1. Enzymes and the digestive system
2.2 Carbohydrates – Test for reducing sugars
2.3 Carbohydrates – Test for non-reducing sugars
15/11/2018

2. Learning Objectives All students can...
Outline the stages of the Benedict’s test for reducing and non-reducing sugars.
Carry out the test for reducing and non-reducing sugars.
I can answer exam questions on testing for reducing and non-reducing sugars.
Specification reference: 3.1.2
15/11/2018

3. Success Criteria
I can describe the stages of the Benedict’s test for reducing and non-reducing sugars.
I can carry out the test for reducing sugars and non-reducing sugars.
Specification reference: 3.1.2
15/11/2018

4. Test for reducing sugars: Benedict’s test
All monosaccharides and some disaccharides (e.g. Maltose) are reducing sugars.
Reduction is a chemical reaction involving the gaining of electrons (OIL RIG).
A reducing sugar is a sugar that can donate electrons to (or reduce) another chemical, in this case Benedict’s reagent.
Benedict’s reagent is an alkaline solution of copper(II) sulphate.
When a reducing sugar is heated with Benedict’s reagent it forms an insoluble red precipitate of copper(I) oxide.

5. The Benedict’s Test
Add 2cm3 of the food sample to be tested to a test tube. If the sample is not already in liquid form, first grind it up in water.
Add an equal volume of Benedict’s reagent.
Heat the mixture in a gently boiling water bath for 5 minutes.
If a reducing sugar is present an orange-brown coloured precipitate is formed.

6. Semi-quantitative nature of the Benedict’s test
The table shows the relationship between the concentration of reducing sugar and the colour of the solution and precipitate formed during the Benedict’s test. The differences in colour mean that the Benedict’s test is semi-quantitative, that is it can be used to estimate the approximate amount of reducing sugar in a sample.
NONE
VERY LOW
LOW
MEDIUM
HIGH
Concentration of reducing sugar
Colour of solution and precipitate
None
Blue
Very low
Green
Low
Yellow
Medium
Brown
High
Red

7. Test for non-reducing sugars
Sucrose is a non-reducing sugar, because it doesn’t change the colour of Benedict’s reagent when heating with it.
In order to detect a non-reducing sugar, it must first be broken down into its constituent monosaccharides by hydrolysis.

8. Test for non-reducing sugars
If the sample is not already in liquid form, it must first be ground up in water.
Add 2cm3 of the food sample being tested to 2cm3 of Benedict’s reagent in a test tube.
Place the test tube in a gently boiling water bath for 5 minutes. If the Benedict’s reagent does not change colour (the solution remains blue), then a reducing sugar is not present.
Add another 2cm3 of the food sample to 2cm3 of dilute hydrochloric acid in a test tube and place the test tube in a gently boiling water bath for 5 minutes. The dilute hydrochloric will hydrolyse any disaccharide present into its constituent monosaccharides.
Slowly add some sodium hydrogencarbonate solution to the test tube to neutralise the hydrochloric acid. (Benedict’s reagent will not work in acidic conditions). Test with pH paper to check that the solution is alkaline.
Re-test the resulting solution by heating with 2cm3 of Benedict’s reagent in a gently boiling water bath for 5 minutes.
If a non-reducing sugar was present in the original sample, the Benedict’s reagent will now turn orange-brown. This is due to the reducing sugars that were produced from the hydrolysis of the non-reducing sugar.

9. Plenary Activity: Sugar Puzzle Practical
You have been given 3 sugar solutions and a protein solution: Glucose, Maltose, Sucrose and Albumen.
Your task is to work out which solution is which using the apparatus available.
Big Hint!!!!! The Benedict’s test is quantitative! Think about the number of reducing centres available and how this will affect the colour of the precipitate!

10. Apparatus Bunsen burner, tripod, gauze and heat proof mat 250ml beaker
Benedict’s reagent
10 Boiling-tubes
Dilute hydrochloric acid
Sodium hydrogencarbonate solution
7 pipettes (one for each chemical)
4 Conical flasks containing: Glucose solution, Maltose solution, Sucrose solution and Albumen (Protein) solution

11. Answers
A = Sucrose solution. A negative result is found when test for reducing sugar is carried out and a positive result is found when a test for a non-reducing sugar is carried out. This is because the reducing centres in sucrose are both in the glycosidic bond so cannot reduce Benedict’s reagent. When sucrose is hydrolysed into its constituent monosaccharides (glucose and fructose), two reducing centres become available. C
CH2OH OH H O 1 2 3 4 5 6
Reducing Centres OH H C
CH2OH O 1 2 3 4 5 6
1-2 Glycosidic bond

12. Answers
B = Maltose solution. A positive result is found when test for reducing sugar is carried out and a positive result is also found when a test for a non-reducing sugar is carried out. The precipitate formed after the non-reducing sugar test is carried out is darker because hydrolysis of the glycosidic bond has doubled the number of reducing centres. This is because one of the reducing centres in maltose is in the glycosidic bond so cannot reduce Benedict’s reagent. When maltose is hydrolysed into its constituent monosaccharides (glucose and glucose), two reducing centres become available. C
CH2OH OH H O 1 2 3 4 5 6
Reducing Centre C
CH2OH OH H O 1 2 3 4 5 6

13. Answers
C = Glucose solution. A positive result is found when test for reducing sugar is carried out and a positive result is also found when a test for a non-reducing sugar is carried out. The precipitate formed after the non-reducing sugar test is carried out is the same colour/intensity as the precipitate formed when the reducing sugar test is carried out. Hydrolysis does not create further reducing centres in this case.
CH2OH 6 H 5 C O H
Reducing Centre H C C 4 1 OH H OH OH C C 3 2 H OH

14. Answers
D = Albumen (protein) solution. A negative result is found when test for reducing sugar is carried out and a negative result is also found when a test for a non-reducing sugar is carried out. No coloured precipitate is formed and Benedict’s reagent remains blue.

15. Learning Objectives All students can...
Outline the stages of the Benedict’s test for reducing and non-reducing sugars.
Carry out the test for reducing and non-reducing sugars.
I can answer exam questions on testing for reducing and non-reducing sugars.
Specification reference: 3.1.2
15/11/2018