QUALITATIVE ANALYSIS FOR CARBOHYDRATES

Advinn Nathaniell S. Celendro

BSN 1-YA-21
March 5, 2022

I. INTRODUCTION
In order for us to identify different types of Carbohydrates. Solubility, and presence of
Starch in the solution, we use different types of testing such as Solubility test, Molisch’s test,
Fehling’s test, Benedict’s test, Tollen’s test, and Iodine test. The Solubility test is used to carried
out to determine a compounds ability to dissolve in a solvent. Next, Molisch test is used to check
for the presence of carbohydrates in a given analyte. Next, Fehling test is used to differentiate
and identify between reducing and non-reducing sugars. It also distinguishes the ketone
functional group. The Benedict test, it is used to detect the presence or absence of reducing sugar
in the given solution, while Tollen’s test is a silver-mirror test that used to distinguishes between
an aldehyde and ketone. Lastly, an Iodine test is a chemical reaction-based identification test for
starch.

II. MATERIALS
(1) Samples d) Fehling’s solution B
a) Glucose e) Benedict’s reagent
b) Sucrose f) Tollen’s reagent
c) Starch g) Iodine solution
(2) Reagents (3) Apparatus
a) Distilled water a) Test tubes
b) Molisch’s reagent b) Dropper
c) Fehling’s solution A c) Beaker

III. METHODS
 (1) Solubility test
a. Take a small amount of the given sample in a test tube and add 3 ml of distilled water
into it and shake the test tube well.
(2) Molisch test
a. Take about 2 ml of aqueous solution of the sample in a test tube and add few drops of
Molisch’s reagent into it. Pour 1 ml conc. H 2SO4 slowly along the side of the test
tube.
(3) Fehling’s test
a. Take 2 ml of aqueous solution of the sample in a test tube and add 1-2 ml each of
Fehling’s solution A and Fehling’s solution B into it. Keep the test tube in a boiling
water bath.
(4) Benedict’s test
a. Take 1-2 ml of aqueous solution of the sample in a test tube and add 1-2 ml of
Benedict’s reagent. Keep the test tube in a boiling water bath.
(5) Tollen’s test
a. Take 2-3 ml of aqueous solution of the sample and in a test, tube add 2-3 ml of
Tollen’s reagent. Keep the test tube in a boiling water bath for 10 minutes.
(6) Iodine test
a. Take a small amount of aqueous solution of the sample in a test tube and add 1-2
drops of iodine solution into it.

IV. DATA

Solubility test

SI No. Sample Observation

1. Glucose Soluble
2. Sucrose Soluble
3. Starch Insoluble

Molisch’ test

SI No. Sample Observation

1. Glucose A red violet (purple) ring is
formed at the junction of two
layers.
2. Sucrose A red violet (purple) ring is
formed at the junction of two
layers.
3. Starch A red violet (purple) ring is
formed at the junction of two
layers.

Fehling’s test

SI No. Sample Observation

1. Glucose A reddish precipitate is
formed
2. Sucrose No formation of reddish
precipitate
3. Starch No formation of reddish
precipitate

Benedict’s test

SI No. Sample Observation

1. Glucose A reddish precipitate is
formed
2. Sucrose No formation of reddish
precipitate
3. Starch No formation of reddish
precipitate

Tollen’s test

SI No. Sample Observation

1. Glucose A shinning silver mirror is
form
2. Sucrose No formation of silver mirror
3. Starch No formation of silver mirror
 Iodine test

SI No. Sample Observation

1. Glucose No characteristic change
2. Sucrose No characteristic change
3. Starch A blue color is formed

V. ANALYSIS

According to the result of the Solubility test, glucose and sucrose are soluble in

water, while starch is insoluble in water. From Fehling’s test and Benedict’s test, glucose has the

presence of reducing sugars because it formed a reddish precipitate, while sucrose and starch

have no presence of reducing sugars. From Tollen’s test, glucose has formed a silver mirror

which indicates that it has the presence of reducing sugar, while sucrose and starch have no

formation of silver mirror, which indicates no presence of reducing sugar. Lastly, in the Iodine

test, glucose and sucrose have no characteristic change, which means no presence of starch. The

starch solution has formed a blue color, which means that it has a presence of starch.

VI. CONCLUSION

In conclusion, students can identify and understand the different types of

carbohydrates/ sugar solution using Fehlings, Benedict’s, and Tollen’s test. Three test has

different types of reagent/ solution, and physical result to the test tubes, but the same outcome

which is used to determine the presence of non-reducing and reducing sugars. The Solubility test

used to determine the determine the solubility of the samples, while in Iodine test used to

determine the presence of starch in different kind of samples.

VII. REFERENCE
https://byjus.com/jee-questions/what-is-solubility-test/#:~:text=Solubility%20test%20is%20a
%20type,compounds%20are%20acidic%20or%20basic.

https://byjus.com/chemistry/molischs-test/

https://byjus.com/chemistry/molischs-test/

https://byjus.com/chemistry/benedicts-test/

QUALITATIVE TESTS FOR CARBOHYDRATES

Advinn Nathaniell S. Celendro
BSN 1-YA-21
March 5, 2022

I. INTRODUCTION
To understand and know the presence of carbohydrates, we have different types
to test and reagent to identify and determine the presence of carbohydrates in the sugar
solutions such as glucose, fructose, lactose, and sucrose. One of these is the Molisch test,
which is used to check the presence of carbohydrates by reaction with conc. H2SO4 gets
dehydrated to form furfural and its derivatives. Second, the Benedict test is used to check
the presence of carbohydrates by reducing its sugar. It used Benedict’s reagent that is
composed of a complex mixture of sodium carbonate, sodium citrate, and copper (II)
sulfate pentahydrate. Third, the Fehling test is used to differentiate reducing and non-
reducing sugar. It also differentiates ketones and aldehydes. Fourth, the Barfoed test is
used to detect the presence of reduced monosaccharide by oxidized by the copper ion in
solution to form a carboxylic acid and a reddish precipitate of copper (I) oxide within
three minutes. Fifth, the Seliwanoff test was used to differentiate between sugars that
have a ketone group (ketoses) and an aldehyde group (aldoses). Lastly, an Iodine test is
used to identify and determine the presence of starch.

II. MATERIALS
 (4) Test tubes (12) Iodine Solution
(5) Dropper
(13) Glucose
(6) Graduated cylinders
(14) Fructose
(7) Fehling’s reagents A and
(15) Lactose
B
(16) Sucrose
(8) Benedict’s reagent
(17) Starch
(9) Seliwanoff’s reagent
(18) Test tube racks
(10) Molisch’s reagent
(19) Marker and paper tape
(11) Barfoed’s reagent

III. METHODS

First, do the Molisch test by putting 2ml of each carbohydrate into the test tubes and
then putting 10 drops of Molisch reagent into each carbohydrates test tube. After putting the
Molisch reagent, carefully add 15 to 20 drops of concentrated sulfuric acid and do not stir.
Second, do Benedict’s test by adding 2ml of Benedict’s reagent/solution to the four empty tubes
and placing them in the water bath for 30 seconds. After heating all the four test tubes with
Benedict’s test, add five drops of different carbohydrates solutions and place them into a boiling
water bath. Third, do the Fehling’s test by adding and mixing the 1ml of Fehling’s solution A
and 1ml of Fehling’s solution B to four empty test tubes. And then, add 3ml of water to all the
test tubes and place it into a boiling water bath for 1 minute. When the solution of four tubes
remains blue, start adding 8 drops of sugar solution to every four tubes and place to the water
bath for 2 minutes. The third is the Barfoed’s test, add 2ml of Barfoed's reagent to every four test
tubes and then add 10 drops of sugar solution such as glucose, fructose, lactose, and sucrose in
each four test tubes. Then, place the four-test tube with sugar solution and barfoed’s reagent to
the boiling water bath for 5 minutes and cool it down for 15 minutes. Fourth, do the Seliwanff
test. Keep in mind that don’t overheat the samples because may it lead to giving positive results
to all the samples. Start with adding 2ml of each sugar solution to the four test tubes and add 3
ml of Seliwanff’s reagent to each test tube. Then, immerse the solutions in a boiling water bath
and observe the color changes during the first 10 minutes of boiling. Lastly, Iodine test for starch
by placing 2ml of the plant starch solution and then adding a drop of iodine solution. And then,
heat the solution and observe the change.
IV. DATA

Molisch Test

Test tube 1 2 3 4
Sugar solution 2ml of Glucose 2ml of 2ml of Lactose 2 ml of
Fructose Sucrose
Molisch 10 drops 10 10 drops 10 drops
reagent drops
Concentrated 10 – 15 drops 10 – 15 drops 10 – 15 drops 10 – 15 drops
Sulfuric Acid

Benedict Test
Test tube 1 2 3 4
Sugar solution 5 drops of 5 drops of 5 drops of 5 drops of
Glucose Fructose Lactose Sucrose
Benedict’s 2ml 2ml 2ml 2ml
solution
Color change Brick-red Brick-red Brick-red None

Fehling Test
Test tube 1 2 3 4
Sugar solution 8 drops of 8 drops of 8 drops of 8 drops of
Glucose Fructose Lactose Sucrose
Fehling’s 1ml 1ml 1ml 1ml
solution A
Fehling’s 1ml 1ml 1ml 1ml
solution B
Water 3ml 3ml 3ml 3ml
Result Small amount Brick-red Small amount Blue/none
Brick-ret Brick-ret

Barfoed Test
Test tube 1 2 3 4
Barfoed’s 2ml 2ml 2ml 2ml
solution
Sugar solution 10 drops of 10 drops of 10 drops of 10 drops of
Glucose Fructose Lactose Sucrose
Result Brick-red Brick-red None None
Seliwanoff Test
Test tube 1 2 3 4
Seliwanoff’s 3ml 3ml 3ml 3ml
reagent
Sugar solution 2ml of Glucose 2ml of Fructose 2ml of Lactose 2ml of Sucrose
Result Clear Red Clear Red

Iodine Test
Test tube 1
Starch 2ml
Iodine Solution 1 drop
Result Blue with dark blue in the bottom

V. ANALYSIS
In the Molisch test, when the test solution creates a purple or purplish-red tinted
layer, it results that the test solution has the presence of carbohydrates. In the Benedict test, when
a test solution change color from blue (no reducing sugar), green/yellow (traces of reducing
sugar), orange-red (moderate), and brick-red (A large amount of reducing sugar). In the Fehling
test, the appearance of a reddish-brown precipitate indicates reducing sugars, while the
appearance of deep blue color indicates a lack of reducing sugars. In the Barfoed test, the
formation of a brick red color in the bottom of the test solution is indicated to be a
monosaccharide. In the Seliwanoff test, the formation of the cherry red-colored complex
indicates that the test samples contain ketoses. In an Iodine test, starch is present when the
sample's color changes ranging from violet to black, while no starch when the yellow color of the
iodine solution.

VII. CONCLUSION

In conclusion, according to the Molisch test, all the sugar solution such as glucose,
fructose, lactose, and sucrose has a presence of carbohydrate. For Benedict’s test, glucose,
fructose, and lactose have a high amount of reducing sugar, while sucrose has no reducing sugar.
Next, according to Fehling’s test, glucose, fructose, and lactose have reduced sugar, while
sucrose has no reducing sugars. In Barfoed’s test, glucose and fructose are a monosaccharide,
while lactose and sucrose are a disaccharide. In the Seliwanoff test, fructose and sucrose are
ketoses, while glucose and lactose are aldoses. Lastly, In the Iodine test, starch is present when
the sample's color changes ranging from violet to black

VIII. REFERENCE

https://www.nku.edu/~whitsonma/Bio150LSite/Lab%203%20Organic/
Bio150LRevMolec.html#:~:text=Interpreting%20Benedict's%20Reagent%20Results&text=The
%20%22hotter%22%20the%20final%20color,is%20a%20very%20strong%20positive.

https://www.vedantu.com/chemistry/molisch-test

https://www.nku.edu/~whitsonma/Bio150LSite/Lab%203%20Organic/
Bio150LRevMolec.html#:~:text=Interpreting%20Benedict's%20Reagent%20Results&text=The
%20%22hotter%22%20the%20final%20color,is%20a%20very%20strong%20positive.

https://microbenotes.com/fehlings-test/#:~:text=the%20test%20tubes.-,Result%20and
%20Interpretation%20of%20Fehling's%20Test,and%20lack%20of%20reducing%20sugars.

https://www.onlinebiologynotes.com/barfoeds-test-objective-principle-reagents-procedure-
and-result/#:~:text=Positive%20Barfoed's%20test%3A%20development%20of,test%3A
%20absence%20of%20red%20color

https://microbenotes.com/seliwanoffs-test/#:~:text=The%20formation%20of%20the
%20cherry,sample%20doesn't%20have%20ketoses.

https://writeonline.ca/annotation.php?s=lr5#:~:text=A%20positive%20result%20for
%20the,colour%20of%20the%20iodine%20solution.

BENEDICT’S TEST FOR REDUCING SUGARS

Advinn Nathaniell S. Celendro
BSN 1-YA-21
March 5, 2022

I. INTRODUCTION
 Stanley Rossiter Benedict was born on March 17, 1884, and died on December 21, 1936.

He was an American chemist and best known for discovering Benedict’s reagent, a solution that

detects certain sugar. Benedict’s test is used to detect the presence or absence of reducing sugar

in the solution. Also, it uses to identify reducing sugars (monosaccharides and some

disaccharides) and simple carbohydrates. Benedict’s solution can be used to test for the presence

of glucose in urine. Some sugars, such as glucose, are known as reducing sugars because they

can transfer hydrogens (electrons) to other compounds, a process known as reduction. When

reducing sugars are heated in the presence of Benedict's reagent, a reduction reaction occurs,

causing Benedict's reagent to change color. Depending on the amount and type of sugar, the

color can range from green to dark red (brick) or rusty-brown. The mixture of Benedict’s

solution and simple carbohydrates such as glucose are heated, the solution changes t orange-red/

brick read due to the reducing property of simple carbohydrates. The copper (II) ions in

Benedict’s solution serves as a reagent that is composed to copper, sodium citrate, sodium

carbonate, and copper II sulphate pentahydrate. The reduced to Copper (I) ions, which causes

the color change.

In the video shown, the students ask to detect the presence of sugar in the given sample of

urine by Benedict’s reagent. Determine the color if it’s blue, green, orange-red, or brick-red, and

identify what symbolizes the change color of the solution.

II. MATERIALS

(1) Benedict’s solution

(2) Test tube

 (3) Test tube holder

(4) Pipette

(5) Urine Sample

(6) Burner

III. METHODS

In the first experiment with the urine and Benedict’s reagent. The first step is using a pipette

to accurately take 5ml of Benedict’s reagent and perfectly transfer it to the test tube. Second, take

5ml of freshly collected urine by pipette and add it to the test tube with Benedict’s reagent.

Third, using test tube holder safety get the test tube to heat it on the burner for about 2 minutes.

Fourth, after 2 minutes observe the changes of the mixture. The second experiment is using

glucose and Benedict’s reagent. First, using pipette again put 5ml of Benedict’s reagent to the

test tube. Second, put a 15ml of glucose by using the pipette to the test tube with Benedict’s

solution. Lastly, as the same in experiment 1, using test tube holder heat it through burner for 2

minutes and observe the changes.

IV. DATA

Test tube # #1 #2

Mixture 5ml of fresh 15 ml of

urine glucose

Benedict’s 5ml 5ml

reagent

Time 2 minutes 2 minutes

Color Green Brick Red

 Change

IV. ANALYSIS

Color Change Reduces sugars

Blue None

Green Traces

Orange-red Moderate amount

Brick-red Large amount

In the first experiment, the mixture of urine and Benedict’s solution change into a color of green

after being heated on the burner for 2 minutes. The Green indicates that the first samples have

traces or a small amount of reducing sugar or sugars. The second sample is composed of 15ml of

glucose. Glucose is a type of simple carbohydrate. The amount of sugar in the sample is high that

it changes into brick-red color. The brick-red color indicates that the sample has a high amount

of sugar or a large amount of sugar was reduced through the process of the test.

V. CONCLUSION

In conclusion, Benedict’s test is to detect the presence of simple carbohydrates in an

unidentified analyte such as urine. When the sample changes its color to blue, it has no sugars.

The green has traces/ small amount of reduced sugar. The orange-red has moderate sugar, and

bright-red contains a large high amount of sugar in the sample. A urine glucose test can be used

to help determine if blood glucose levels are too high, which may be a sign of diabetes. We can
use Benedict’s solution to detects diabetes mellitus because it used to check the reduce sugars in

the reaction.

VI. REFERRENCES

https://en.wikipedia.org/wiki/Stanley_Rossiter_Benedict
https://www.webmd.com/diabetes/glucose-diabetes
https://byjus.com/chemistry/benedicts-test/
https://europepmc.org/articles/pmc1503294/pdf/califmed00020-0002.pdf
https://www.youtube.com/watch?v=PAKCgrnKeBA

BARFOED’S TEST
Advinn Nathaniell S. Celendro
BSN 1-YA-21
March 5, 2022

I. INTRODUCTION
The Barfoed’s test was invented by Danish chemist named Christen Thomsen Barfoed.

Barfoed’s test use a Barfoed reagent which is made up of copper acetate in a dilute solution of

acetic acid. The appearance of precipitate thus helps distinguish reducing monosaccharides from

reducing disaccharides.

II. MATERIALS

(1) Maltose

(2) Barfoed’s reagent

(3) Glucose

(4) Test tubes

 (5) Measuring cylinder

(6) Pipette

III. METHODS

First, measure 3ml of freshly prepared Barfoed’s reagent into labelled test tubes. Second,

add 1ml of the test solution to Barfoed’s reagent that result to Maltose + Barfoed’s reagent and

Glucose + Barfoed’s reagent. Third, the test tubes into a boiling water bath and heat for 3

minutes.

IV. DATA

Samples 1 2

Carbohydrates Maltose Glucose

Reagent Barfoed’s Barfoed’s

Color None Brick red

formation
V. ANALYSIS

The first sample contains maltose and Barfoed’s reagent turn in to color blue, while second

sample contains glucose and Barfoed’s reagent forms a red or yellow orange below precipitate

below the test tube.

VII. CONCLUSION

In conclusion, the formation of red or yellow to orange precipitate is a positive test for

reducing monosaccharides. In result, maltose is disaccharides, while glucose is monosaccharides.

VIII. REFERRENCES

https://www.youtube.com/watch?v=yQfMqvOxPrc

https://microbenotes.com/barfoeds-test/

https://en.wikipedia.org/wiki/Barfoed%27s_test

TOLLEN’S TEST
Advinn Nathaniell S. Celendro
BSN 1-YA-21
March 5, 2022

I. INTRODUCTION
 The Tollens’s test is made by Bernhard Christian Gottfried Tollens (1841 – 1918). He was

a German chemist and the one who was recognized through the silver mirror test using Tollens’s

reagent. He also developed to differentiate between aldose and ketose sugars. Tollens’s test is a

chemical test used also to distinguish reducing sugars from non-reducing sugars. This test is

known as the silver mirror test since the free silver meal is formed at the of the test reaction. It

used an alkaline solution of silver nitrate (AgNO3) mixed with liquid ammonia (NH3). The

Tollens’s reagent is a strong oxidizing agent that oxides the aldehyde group present in some

carbohydrates to form a carboxylic acid. Silver ions in the presence of hydroxide ions come out

of the solution as a brown precipitate of silver(I) oxide, Ag2O(s). This precipitate dissolves in

aqueous ammonia, forming the diamine silver(I) ion, [Ag (NH3)2] +. Ketones do not react with

Tollens' reagent.

In this study, the student aims to detect the presence of reducing sugars in given sample

by Tollens’s test.

II. MATERIALS

(1) Tollens’s reagent (Ammoniacal solution of silver nitrates)

(2) Test tubes

(3) Pipette

(4) Water bath

III. METHODS

The first step to this experiment is to create Tollens’s reagent/ solution. Step 1, take 1 ml

of aqueous silver nitrate solution in a test tube. Step 2, add sodium hydroxide solution drop wise

to get the precipitate of silver oxide. Last step, now ass 2 drops ammonium hydroxide solution
while shaking the mixture and formed precipitate dissolve. After creating Tollens’s reagent, in

different test tube take a 2 ml of sugar solution. Then, take 2 ml of Tollens’s reagent to it and

observe the changes.

IV. DATA

Sample # 1

Mixture 2ml of Tollens’s

reagent

Solution 2ml of sugar

solution

V. ANALYSIS

The sample with 2ml of Tollens’s reagent and 2ml of sugar solution forms a silver mirror on

the wall of the test tube that indicates the presence of reducing sugars.

VI. CONCLUSION

In conclusion, Tollens’s test can be used to identify and distinguishes aldehydes from

ketones. It also used to help and differentiate of reducing sugars from non-reducing sugars. From

the result given, the student able to understand that aldehydes are readily oxidized, whereas

ketones are not. Lastly, using Tollens’s test we can identify and detect the presence of aldehyde

containing carbohydrates like glucose. A sample contains reducing sugars or aldoses when there

is a formation of dark grey precipitate or silver mirror on the bottom and sides of the test tube.

V11. REFERENCE
https://edu.rsc.org/exhibition-chemistry/the-silver-mirror-test/2020077.article
https://onlinesciencenotes.com/tollens-test-or-silver-mirror-test-principle-requirements-
procedure-result-interpretation-and-limitations/
https://www.youtube.com/watch?v=8HjtN5vbLj8
https://byjus.com/chemistry/tollens-test/#:~:text=What%20color%20is%20a%20positive,sample
%20contains%20reducing%20sugars%2F%20aldoses.