GROUP 2

Leader: Bucatcat, Casandra Jnesen

Members:
Bordo, Shiela Mae
Buzon, Ethand Erique
Bendebel, Jenny D.
Beltrano, Marydin F.

1.1 SPECIFIC REACTION TESTS FOR CARBOHYDRATES

1.1.1 BASED ON THE FURFURAL FORMATION AND ITS DERIVATIVES

1.1.1.1 SELIWANOFF’S TEST
Maerial
a. Seliwanoff’s reagent
b. 0.1M Sugar solutions: Glucose , Xylose , Fructose
Procedure:
1. In separate test tubes, add 1mL of each of the sugar solutions to 5mL of the seliwanoff’s reagent.
2. Immerse in a boiling bath.
3. Observe the color changes in each tube during the first 15 minutes of boiling.
4. Record the color and the time of formation for each solution tested.

Principle:
The Seliwanoff's test is a method used to differentiate between ketose and aldose sugars according to the
Biology Notes Online. This test involves the use of hydrochloric acid as a dehydrating agent and resorcinol as
a condensing agent. Ketoses undergo dehydration more rapidly in the presence of HCl, forming a furfural
derivative (5-hydroxymethylfurfural) that reacts with resorcinol to produce a cherry-red colored compound.
On the other hand, aldoses also yield the same product but at a slower rate, leading to the formation of a
pale pink to yellow colored complex.

Result:
Based on the result conducted during experiment, there are two clesr result which is the Glucose
and Xylose, which represent as a negative result. While the Frtuctose result was a light orange or a
cherry color which means ketosugar (Fructose along with Sucrose) exists in the solution.

Reference/s:
Biology Notes Online. (n.d.). Principle of Seliwanoff's Test. Biology Notes
Online. https://biologynotesonline.com/seliwanoffs-test-principle-procedure-result/

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 1.1.1.2 BIAL’S ORCINOL TEST
Material
a. Orcinol reagent
b. 0.1M Sugar solutions: Glucose, Xylose, Fructose
Procedure:
1. Pipette 0.5mL of each of the sugar solution into separate test tube.
2. Add 4.5mL of the orcinol reagent into each test tube.
3. Place the tubes in a boiling water bath.
4. Note the color formed with each reactant during 15 minutes of heating.

Principle:
This test is based on the principle that under hydrolysis pentosans are hydrolyzed into pentoses.
Further, pentoses are dehydrated to yield furfural, which in turn condense with orcinol to form a
blue-green precipitate. In the presence of hexoses, hydroxyfurfural is formed instead of furfural
which upon condensation with orcinol forms a muddy brown colored precipitate. The intensity of
the precipitation is directly proportional to the concentration of the pentoses in the sample. The
intensity of the color developed depends on the concentration of HCl, ferric chloride, orcinol, and
the duration of boiling. The concentration of the sugars is determined by measuring the absorbance
of 620 nm wavelength in a spectrophotometer or in a red filter colorimeter.

Result:
Therefore, the Bial’s Test result was clearly distinguish pentoser sugar Xylose which changes from
green to Blue-Green, while Fructose and the Glucose showed a dark green color transformation.
This means the Dark Blue-Green represent as a positive result of the experiment while the two
Dark Green result is negative experiment result. The Xylose positive ruselt also represent the
indicates the presence of pentoses in the sampleic.ates the presence of pentoses in the sampl

Reference/s:
Sapkota, A. (2022, April 18). Bial’s Test- Definition, Principle, Procedure, Result, Uses. Microbe Notes.
https://microbenotes.com/bials-test/

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 1.1.1.3 FEHLING’S TEST
Material
a. Fehling’s reagent (A and B)
b. 0.1M Sugar solutions: Glucose , Fructose, Galactose, Lactose, Maltose, Sucrose

Procedure:
1. Mix 1mL of the Fehling’s A and 1mL of the Fehling’s B. Dilute with 4mL of water.
2. Boil the solution for 1 minute. The solution should be clear blue.
3. Place 8 drops of sugar solution in a separate test tubes.
4. Add 1mL of the mixed Fehling’s solution to each of the tubes.
5. Boil the tubes in a water bath for 5minutes and note the changes produced.

Principle:
The carbohydrates having free or potentially free carbonyl groups (aldehyde or ketone) can act as
reducing sugars.The Fehling’s solution appears deep blue in color and consists of copper sulfate
mixed with potassium sodium tartrate and strong alkali, which is usually sodium hydroxide.

Result:
Based on our experiment for Glucose, Fructose, Galactose, Maltose, Sucrose and Lactose was
resulted a clear to deep blue color, means that the result was negative which is the absence of the
reddish precipitate or the appearance of deep blue color indicates a negative result and lack of
reducing sugars.

Reference/s :
Sapkota, A. (2022, April 18). Fehling’s Test- Definition, Principle, Procedure, Result, Uses. Microbe Notes.
https://microbenotes.com/fehlings-test/

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 1.1.1.4 BARFOED’S TEST
Material
a. Barfoed’s reagent
b. 0.1M Sugar solutions: Glucose , Fructose, Galactose, Lactose, Maltose, Sucrose

Procedure:
1. To 3mL of Barfoed’s reagent add 10 drops of the sugar solution.
2. Place in a boiling water bath for 15 minutes.
3. Remove and allow to cool spontaneously.
4. After 15 minutes observe and record the results.

Principle:
The Barfoed reagent is made up of copper acetate in a dilute solution of acetic acid. Since acidic pH is
unfavorable for reduction, monosaccharides, which are strong reducing agents, react in about 1-2
min. However, the reducing disaccharides take a longer time of about 7-8 minutes, having first to get
hydrolyzed in the acidic solution and then react with the reagent. Once the reaction takes place, thin
red precipitate forms at the bottom of the sides of the tube. The difference in the time of appearance
of precipitate thus helps distinguish reducing monosaccharides from reducing disaccharides.

Result:
The result of Glucose, Fructose, Maltose is Darkish Blue-Red which is the positive result of
Barfoed’s Test result, while the Maltose, Sucrose and Lactose was resulted only a Darkish Blue and
this represent a negative result of our experiment. Therefore, the positive result that has a Red in
the last part or the below of Darkish Blue color in the tube is represent as a detects the presence
of reducing monosaccharides in the sample.

Reference/s:
Sapkota, A. (2022, April 18). Barfoed’s Test- Definition, Principle, Procedure, Result, Uses. Microbe Notes.
https://microbenotes.com/barfoeds-test/

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 1.1.1.5 BENEDICT’S TEST
Material
a. Benedict’s reagent
b. 0.1M Sugar solutions: Glucose , Fructose, Galactose, Lactose, Maltose, Sucrose
Procedure:
1. To 2mL of the benedict’s solution, add 5 drops of the sugar solution.
2. Place in a boiling water bath for 2 minutes
3. Cool. Record the results.

Principle:
When a reducing sugar is subjected to heat in the presence of an alkali, it gets converted into an enediol (which is a
relatively powerful reducing agent). Therefore, when reducing sugars are present in the analyte, the cupric ions
(Cu2+) inBenedict’s reagent are reduced to cuprous ions (Cu+). These cuprous ions form copper(I) oxide with
the reaction mixture and precipitate out as a brick-red coloured compound.

Result:
The result of our experiment was fice (5) positive while one (1) negative. The Glocuse, Fructose, Galactose,
Maltose and the Lactose was Brown-White, which represent the positive result while the Sucrose stay clear
blue.

Reference/s:
BYJU's. (2022). Benedict’s Test - Reagent Preparation, Principle, Procedure, Reaction. BYJU’S.
https://byjus.com/chemistry/benedicts-test/

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 Name: BUCATCAT, CASANDRA JENSEN A. Group No: #02
Schedule: THURSDAY 402 4:00PM-7:00PM Date: 08-14-24

ACTIVITY NO. 1.4

SPECIFIC TESTS FOR CARBOHYDRATES: Tabulation of results
SELIWANOFF’S TEST BIAL’S ORCINOL TEST
SUGAR SOLUTION
T.R A.R T.R A.R
Glucose (-) CLEAR CLEAR (-) CARK GREEN GREEN
Xylose (-) CLEAR CLEAR GREEN
(+) BLUE-GREEN
Fructose (+) LIGHT ORANGE CLEAR (-) DARK GREEN GREEN

Interpretation and conclusion:

Both Seliwanoff's test and Bial's test provide valuable insights into the chemical properties of sugars. They demonstrate the
importance of understanding the specific reactions of different sugar types and the usefulness of these reactions for
identification purposes.

SUGAR FEHLING’S TEST BENEDICTS TEST BARFOED’S TEST

SOLUTION
T.R A.R T.R A.R T.R A.R
Glucose (+) CLEAR SKY BLUE (+) BROWN- CLEAR (+) DARKISH BLUE
WHITE BLUE BLUR-RED
Fructose (+) CLEAR SKY BLUE (+) BROWN- CLEAR (+) DARKISH BLUE
WHITE BLUE BLUR-RED
Galactose (+) CLEAR SKY BLUE (+) BROWN- CLEAR (+) DARKISH BLUE
WHITE BLUE BLUR-RED
Maltose (+) CLEAR SKY BLUE (+) BROWN- CLEAR (+) DARKISH BLUE
WHITE BLUE BLUR-RED
Sucrose (+) CLEAR SKY BLUE (+) BROWN- CLEAR (+) DARKISH BLUE
WHITE BLUE BLUR-RED
Lactose (+) CLEAR SKY BLUE (+) BROWN- CLEAR (+) DARKISH BLUE
WHITE BLUE BLUR-RED

Interpretation and conclusion:

Fehling's, Benedict's, and Barfoed's Tests Interpretation and Conclusion The Fehling's, Benedict's, and Barfoed's
tests are essential tools for identifying and distinguishing between various types of sugars based on their
reducing properties. In the Fehling's test, the clear to deep blue color change observed for glucose, fructose,
galactose, maltose, sucrose, and lactose indicated the absence of reducing sugars, highlighting a negative result.
Similarly, the Benedict's test resulted in positive outcomes for glucose, fructose, galactose, maltose, and lactose,
with a brown-white color change, while sucrose exhibited a clear blue color, further confirming the presence of
reducing sugars. Additionally, the Barfoed's test successfully differentiated between reducing monosaccharides
and disaccharides, with glucose, fructose, and maltose showing a darkish blue-red color as a positive result, and
sucrose and lactose displaying a darkish blue color, signaling a negative outcome. These tests collectively
demonstrate their efficacy in identifying specific sugar types based on their distinct chemical reactions, providing
valuable insights into the composition and characteristics of various sugars.

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DOCUMENTATION

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