-Definition and Color Reactions of Reducing Sugars-

PROCEDURE

A) Benedict Test B) Tollens Test

1) Sodium citrate and anhydrous sodium 1) Solution A, consisting of a 5%
carbonate are dissolved in AgNO3 solution in distilled water,
approximately 800 ml of water in a and Solution B, a 5% KOH or NaOH
separate container. solution, are prepared.

2) Copper(II) sulfate pentahydrate is 2) Equal volumes of solutions A and B

dissolved in 100 ml of water in another are taken into separate test tubes.
container.
3) Upon mixing, white AgOH
precipitate is immediately formed.
3) Both solutions are mixed together.

4) The volume is diluted to 1 L with 4) NH4OH is added dropwise to the

distilled water. precipitate until complete dissolution
occurs.

5) If necessary, the solution is filtered to 5) The obtained silver diamine solution

ensure clarity. is combined with 1 ml of the sample
to be examined.
6) Benedict solution is taken into a test
tube.
6) The mixture is heated in a water bath.

7) 1 ml of the sample is added to the 7) The formation of a silver mirror on

Benedict solution. the inner surface of the tube is
observed.
8) The test tube is heated on a heater.

9) A reddish-brown precipitate of Cu2O

(copper II oxide) is observed.
 C) Picric Acid Test D) Methylene Blue Test

1) A 5% solution of picric acid in 1) Into a test tube, pure water (1 ml)

alcohol or a saturated solution and 1 drop of 1% methylene blue
of picric acid in water is are added.
prepared.
2) Then, 1 ml of 0.1 N NaOH is
2) A 10% solution of NaOH in added to the test tube.
distilled water is prepared.

3) When heated, no color change is

3) Into a test tube, the sample is observed.
added (1 ml), and 3-4 drops of
picric acid are dripped.

4) The test tube is heated in a

water bath.

5) During heating, 3-4 drops of

NaOH are dripped into the
solution.

6) The formation of a red color is

observed.
DATA AND CALCULATION

In the Benedict trial, a reddish-brown precipitate of Cu2O (copper II oxide) was observed due to the
presence of reducing sugars in the sample solution.

In the Tollens trial, the formation of a silver mirror on the inner surface of the tube indicated the
presence of reducing sugar.

During the Picric Acid trial, the appearance of a red color in the tube indicated the presence of reducing
sugar.

In the Methylene Blue experiment, initially, a blue color was observed in the tube. This color change
occurred because methylene blue was reduced in the presence of reducing sugars in the sample, leading
to the disappearance of the blue color. However, upon depletion of reducing sugars, shaking the tube
open allowed the reduced form of methylene blue to be oxidized by atmospheric oxygen, leading to the
reappearance of the blue color.

RESULT AND DISCUSSION

 The experiments conducted aimed to detect the presence of reducing sugars using four different
tests: Benedict test, Tollens test, Picric Acid test, and Methylene Blue test. In the Benedict test, a brick-
red precipitate of copper (I) oxide (Cu2O) was observed in the presence of reducing sugars. This color
change occurs due to the reduction of copper (II) ions to copper (I) ions, which forms the insoluble
Cu2O precipitate. The alkalinity required for this reaction is provided by sodium carbonate, while
sodium citrate prevents premature reduction of copper (II) ions. In the Tollens test, the formation of a
silver mirror on the inner surface of the test tube indicated the presence of reducing sugars. Tollen's
reagent reacts with reducing sugars or aldehydes to produce free silver metal. The silver ions in Tollen's
reagent are reduced to metallic silver, resulting in the formation of a silver mirror. The Picric Acid test
also confirmed the presence of reducing sugars. When reducing sugars react with picric acid, a red-
colored picramic acid is formed. The appearance of a red color in the test tube indicated a positive result
for the presence of reducing sugars.Lastly, in the Methylene Blue test, the blue color of methylene blue
disappeared when reducing sugars were present in the sample due to their reduction. However, when
reducing sugars were exhausted, shaking the tube allowed the reduced form of methylene blue to be
oxidized by atmospheric oxygen, resulting in the reappearance of the blue color. Overall, all four tests
provided positive results for the presence of reducing sugars in the samples tested. These tests are
valuable tools for qualitative analysis of carbohydrates, allowing for the identification of compounds
with free ketone or aldehyde functional groups.

CONCLUSION

The experiments employing the Benedict, Tollens, Picric Acid, and Methylene Blue
tests effectively identified the presence of reducing sugars in the samples. Distinct reactions
such as precipitate formation or color changes confirmed the presence of compounds with
free ketone or aldehyde groups. These qualitative methods provide valuable insights into
sample composition, demonstrating their utility in carbohydrate analysis and beyond.