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Complete report of Basic Biology with the title Influence of PH to Enzyme

Activities, created by :
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: Bertha Tandi
reg. Number : 1414442010
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: ICP B Biology
after its checked and consulted by Assistant and Assistant Coordinator, it has
fulfilled requirement.
Makassar, January
Assistant Coordinator

Djumarirmanto S.Pd

2014

Assistant

Muh. Nur Arsyad

ID . 09104158
Known by,
Lecturer of Responsibility

Drs. H. Hamka L.Ms

NIP. 19621231 198702 1 005

CHAPTER I
INTRODUCTION
A. Background
Metabolism includes all the chemical reactions that occur in the body of
organisms. The metabolic process is the enzymatic reaction, meaning that the
reaction involving the enzyme. Enzymes are substances in each of the reactions in
living organisms that act as biocatalisator, which is a catalyst in chemical

reactions of living cells. As a catalyst, the enzyme is a chemical agent that can
change the rate of a reaction without he himself had to change because of the
reaction. With the enzyme, the traffic through the chemical - metabolic pathways
run more smoothly. In these reactions are described substances and substances
that are formed, the substances described by the reaction are called substrates, and
the newly formed from the reaction are called products.
In a chemical reaction, the enzyme did not change shape, do not affect the
balance of the reaction or change the product. The reaction of complex chemical
reactions in the body will last a very slow if without enzymes. This enzyme works
in a liquid aqueous solution, temperature, and pH are in accordance with the
biological physiological conditions. Through its activities, the enzymes are well
coordinated so as to produce a good relationship between a number of different
metabolic activity, which refers to support in our life. Enzyme not determine the
direction of the reaction (back - back), can be used repeatedly for not damaged,
needed in small amounts. The enzyme has a different temperature and pH different to be able to work optimally, because if the temperature and acidity are
not in accordance with the nature of an enzyme, the enzyme cannot work
optimally, inactive, even experienced denaturation (damage). The enzyme can be
found in animals and plants. One such enzyme is an enzyme amylase (diastase)
contained in the plant. Enzyme can hydrolyze starch into sugar. Amylase
produced by the leaves or seeds which are for prove the above statement, we
conducted an experiment entitled "effect of pH on enzyme activity".
B. Purpose
The purpose of this practicum, to showing influenced pH about amylase
enzyme activity.
C. Benefit
Based on this practicum the benefit of this practicum the university
student will know about influenced pH about amylase enzyme activity.

CHAPTER II
PREVIEW OF LITERATURE
An enzyme is a macromolecule that acts as a catalyst, chemical agent that
speeds up a reaction without being consume by the reaction. (In this chapter, we are
focusing on enzymes that are proteins. RNA enzymes, also called ribozymes, without
regulation by enzymes, chemical traffic through the pathways of metabolism would
become terribly congested because many chemical reactions would take such a long

time. In the next two sections, we will see what prevents a spontaneous reaction from
occurring faster and how an enzyme changes the situation (Campbell, 2011: 152).
Enzymes can be found both in animals and in plants. One of the enzymes
found in plants is amylase. Other names of amylase is amylase hydrolyze latase. The
enzyme can amylase hydrolyze starch into sugar. Amylase produced by the leaves or
seeds germinating. Amylase activity is affected by inorganic salts, pH, temperature,
and light. The optimum pH of amylase hydrolyze according to Hopkins, Cole, and
Green is 4.5 to 4.7 (Tim Dosen, 2014: 32).
An enzyme catalyzes a reaction by lowering the EA barrier enabling the
reactant molecules to absorb enough energy to reach the transition state even at
moderate temperatures. An enzyme cannot change the G for a reaction; it cannot
make an endergonic reaction exergonic. Enzymes can only hasten reactions that
would eventually occur anyway, but this function makes it possible for the cell to
have a dynamic metabolism, routing chemicals smoothly through the cells metabolic
pathways. And because enzymes are very specific for the reactions they catalyze, they
determine which chemical processes will be going on in the cell at any particular
time. The reactant an enzyme acts on is referred to as the enzymes substrate. The
enzyme binds to its substrate (or substrates, when there are two or more reactants),
forming an enzyme substrate complex. While enzyme and substrate are joined, the
catalytic action of the enzyme converts the substrate to the product (or products) of
the reaction. The overall process can be summarized as follows:
Enzyme _ Enzyme- Enzyme _
Substrate(s) substrate Product(s) complex
For example, the enzyme sucrase (most enzyme names end in -ase) catalyzes the
hydrolysis of the disaccharide sucrose into its two monosaccharides, glucose and
fructose.
Sucrase _ Sucrase- Sucrase _
Sucrose _ sucrose-H2O Glucose _H2O complex Fructose

The reaction catalyzed by each enzyme is very specific; an enzyme can recognize its
specific substrate even among closely related compounds. For instance, sucrase will
act only on sucrose and will not bind to other disaccharides, such as maltose. What
accounts for this molecular recognition? Recall that most enzymes are proteins, and
proteins are macromolecules with unique three-dimensional configurations. The
specificity of an enzyme results from its shape, which is a consequence of its amino
acid sequence (Campbell, 2011: 152).
Most enzymes are active only over a narrow pH range and have an optimal
pH, at which the rate of reaction is fastest. The optimal pH for most human enzymes
is between 6 and 8. (Recall from Chapter 2 that buffers minimize pH changes in cells
so that the pH is maintained within a narrow limit.) Pepsin, a protein digesting
enzyme secreted by cells lining the stomach, is an exception; it works only in a very
acidic medium, optimally at pH 2. In contrast, trypsin, a protein-splitting enzyme
secreted by the pancreas, functions best under the slightly basic conditions found in
the small intestine. The activity of an enzyme may be markedly changed by any
alteration in pH, which in turn alters electric charges on the enzyme. Changes in
charge affect the ionic bonds that contribute to tertiary and quaternary structure,
thereby changing the proteins conformation and activity. Many enzymes become
inactive, and usually irreversibly denatured, when the medium is made very acidic or
very basic (Salomon, 2008: 164).
Effects of Temperature and pH Recall from Chapter 5 that the threedimensional structures of proteins are sensitive to their environment. As a
consequence, each enzyme works better under some conditions than under other
conditions, because these optimal conditions favor the most active shape for the
enzyme molecule. Temperature and pH are environmental factors important in the
activity of an enzyme. Up to a point, the rate of an enzymatic reaction increases with
increasing temperature, partly because substrates collide with active sites more
frequently when the molecules move rapidly. Above that temperature, however, the
speed of the enzymatic reaction drops sharply. The thermal agitation of the enzyme

molecule disrupts the hydrogen bonds, ionic bonds, and other weak interactions that
stabilize the active shape of the enzyme, and the protein molecule eventually
denatures. Each enzyme has an optimal temperature at which its reaction rate is
greatest. Without denaturing the enzyme, this temperature allows the greatest number
of molecular collisions and the fastest conversion of the reactants to product
molecules. Most human enzymes have optimal temperatures of about 3540C (close
to human body temperature). The thermophilic bacteria that live in hot springs
contain enzymes with optimal temperatures of 70C or higher. Just as each enzyme
has an optimal temperature, it also has a pH at which it is most active. The optimal
pH values for most enzymes fall in the range of pH 68, but there are exceptions. For
example, pepsin, a digestive enzyme in the human stomach, works best at pH 2. Such
an acidic environment denatures most enzymes, but pepsin is adapted to maintain its
functional three-dimensional structure in the acidic environment of the stomach. In
contrast, trypsin, a digestive enzyme residing in the alkaline environment of the
human intestine, has an optimal pH of 8 and would be denatured in the stomach
(Campbell, 2011: 155).
In an enzyme-catalyzed reaction, the reactants are called substrates. Substrate
molecules bind to a particular site on the enzyme, called the active site, where
catalysis takes place. The specificity of an enzyme results from the exact threedimensional shape and structure of its active site, into which only a narrow range of
substrates can fit. Other moleculeswith different shapes, different functional
groups, and different propertiescannot properly fit and bind to the active site. The
names of enzymes reflect the specificity of their functions a reaction is recovered
during the ensuing downhill phase of the reaction, so it is not a part of the net free
energy change and often end with the suffix -ase. For example, the a reaction is
recovered during the ensuing downhill phase of the reaction, so it is not a part of
the net free energy change (Heller, 2011: 113-114).
Almost every biological process is pH dependent; a small change in pH
produces a large change in the rate of the process. This is true not only for the many

reactions in which the H_ ion is a direct participant, but also for those in which there
is no apparent role for H_ ions. The enzymes that catalyze cellular reactions, and
many of the molecules on which they act, contain ionizable groups with characteristic
pKa values. The protonated amino and carboxyl groups of amino acids and the
phosphate groups of nucleotides, for example, function as weak acids; their ionic
state depends on the pH of the surrounding medium. As we noted above, ionic
interactions are among the forces that stabilize a protein molecule and allow an
enzyme to recognize and bind its substrate. Cells and organisms maintain a specific
and constant cytosolic pH, keeping biomolecules in their optimal ionic state, usually
near pH 7. In multicellular organisms, the pH of extracellular fluids is also tightly
regulated. Constancy of pH is achieved primarily by biological buffers: mixtures of
weak acids and their conjugate bases. We describe here the ionization equilibria that
account for buffering, and we show the quantitative relationship between the pH of a
buffered solution and the pKa of the buffer. Biological buffering is illustrated by the
phosphate and carbonate buffering systems of humans. Although many aspects of cell
structure and function are influenced by pH, it is the catalytic activity of enzymes that
is especially sensitive. Enzymes typically show maximal catalytic activity at a
characteristic pH, called the pH optimum). On either side of the optimum pH their
catalytic activity often declines sharply. Thus, a small change in pH can make a large
difference in the rate of some crucial enzyme-catalyzed reactions. Biological control
of the pH of cells and body fluids is therefore of central importance in all aspects of
metabolism and cellular activities (Salomon, 2008: 160).
PH affects enzyme activity the rates of most enzyme-catalyzed reactions
depend on the pH of the medium in which they occur. Each enzyme is most active at
a particular pH; its activity decreases as the solution is made more acidic or more
basic than its ideal (optimal) pH several factors contribute to this effect. One is the
ionization of carboxyl, amino, and other groups on either the substrate or the enzyme.
In neutral or basic solutions, carboxyl groups (COOH) release H+ to become
negatively charged carboxylate groups (COO). Similarly, amino groups (NH2)

accept H+ ions in neutral or acidic solutions, becoming positively charged NH3+

groups (see Chapter 2). Thus, in a neutral solution, a molecule with an amino group is
attracted electrically to another molecule that has a carboxyl group, because both
groups are ionized and the two groups have opposite charges. If the pH changes,
however, the ionization of these groups may change. For example, at a low pH (high
H+ concentration), the excess H+ may react with the COO to form COOH. If this
happens, the group is no longer charged and cannot interact with other charged
groups in the protein, so the folding of the protein may be altered. If such a change
occurs at the active site of an enzyme, the enzyme may no longer have the correct
shape to bind to its substrate (Heller. 2011: 122).
Cells regulate the rates of chemical reactions with enzymes, which are
biological catalysts that increase the speed of a chemical reaction without being
consumed by the reaction. Although most enzymes are proteins, scientists have
learned that some types of RNA molecules have catalytic activity as well . The
catalytic ability of some enzymes is truly impressive. For example, hydrogen
peroxide (H2O2) breaks down extremely slowly if the reaction is un catalyzed, but a
single molecule of the enzyme catalase brings about the decomposition of 40 million
molecules of hydrogen peroxide per second! Catalase has the highest catalytic rate
known for any enzyme. It protects cells by destroying hydrogen peroxide, a
poisonous substance produced as a by-product of some cell reactions. The
bombardier beetle uses the enzyme catalase as a defense mechanis (Salomon, 2008:
163).

CHAPTER III
OBSERVATION METHOD

A. Place and Date

Day / date
: Monday/ January th 2010
Time
: 16.00 Wita 18.00 Wita
Place
: Laboratory of Biology, 3rd west floor Mathematics and Science
Faculty State University of Makassar
B. Tools and Materials
1. Tools
a. 10 pieces Test tube
b. 1 piece Test tube rack
c. 5 pieces Dropping pipette
d. 1 piece Bunsen burner
e. 3 pieces Universal indicator
f. 1 piece Beaker glass
2. Material
a. Extract of phaseolus sprout
b. Amylum solution
c. Fehling solution A and B
d. HCl solution (10%)
e. NaOH solution (1%)
f. PH paper
g. Filter paper
h. Aquadest
i. 1 piece Matches
C. Work Procedure
1. Prepared 10 units of test tube and its rack. Divided the tubes in to 4 groups
(Tube A, B, C, and D). Tube A, B and C were classified in to 3 tubes and gave
them (A1/B1/C1, A2/B2/C2, And A3/B3/C3). For tube D, it was independent.
2. For A tubes, dropped 2 drops of starch solution and then dropped 1 mL of
extract.
3. Added 2 drops of fehling A and B, measured the solution pH using pH meter.
Observed the initial color.
4. Let the 3 tubes about a few minutes. A1 during 5 minutes, A2 during 10
minutes, A3 during 15 minutes. After that heated each tubes.
5. After heating the tubes, observed each colors.
6. Wrote down your observation.
7. For B tubes, dropped 2 drops of starch solution and then dropped 1 mL of
extract, after that added 2 drops of NaOH, done the same procedures from
step 3 to step 6.

8. For C tubes, dropped 2 drops of starch HClsolution and then dropped 1 mL of

extract, after that added 2 drops of. Done the same procedures from step 3 to
step 6.
9. For D tube, dropped 2 drops o starch solution and then dropped 1 mL of
extract. Gave addition of fehling A and B about 2 drops. And then, observed
the indicial color.
10. Heated the tube, and then observed the colors changing.

CHAPTER IV
RESULT AND DISCUSSION
A. Observation Result
1. Observation Table
No
Tube

pH

First Color

Last Color

Blue

Yellow Orange

Blue

Orange Turbid

.
A1
1.

A2

12

A3

Blue

B1

2.

3.

B2

Light Yellow
White Turbid +

White

11

Sediment
White Turbid +

White

Sediment
White Turbid +

B3

White

C1

Clear Yellow

Light Yellow

Clear Yellow

Light Yellow

Clear Yellow

Light Yellow

C2
C3

Sediment

4.

13

Blue

Light Blue

2. Figures of Observation
1)Tube I
a. Tube I A

Note :
1. The addition of 1 ml of starch
and germ extract.
2. The addition of 2 drops of
Fehling A and B.
3. After heated for 5 minutes.
1

b. Tube I B

Note :
1. The addition of 1 ml of
starch and germ extract.
2. The addition of 2 drops

of Fehling A and B.
3. After heated for

10

minutes.

c. Tube I C

Note :
1. The addition of 1 ml of
starch and germ extract.
2. The addition of 2 drops of
1

Fehling A and B.
3. After heated for
minutes.

15

2) Tube II
a. Tube II A

Note :
1. The addition of 1 ml of starch

and germ extract.

2. The addition of 3 drops of
NaOH.
3. After heated for 5 minutes.

b. Tube II B

Note :
1. The addition of 1 ml of starch
and germ extract.
2. The addition of 3 drops of

NaOH.
3. After heated for 10 minutes.

c. Tube II C

Note :
1. The addition of 1 ml of starch
and germ extract.
2. The addition of 3 drops of
NaOH.
3. After heated for 15 minutes.
1

3) Tube III
a. Tube III A

Note :
1. The addition of 1 ml of
starch and germ extract.
2. The addition of 3 drops of
1

HCl.
3. After heated for 5 minutes.

b. Tube III B

Note :
1. The addition of 1 ml of starch
and germ extract.
2. The addition of 3 drops of HCl.
3.

4. After heated for 10 minutes.

c. Tube III C

Note :
1. The addition of 1 ml of starch
and germ extract.
2. The addition of 2 drops of
1

HCl
3. After heated for 15 minutes.

4. Tube IV

Note :
1. The addition of 1 ml of starch
and germ extract.
2. The addition of 2 drops of
1

Fehling A and B.
3. This direct heated.

B. DISCUSSED
In the tube 1 in the tube given amylum, added fehling A and B, and added
green peal obtained the 12 pH and we heated the tube 1A. 1B, and 1C with
different time and the color was change in the tube 1A from blue became the
yellow orange, tube 1B from blue into orange turbir, and the 1C from dark blue
into light yellow. So for the tube 1 is basa.
In the tube 2 in the tube given amylum, added fehling A and B, added
extract green peal and 3 drop NaOH obtained the 11 pH and we heated the tube
2A. 2B, and 2C with different time and the color was change in the tube 2A from
white became the White Turbid + Sediment, tube 2B from white into White
Turbid + Sediment, and the 2C from white into White Turbid + Sediment. So it
mean the tube 2 is base.

In the tube 3 in the tube given amylum, added fehling A and B, added
extract green peal and 3 drop HCl solution obtained the 1 pH and we heated the
tube 3A. 3B, and 3C with different time and the color was change in the tube 3A
from clear yellow became the light yellow, tube 3B from clear yellow into light
yellow, and the 3C from clear yellow into light yellow. So it mean the tube 3 is
acid.
The last tube 4 wasnt done anything just give fehling A and B. So the
result in the tube 4 is bases.

CHAPTER V
CONCLUSION AND SUGGESTION
A. Conclusion
Based on the practicum about the influence the PH of enzyme activity, the
practicum draw the conclusion as follows:
The pH of the substrate it can be evidence the enzyme activity which is
enzyme function is to make fast the reaction.
B. Suggestion
In conducting the experiment, must be done seriously, meticulous when
dripping the solution until the solution is dripped do too much. Perform
experiments in accordance with workplace procedures.

BIBLIOGRAPHY
Neil A, Campbell. Reece, Jane B. 2009. Campbell biology ninth edition. Amerika
Serikat : MasteringBiology and BioFlix.
Orians, Heller. 2011. The Science of Biology 7Th Edition. New York: Porves Sadava.
Solomon, Eldra P. Berg, Linda R. 2008. Biology eighth edition. Amerika Serikat: The
part of Thomson Corporation.
Tim Pengajar Biologi. 2010. Penuntun Praktikum Biologi Dasar. Makassar:
Laboratorium FMIPA UNM.

APPENDIX
A. Question
1. What to Fehling's solution A and Fehling B and JKJ?
2. Why the enzyme extract from the seeds need centrifuge?

3. What is the function of HCl and NaOH in this practicum?

B. Answer
1. Usefulness of Fehling's solution A and Fehling B to test whether an extract or a
solution containing glucose or not. JKJ solution is used to test whether a
solution containing starch or not.
2. Enzymes from seeds that need centrifuge mix solid substances and liquid can be
separated so that the solids can settle to the bottom while the liquid remains in
the liquid part. Or is this liquid that is part amylase containing extracts on
germination.
3. The function of HCl and NaOH in this practicum to influence if the solution
acid and base.