SUBMITTED

BY,

MERIN MARIAM
JOSE
Class:-XII A
Roll no: -36

It is said that a thankful heart is not only the greatest

virtue but also the parent of all other virtues. My efforts bore
fruit with the successful completion of this project. However
there are many
others who share the reward of this effort
simply because it would never have been this good without this
help.
First and foremost, I thank God almighty for been able
to complete this project with success. I acknowledge the
cooperation, encouragement, austerity of my chemistry teacher
Mrs.Bindhu.P
whose guidance did half the magic of keeping me
thrilled
throughout this project. I extend my sincere gratitude
to my teammate, who worked on this project with me, friends,
classmates, batch mates and also the lab assistants who have
helped me whenever I got stuck and for being patient with me
even at the toughest hours. Each of you played a very crucial role
in the development of this project.
A special thanks to my parents and family for their
support and understanding and of course for bearing with me
when I was all too busy with the project.

AIM-------------------6
INTRODUCTION-----7
THEORY-------------8,9
REQUIREMENTS-----10
PROCEDURE-1-------11
PROCEDURE-2-------12

PROCEDURE-3-------13
OBSERVATIONS-----14,15
CONCLUSI ON-------16
REFERENCES------17

To study the digestion

of starch by salivary
amylase and the

effect of temperature
and pH on it.

Every Health book insists on the chewing of food.

The act of chewing stimulates the excretion of
saliva. Saliva mixes up with the food and helps its
digestion. That is, the enzyme ptyalin or amylase
present in human saliva hydrolyzes the big
molecules of food into many molecules. For
example, starch into mono-saccnaricies, maltose
and glucose; proteins into amino acids and fats

into fatty acids and glycerol. Thus saliva not only

helps in digestion of food but convert it into
energy generating substances. Further, enzymes
and their activity are very sensitive to
temperature and pH. Even a slight variation in
these two factors, can disturb the action of
enzymes. In other words, digestion of food by
salivary amylase is also effected by pH and
temperature and can be verified experimentally.

An enzyme is a protein molecule that is a

biological catalyst with three characteristics.
First, the basic function of an enzyme is to
increase the rate of a reaction. Second, most
enzymes act specifically with only one reactant,
called a substrate, to produce products. The third
and most remarkable characteristic is that
enzymes are regulated from a state of low
activity to high activity and vice versa. The
activity of enzymes is strongly affected by
changes in pH and temperature. Each enzyme
works best at a certain pH and temperature, its
activity decreasing at values above and below
that point due to denaturation. For enzymes,

denaturation can be defined as the loss of

enough structure rendering the enzyme inactive.
This is not surprising considering the importance
of tertiary structure in enzyme function and noncovalent forces in determining the shape of
enzymes.
(P.T.O)

Salivary amylase is the enzyme produced by the

salivary glands. Formerly known as ptyalin, it
breaks down starch into maltose and isomaltose.
Amylase, like other enzymes, works as a catalyst.
All catalysts are enzymes, but not all enzymes
are catalysts. A catalyst is a substance that
hastens a chemical reaction but does not become
part of the end product. Amylase digests starch
by catalyzing hydrolysis, which is splitting by the
addition of a water molecule. The presence and
absence of starch can be confirmed by several
tests such as the iodine test. In general, a blueblack colour indicates the presence of starch.

1) Test tubes
2) Test tube stand
3) One dropper
4) Beaker
5) Starch and Iodine solution
6) Salivary amylase solution
7) Thermometer
8) Stop watch
9) Dil. HCl and NaOH solution
10) Heating arrangement

1) Collection of Saliva: - Rinse mouth thoroughly with cold

water and ensure that it does not contain any food
particles. Now take about 20ml of lupe warm water in the
mouth and gargle for about three minutes so that saliva
mixes up well with it. Spit this into a beaker. Filter, if
there is any suspended impurity. Clear filtrate is saliva
solution and contains enzyme ptyalin.
2) Preparation of starch solution - Take about 0.5g of starch
in a 100ml beaker and add enough water to make a
paste. Dilute the paste by adding 50ml water and boil for
about 5 min.
3) Digestion of starch

a. Take 5ml of the starch solution in a test tube. Add 2 ml

of saliva solution into it. Mix the solutions well by
shaking the tube carefully and start a step watch.
b. After one minute take out two drops of the mixture
solution from the test tube with the help of a dropper
and transfer it into another test tube containing about
one ml of 1% iodine solution. Note the colour
produced, if any.
c. Repeat this test after every one minute taking two

drops of the mixture solution and fresh 1% iodine

solution continue until the test shows no blue colour.
Record the time and blue colour intensity.

Take three test tubes and label these 1, 2,

and 3.
Take 5ml of the starch solution, 2ml of the
saliva solution and 5 ml of water in each test
tube.
Place test tube No. 1 in water at room
temperature, test tube No.2 in a beaker
containing water at 500 C and test tube No.3
in boiling water.
After 5 minutes, observe the colour change on
mixing two drops of the mixture of every tube
with one ml of 1% iodine solution. Note the
intensity of blue coloured form.

 Take three test tubes and label these 1, 2, 3.Add

5ml of the starch solution, 2ml of the saliva

solution in each test tube.
Now add 2 ml of water in test tube No. 1,
2 ml
of dil HCl in test tube No. 2 and 2ml of NaOH
solution in test tube No. 3 and shake carefully.
Keep the three test tubes in water at room
temperature for about 10 minutes.
Add two drops of the solution of each test tube
with 1% iodine solution and observe the colour
change.

TIME PASSED
AFTER
MIXING
COLOUR
INTENSITY

1 MINUTE

2 MINUTES

3 MINUTES

4 MINUTES

DEEP BLUE

BLUE

LIGHT BLUE

NO COLOUR

Absence of blue colour on addition to iodine solution

means absence of starch in the mixture solution. That is
whole of the starch has got digested or hydrolysed.

TEMPERATURE

COLOUR
INTENSITY

320C-370C
(ROOM
TEMPERATURE)
DEEP BLUE

500C-550C

LIGHT BLUE

700C-850C
(BOILING
WATER)
NO COLOUR

As the temperature increases, its enzymatic also

increases up until the optimum temperature.From the

experiment it is observed that the optimum temperature

of salivary amylase ranges from 32C to 37C.This
applies to the human body since salivary amylase is
suitable to function within these temperatures. After
37C, there is loss of activity indicated by colour change
from blue-black to light yellow colour of the solution. At
50C and 70C, salivary amylase is denatured. The
molecular conformation of the enzyme becomes altered
as the hydrogen bonds responsible for its secondary,
tertiary and quaternary structures are broken.

pH
COLOUR
INTENSITY

Dil. HCl (5.5)

Water(6.5-7)

DEEP BLUE

DEEP BLUE

NaOH
solution(9-10)
NO COLOUR

The optimum pH for the action of salivary amylase

ranges from 5.6 to 6.9.This is consistent with
observation between pH 4 and 6 (dil. HCl and water) in
table-3. Ideally at pH 10(NaOH solution), salivary
amylase is denatured due to high alkalinity. As pH
increases, certain amino acids such as lysine and
arginine are deprotonated, causing them to lose their
net positive charge which also results to enzyme
denaturation.

Several factors affect

the activity of enzymes.
Among these are the
temperature and pH. At
optimum levels of these
factors, enzymes
perform their function
best. Optimum
temperature and pH
differ from one enzyme
to another. Salivary
amylase is an enzyme

found in human saliva

which functions to break
down starch to simpler
compounds. Through
the experiment, it was
found out that the
optimum temperature of
salivary amylase ranges
from 32C to 37C and
its optimum pH ranges
from 5.6 to 6. 9.

Comprehensive lab manual class XII

www.britannica.com
www.google.com
Report on Study of effect of temperature
and pH on the enzymatic activity of
salivary amylase by Gae Khalil Rodillas,
Nonia Carla Ysabel Samson, Raphael Jaime
Santos and Brylle Tabora (Department of
Biological Sciences, College of Science,
University of Santo Tomas, Manila,
Philippines).