive control respectively.
2. Pippete 5 ml of Benedicts qualitative reagent in a test tube.
3. To this add 8 drops of negative control, test solution and positive control respectively.
4. Boil the test tubes over the flame or in a boiling water bath for 2 min.
5. Cool the solutions.
6. Observe the color of the reaction mixture in the test tube.
Result: The color of the solution or the precipitate gives an approximate amount of reducing sugar present in the test. Negative control will remain blue.
Figure1: A diagrammatic representation of different color in Benedicts qualitative test

Observation:

Discussion: Of the given test solution, potato juice, glucose solution, and reducing sugar are able to reduce cuprous to cupric in the benedicts reagents. Hence they are positive for reducing substances. Glucose is a reducing sugar, while starch is not a reducing sugar.
Conclusions: Reducing sugar is present in potato juice, glucose solution and reducing sugar solution.

References:

1. Chawla, Ranjna. Practical Clinical Biochemistry. 4th ed. Kundii: Jaypee Publishers,2014.Print.