BIO270 / OKT 2021 -FEB 2022

UNIVERSITI TEKNOLOGI MARA (UiTM)

BIO 270
ANIMAL & PLANT PHYSIOLOGY
Experiment II

DETERMINATION OF STARCH AND SUGAR IN SEEDS AND

HOW THOSE STARCHES BEEN DIGESTED IN HUMAN
GROUP NAME: A4AS1203C

LECTURER: DR MIMI SOPHIA SARBANDI

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We declare that the lab report submitted is not plagiarised and is entirely our own work,
and that no part of it has been copied from any work produced by other person(s)/
source(s) or provided by any other student(s). We understand that issuing a false
declaration can result in severe penalties and we are willing to be penalized if any form of
copying found valid.

No. FULL NAME OF TEAM MEMBER & ID TASK ASSIGNED SIGNATURE

NURUL FATINI BINTI NOR HISHAM LEADER IN
(2021126981) EXPERIMENT,
CONDUCT THE
EXPERIMENT,
RESEARCH,
INSERTING AND
LABELING PICTURES.

NURUL FATHIAH BINTI NOR HISHAM CONDUCT THE

(2021126967) EXPERIMENT,
COMPILE, RECHECK
LAB REPORT,
RESEARCH.

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GRADING

FOR LECTURER USE ONLY

Section Marks Total

INTRODUCTION
Introduction / Background 0.4 0.8 1.2 1.6 2.0

Problem Statement 0.1 0.2 0.3 0.4 0.5

Objectives 0.2 0.4 0.6 0.8 1.0

Hypothesis 0.1 0.2 0.3 0.4 0.5

MATERIALS & METHOD
Word structure 0.2 0.4 0.6 0.8 1.0

Sequence 0.2 0.4 0.6 0.8 1.0

RESULTS
Data Analysis (Figure & Table) 0.4 0.8 1.2 1.6 2.0

Description 0.4 0.8 1.2 1.6 2.0

DISCUSSION
Explanation (including questions) 0.4 1.2 2.8 4.4 6.0

Citation 0.2 0.4 0.6 0.8 1.0

Sources of error 0.2 0.4 0.6 0.8 1.0

CONCLUSION 0.2 0.4 0.6 0.8 1.0

REFERENCES 0.2 0.4 0.6 0.8 1.0

TOTAL /20

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1.0 Introduction

Starch is a crucial source of carbohydrates in the human diet because it is a complex

carbohydrate, (Bertoft, 2017). It must be digested for the body's absorption process to be
facilitated. Starch digestion is the process of breaking down starch. Sugar molecules are
broken into smaller sugar particles. The start of the process will be defined by chewing by the
mouth. The process of tearing down the long chain of starch that was held together by multiple
bonds. The mouth's salivary glands will be activated, and salivary amylase will break down
the food. Hydrolysis is a process that breaks down starch into simpler carbohydrates.

Sugar that contains aldehyde and ketone groups is recognized as reducing sugar, (Kunz,
2011). After that, this group will operate as a reducing agent. Benedict's test is done to
evaluate the presence of reducing sugar in a sample. Benedict's reagent is heated with a
sample that contained reducing sugars. The color of the solution will change from blue to
green, yellow, orange, red, and dark red. Based on the quantity of reducing sugar in the
sample, it will turn red or brown.

The iodine test is used to determine the presence of starch and to differentiate
monosaccharides and disaccharides from polysaccharides, which are composed of a long
chain of sugar subunits that are bound together, (Niaz, 2020). The presence of starch can be
seen when the color changes. When iodine is applied, the color changes from yellow-brownish
to bluish-black. The example when starch is hydrolyzed into a more basic form of
carbohydrates, there is no color change. This indicates that there is no longer any presence
of starch in the sample.

1.1 OBJECTIVES
1. To determine whether seeds contain starch or reducing sugars.
2. To see if saliva contains an enzyme that converts starch to sugar.
3. To discover more about the enzyme's properties and the conditions in which it works
best.

1.2 PROBLEM STATEMENT

To study whether there is any starch or sugar reduction chemical presence in seeds.

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1.3 HYPOTHESIS
1. Starch and reducing sugar are found in seeds.
2. Enzyme contained in the saliva breaks down starch to sugar.

2.1 MATERIALS/ APPARATUS:

• Balancing scale
• Measuring cylinder
• Test tubes
• Test tube rack
• Water bath
• Razor blade
• Benedict’s reagent
• Iodine solution
• Hydrochloric acid (1 mol dm-3)
• Forceps
• Peanut (ground) and Peanut (soaked)
• Mung bean (ground) and Mung bean (soaked)
• Corn grain (ground) and Corn grain (soaked)
• Starch suspension,
• Dextrose (glucose).

2.2 METHODS/ PROCEDURES:

A. Food reserves test

1. The whole of the cotyledon in iodine solution was stained.

2. Result was recorded in table 2.
3. Blender was used to prepare the sample for reducing sugar test.
4. The test tube was filled with 1 gram sample.
5. 4 ml of benedict’s reagent was added into test tube and the initial colour was recorded.
6. The test tube was placed in a boiling water bath for 3 minutes.

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7. The test tube from hot water bath was removed with the test tube holder and placed in
test tube rack. Colour changes was recorded in table 2.
8. The contents of the test tube were discarded and rinsed properly. Excess water from
the test tube was shacked out.
9. The same procedure was carried for other samples.

B. Effect of amylase on a starchy substance

1. 10 ml of uncontaminated saliva was collected in a measuring cylinder.

2. 2ml of the saliva was transferred to a test tube and placed in a boiling water bath for
15 minutes.
3. A further 2ml of saliva was transferred to another test tube and added with 2 drops of
hydrochloric acid (HCl). It was mixed well and leave for at least 15 minutes. The rest
of the other untreated saliva was kept at room temperature.
4. 6 pairs of test tube were set up and each test tube was labelled with code number as
in table 1.

Table 1: test tube setup

LABEL CONTENT
Pair A 1 2 ml sugar only
2 2 ml sugar only
Pair B 3 2 ml starch only
4 2 ml starch only
Pair C 5 2 ml untreated saliva only
6 2 ml untreated saliva only
Pair D 7 2 ml starch + 1 ml untreated saliva
8 2 ml starch + 1 ml untreated saliva
Pair E 9 2 ml starch + 1 ml preheated saliva
10 2 ml starch + 1 ml preheated saliva
Pair F 11 2 ml starch + 1 ml acidified saliva
12 2. ml starch + 1 ml acidified saliva

5. The test tubes were leave at least 10 minutes before the experiment proceeded.

1. The contents of each pair of tube were tested:

a. For starch, 2 drops of iodine solutions were added.
b. For sugar, 3ml of benedict’s reagent was heated.

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2. The starch test was conducted on the first of each pair of test tube, and the sugar test
on the second of each pair.
3. The test tube in the rack was arranged in numerical order to facilitate comparison.
4. The colour changes were recorded in a table 3. Test tubes were indicated on which
one showed positive results and which ones showed negative results for each starch
and sugar contents.

3.0 RESULTS

Table 2: Analysis of Seed Characteristics

Food Reserve Test (present or not present)

Sample
Starch Reducing Sugar

Peanut Not present Not present

Mung bean Not present Not present

Corn Present Present

Table 3: Enzyme Reactions for Sugar and Starch

TEST CONTENT OBSERVATI CONCLUSION

TUBE ON
LABEL
Pair A 1 2 ml sugar only Cloudy white Iodine test result:
colour negative
remains
unchanged Starch is absent
2 2 ml sugar only Colour Benedict’s reagent
change from result: positive
blue to brick
red High amounts of
reducing sugar present
Pair B 3 2 ml starch only Colour Iodine test result:
change from positive

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cloudy white
to bluish black Starch is present
4 2 ml starch only Colour blue Benedict reagent result:
remain negative
unchanged
Reducing sugar absent
Pair C 5 2 ml untreated saliva only Colour Iodine test result:
change from negative
cloudy white
to light brown Starch is absent

6 2 ml untreated saliva only Colour blue Benedict’s reagent

remain result: negative
unchanged
Reducing sugar are
absent
Pair D 7 2 ml starch + 1 ml untreated Colour Iodine test result:
saliva change from negative
cloudy white
to light brown Starch is absent

8 2 ml starch + 1 ml untreated Colour Benedict’s reagent

saliva change from result: positive
blue to
yellowish There is low amount of
reducing sugar present
Pair E 9 2 ml starch + 1 ml pre-heated Colour Iodine test result:
saliva change from positive
cloudy white
to bluish black Starch is present
10 2 ml starch + 1 ml pre-heated Colour blue Benedict’s reagent
saliva remain result: negative
unchanged
Reducing sugar are
absent
Pair F 11 2 ml starch + 1 ml acidified Colour Iodine test result:
saliva change from positive
cloudy white
to bluish black Starch is present

12 2 ml starch + 1 ml acidified Colour blue Benedict’s reagent

saliva remain result: negative
unchanged
Reducing sugar are
absent

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Food reserves test using iodine solution

Reducing sugar test before being put bath water

Reducing sugar test after being put in bath water

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Starch test with iodine reagent

Sugar test with benedict reagent

4.0 DISCUSSION

After the experiment is done and conducted, we can summarize all the results that we gained
from the experiment. There are a few tests that we run to determine whether seeds contain
starch or reducing sugars, to see if saliva contains an enzyme that converts starch to sugar,
and to discover more about the enzyme's properties and the conditions in which it works best.

The food reserve test was run to determine whether seeds contain starch or reducing sugars
in dry and soaked corn, peanut, and mug bean. Benedict’s reagent and iodine solution were
used in the first experiment as an indicator of the presence of reducing sugars and starch. The
iodine solution color changed from yellow-brown to blue-black after was stained with iodine

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solution and left for a while. This indicates that starch is present in corn. Next, benedict’s tests
were conducted on a corn sample test tube and the result was the color of the solution
changed from light green to yellowish green after getting boiled in the water bath for 3 minutes
and benedict’s reagents were added to the test tube. Test tubes for peanuts and mug beans
show no reaction and no starch present. From this experiment, we conclude that corn’s
cotyledon stored or contains a high level of starch because it changed from yellow-brown to
intense blue-black an iodine test.

For the second experiment, for test tubes 1 and 2, only sugar was added in pair A meanwhile
test tubes 3 and 4 in pair B, only starch was added. In pair C, test tubes 5 and 6, only 2 ml of
untreated saliva were added. Test tubes 7 and 8 in pair D, 2ml starch, and 1 ml untreated
saliva were added. Test tubes 9 and 10 in pair E, 2ml starch, and 1 ml preheated saliva was
added. Test tubes 11 and 12 in pair F, 2ml starch, and 1 ml acidified saliva were added. These
test tubes need to leave undisturbed for 10 minutes before the observation to give some time
for the reaction of the saliva and any breaking down starch actions that exist in the seed of
reducing sugar. In test tubes 5 and 6, the presence of starch and sugar was tested. In test
tubes 7 and 8, the test was conducted to test the hypothesis that saliva can break down the
starch to sugar due to an enzyme in it and due to active enzymes in untreated saliva. From
the observation, in test tube 7, to indicate the presence of starch in the solution was tested
with an iodine test. The color in the test tube changed from cloudy white to light brown due to
no starch being present in it after a few drops of iodine were added to the test tube. Meanwhile,
the benedict test was conducted in test tube 8. The color of the solution changes from blue to
yellowish due to the presence of sugar in the solution after the benedict reagent was added
to starch with untreated saliva and was left undisturbed. We conclude that the amylase
enzyme from salivary glands breaks down starch into sugar and break starch into the smaller
molecule which is maltose which consists of 2 glucose molecules that have been split by
maltase.

In test tubes, 7, 8, 9, 10, 11, and 12 were to discover more about the enzyme's properties and
the conditions in which it works best. Iodine tests were run in test tube 9 which contains 1 ml
of preheated saliva was added to starch from this experiment, the color changed from cloudy
white to bluish-black due to the presence of starch in it because when an enzyme in the saliva
has been denatured due to high temperature, (Viitolo, 2020). Next, the iodine test was run in
the test tube 11 which contains 1 ml of acidified saliva and was added to the starch. The color
changed from cloudy white to bluish-black due to the presence of starch and low pH that
denatures the enzyme. Based on the datasheet and experiment, the hypothesis is proven
correct that the amylase enzyme in the saliva is capable to break down starch into sugar as

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proof color in test tube 8 changed from blue to yellowish-green due to sugar that contains
starch.

Lastly, we need to have a lot of food or sample varieties so that we can have a lot of variant
results that we can obtain and do at least 3 retests on each experiment or test so that we can
lower the percentage of error in the experiment or data.

1. Does the endosperm or cotyledon contain more stored starch?

• The cotyledon contains more stored starch rather than the endosperm due to cotyledon
is some sort of the leaf of embryo and its main job is to provide nutrients needed for
sprouting plants meanwhile endosperm is non-living tissue and is also named as a
seed storage organ for food.

2. What is the purpose of food reserve test?

• The purpose of the food test is to determine or identify if seeds contain reducing sugar
or starch. Reserve food material contains a reserve of carbs, lipids, or even proteins
in tissue or cells that can work as energy stores, used or released for synthesis (ATP)
when needed. Besides, the close reserve food material to humans is fungi.

3. Why must the contents of test tubes be left undisturbed for at least 10 minutes before
starting the analysis?

• The test tubes were left undisturbed for at least 10 minutes before the analysis to make
sure that the molecule of starch goes through hydrolysis first and next it (starch) will
get catabolized and converted with the help of enzyme to become sugars.

4. What is the purpose of testing test tubes 1 to 6?

• The purpose is as a control group to make sure the experiment goes well.

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5. Based on your experiment, which of the test tube are used to test the hypothesis that
saliva contains an enzyme which breaks down starch to sugar?

• Test tube pair D because it incorporates starch and saliva solution.

6. What is the name of the enzyme in saliva and what compound does it produce from
starch?

• The name of the enzyme is amylase that produces maltose.

7. Did the hypothesis turn out to be correct? Would you say it was proven?

• Yes, the hypothesis is correct. It can be proven when combining untreated saliva and
starch, it shows no reaction or the iodine colour which is orange remain the same.

8. Which test tube provides information on the properties of the enzyme?

• Test tube that provides information on the properties of enzyme are test tubes 9 and
10 which contains preheated saliva, and test tubes 11 and 12 which contains acidified
saliva due to pH level and temperature as a factor that affect the properties and activity
of the enzyme once they got introduced into the process of the reaction. This is also
because high temperature can alter or change the shape or denaturing the enzyme
meanwhile optimum pH level can make the enzyme works well and both of these can
affect their properties.

5.0 CONCLUSION

All the Hypotheses and objectives in this experiment were accepted. Reducing sugars and
starch presence was identified in seeds which are mug bean, peanut, and corn. The result in
corn grain shows that starch presence after getting tested in an iodine solution, the color
changed from cloudy white to bluish-black meanwhile mug bean and peanut do not have any

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reaction due to absence of starch. Mug beans and peanuts contain reducing sugar, and this
was shown when the color changed from blue to bluish-black after has been removed from
the water bath in the reducing sugar test. Meanwhile, there was no presence of reducing sugar
in corn grain due to the blue color remained unchanged after being removed from the water
bath. Different types of seeds also can cause varieties resulting in the presence of reducing
sugar and starch.

Next, it is having been proven that the enzyme amylase in saliva is capable to break down to
sugar in the inspection of saliva contains an enzyme that converts starch to sugar. In test tube
9 in pair E and test tube 11 in pair F, the color changed from cloudy white to bluish-black as
in the experiment.

Lastly, the enzyme's properties and the conditions in which it works best were discovered are
enzyme works efficiently at 37 degrees Celsius (body temperature) and good in non-acidic
solutions. The amylase in preheated saliva was denatured due to high temperature and the
amylase denatured due to low pH (acidic solution) in the test tube that contains hydrochloric
acid. When the temperature of the enzyme and pH level increase, the rate of enzyme work
increases as in pair E and F. Enzyme functions well when they are in their optimal pH and
temperature.

REFERENCES

Bertoft, E. (2017). Understanding Starch Structure: Recent Progress. Agronomy, 7, 56.

Kunz, T. a.-J. (2011, July). Glucose - a Reducing Sugar? Reducing Properties of sugar in
Beverages and Food. Brewing Science, 64.

Niaz, K. a. (2020). Analysis of carbohydrates (monosaccharides, polysaccharides). 621-633.

Viitolo, M. (2020). BRIEF REVIEW ON ENZYME ACTIVITY. World journal of pharmaceutical

research , 9, 60.

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