CHAPTER I
INTRODUCTION
A.
Background
Inside the body
of life creatures, chemistry reaction was happened. Reactions which were
happening in their bodies happen in temperature 27 oC (room
temperature), for example on plant bodies. Beside that, it also can happen in
teperature of 39 oC, for example inside the body of awarm blood
animal. In that temperature , oxidation process will be happen slowly
or even
doesn’t happen. So that the reaction will be happen faster we need catalysator
for it. Catalysator is a subtance which can make the reaction work faster than
before without join on that reactions. Catalysator inside the cells of life
creature is called biocatalysator or we know that as enzyme.
Because of
enzyme’s function is to make the reaction more faster and it doesn’t join to
the reaction, so the amount is not too much. One molecule of enzyme can work
many times during the enzyme was not damaged. Working of an enzyme was influenced
by some factors, that temperature, pH,product, and inhyibitor. For knowing how
far the factors influence the work of enzyme, where we took pH to observed, so
we did this experiment to know about the factors which influence the work of
enzyme.
A chemistry
reaction, especially between organic compound, which do in laboratory need a
condition which is determined by some factors which are temperature, pressure,
time, and so on. When one of the
condition is not comfort with what should we need so the reaction doesn’t work
well. Our bodies is a complicated laboratory, because in that body happen
various chemistry reaction. Decomposition of subtances there are in our food, use result of description to get energi, Penguraian zat-zat yang terdapat
dalam makanan kita, penggunaan hasil uraian untuk memperoleh energi, affiliation return result of description
to form the food supply in body and also a lot of kinds of other reaction which
is if done in laboratory or in vitro require the specialty and also time old
ones,
can take place better
in body or in vivo not requiring high temperature and can be happened during
which relative shorten. React or chemical process that goes on better in our
body this enabled [by] caused by [his/its] [is] katalis of[is so-called enzyme.
Chemical process that happened by force of enzyme have been recognized [by]
since former epoch for example out of job making by ferment or fermentation.
Pursuant to that m atter, praktikan will [do/conduct] this praktikum with the
title of praktikum of influence pH to enzyme activity to know the influence pH
to aktifitas enzyme by manipulation of pH amilum with the addition HCL, NaOH,
and also Fehling A and Fehling B.
B. Purpose
To
prove the effect of pH on amylase enzyme activity.
C. Benefits
The
student can prove the effect of pH on amylase enzyme activity.
CHAPTER
II
PREVIEW OF
LITERATURE
Enzymes
can be found both in animals and in plant. At an enzymes found in plants is
amylase. Another name of aylase is Diastase. Enzymes can hydrolyze starch into
sugar. Amylase produced by the leaves ore seeds are germinating. Amylase
activity is affected by inorganic salts, pH, temperature, and light. The
optimum pH of amylase according to Hopkins, Cole, and Green (Miller,1938) is
4,5 to 4,7 (Team Lecturer,2012).
For
every enzyme, there is an optimum pH value at which the specific enzyme
functions most actively. Any change in this pH significantly affects the enzyme
activity and/or the rate of reaction. To know more about the relation between
pH and enzymes and/or pH effect on enzymes, read on (Ningthoujam Sandhyaran, 2011).
Enzymes
are proteinaceous catalysts, which speed up the rate of a biochemical reaction.
They reduce the activation energy that is essential for starting any type of
chemical reaction. With a low energy requirement for activation, the reaction
takes place faster. The overall performance of an enzyme depends on various
factors, such as temperature, pH, cofactors, activators and inhibitors. You
might have a fair idea regarding pH effect on enzymes. But, why and how does pH
and temperature affect enzymes? This article highlights on the enzyme activity
with reference to change in the pH level.The rate of a chemical reaction and/or
the enzyme activity is greatly influenced by the structure of the enzyme. Or in
other words, a change in the structure of the enzyme affects the rate of reaction.
When pH of a particular medium changes, it leads to alteration in the shape of
the enzyme. Not only on enzymes, the pH level may also affect the charge
properties and shape of the substrate. Within a narrow pH range, changes in the
structural shapes of the enzymes and substrates may be reversible. But for a
significant change in pH levels, the enzyme and the substrate may undergo
denaturation. In such cases, they cannot identify each other. Consequently,
there will be no reaction as such. This the reason why, pH affect enzyme
activity.Speaking about the connection between pH and enzymes and/or pH effect
on enzymes, each and every enzyme is characterized by an optimum pH. At this
specific pH level, a particular enzyme catalyzes the reaction at the fastest rate
than in any other pH level (Ningthoujam Sandhyaran,2011).
For example, the enzyme pepsin (a protease
enzyme) that catalyzes proteins is most active at an acidic pH, whereas the
enzyme trypsin (another protease enzyme) performs best at a slightly alkaline
pH. Thus, the optimum pH of an enzyme is different from that of another
enzyme.When we study pH, it is clearly defined as the measurement for the
acidic or alkaline nature of a solution. To be more precise, pH indicates the
concentration of dissolved hydrogen ions (H+) in the particular
solution. An increase or decrease in the pH changes the ion concentration in
the solution. These ions alter the structure of the enzymes and at times, the
substrate either due to formation of additional bonds or breakage of already
existing bonds. Ultimately, the chemical makeup of the enzyme and substrate are
changed. Also, the active site of the enzyme is changed, after which the
substrate can no longer identify the enzyme. For more information on enzymes,
you can refer to enzyme substrate complex.Consider
a case, when the reaction is adjusted at a pH level different from the optimum
value. Over here, the rate of reaction or the activity of enzymes will not be
the same as the previous one. At times, you will notice that there is no
reaction at all. This occurs, when there is changes in the structure of the
active site and the substrate. Hence, for the chemical reaction to take place,
you need to adjust the pH of the solution in such a way that it is suitable for
both the enzyme and the substrate. This way, pH effect on enzymes activity can
be studied practically (Ningthoujam Sandhyaran, 2011).
pH
is a measure of the concentration of hydrogen ions in a solution.
The higher the hydrogen ion concentration, the lower the pH. Most enzymes function efficiently over a narrow pH range. A change in pH above or below this range reduces the rate of enzyme reaction considerably. Changes in pH lead to the breaking of the ionic bonds that hold the tertiary structure of the enzyme in place. The enzyme begins to lose its functional shape, particularly the shape of the active site, such that the substrate will no longer fit into it, the enzyme is said to be denatured. Also changes in pH affect the charges on the amino acids within the active site such that the enzyme will not be able to form an enzyme-substrate complex. The pH at which an enzyme catalyses a reaction at the maximum rate is called the optimum pH. This can vary considerably from pH 2 for pepsin to pH 9 for pancreatic lipase (Anonymous, 2012).
The higher the hydrogen ion concentration, the lower the pH. Most enzymes function efficiently over a narrow pH range. A change in pH above or below this range reduces the rate of enzyme reaction considerably. Changes in pH lead to the breaking of the ionic bonds that hold the tertiary structure of the enzyme in place. The enzyme begins to lose its functional shape, particularly the shape of the active site, such that the substrate will no longer fit into it, the enzyme is said to be denatured. Also changes in pH affect the charges on the amino acids within the active site such that the enzyme will not be able to form an enzyme-substrate complex. The pH at which an enzyme catalyses a reaction at the maximum rate is called the optimum pH. This can vary considerably from pH 2 for pepsin to pH 9 for pancreatic lipase (Anonymous, 2012).
All biological systems are based on the same organic
molecules, a similarity that is one of many legacies of life’s common origin.
However, the details of those molecules differ among organisms. Simple organic
building blocks bonded in different numbers and arrangements form different
versions of the molecules of life, just as atoms bonded in different numbers
and arrangements form different molecules. Cells maintain reserves of small
organic molecules that they can assemble into complex carbohydrates, lipids,
proteins, and nucleic acids. When used as subunits of larger molecules, the
small organic molecules (simple sugars, fatty acids, amino acids, and
nucleotides) are called monomers.
Molecules that consist of multiple monomers are called polymers. Cells build polymers from monomers, and break down
polymers to release monomers (Starr,2011).
Metabolism refers to activities by
which cells acquire and use energy as they make and break apart organic
compounds. These activities help cells stay alive, grow, and reproduce.
Metabolism also requires enzymes,
which are organic molecules that speed up reactions without being changed by
them (Starr,2011).
“Saccharide” is from sacchar, a Greek word that
means sugar.Monosaccharides (one sugar unit) are the simplest type of
carbohydrate, but they have extremely important roles as components of larger
molecules. Common monosaccharides have a backbone of five or six carbon atoms,
one carbonyl group, and two or more hydroxyl groups. Enzymes can easily break
the bonds of monosaccharides to release energy . The solubility of these
molecules also means that they move easily throughout the water-based internal
environments of all organisms. Monosaccharides that are components of the
nucleic acids DNA and RNA have five carbon atoms. Glucose (at left) has six.
Glucose can be used as a fuel to drive cellular
processes,
or as a structural material to build larger molecules. It can also be used as a
precursor, or parent molecule, that is remodeled into other molecules. For
example, cells of plants and many animals make vitamin C from glucose (human
cells cannot, so we need to get our vitamin C from food). Cellulose does not
dissolve in water, and it is not easily broken down. Some bacteria and fungi
make enzymes that break it apart into its component sugars, but humans and
other mammals do not. When we talk about dietary fiber, or “roughage,” we are
usually referring to the cellulose and other indigestible polysaccharides in
our vegetable foods. Bacteria that live in the gut of termites and grazers such
as cattle and sheep help these animals digest the cellulose in plants
(Starr,2011).
All living things are
chemical factories driven by chemical reactions. However, these chemical
reactions proceed very slowly when carried out in the laboratory because the
activation energy is high. To be useful to living organisms, additional
substances must be present where the chemical reactions occur to reduce the
activation energy and allow the reaction to proceed quickly. A catalyst is a substance that lowers
the activation energy needed to start a chemical reaction. Although a catalyst
is important in speeding up a chemical reaction, it does not increase how much
product is madeand it does not get used
up in the reaction (Biggs,2008).
In our overview of food processing, we have seen
that digestive enzymes hydrolyze
the same biological materials (such as proteins, fats, and carbohydrates) that
make up the bodies of the
animals themselves. How, then, are animals able to digest food without digesting
their own cells and tissues? The evolutionary adaptation found across a wide
range of animal species
is the processing of food within specialized compartments. Such compartments can
be intracellular, in the form
of
food vacuoles, or extracellular, as in digestive organs and systems. Ingestion and
the initial steps of digestion occur in the mouth, or oral cavity. Mechanical
digestion beginsas teeth of various shapes cut. smash, and grind food. making
the food easier to
swallow
and increasing its surface area. Meanwhile, the presence of food stimulates a
nervous reflex that causes the salivary
glands to deliver saliva through ducts to theoraJ cavity. Saliva may also be
released before food enters the mouth, triggered by a learned association
between eating and the time of day, a cooking odor, or another stimulus. Saliva
initiates chemical digestion while also protecting the oral cavity. Amylase. an
enzyme in saliva, hydrolyzes starch (a glucose polymer from plants) and
glycogen (a glucose polymer from animals) into smaller polysaccharides and the
disaccharide maltose. Mucin, a slippery glycoprotein (carbohydrateprotein
complex) in saliva. protects the lining of the mouth from abrasion. Mucin also
lubricates food for easier swallowing. Additional components of saliva include
buffers, which help prevent tooth decay by neutralizing acid, and antibacterial
agents, which protect against microorganisms that enter the mouth with food
(Campbell,2008).
CHAPTER III
PRACTICUM METHOD
A.
Time
and Place
Day / Date : Friday / December 7th
2012
Time :
at 08.10 - 09.10 am
Place : Laboratory of Biology at
3rd floor of BiologyDepartement of Science and Mathematic Faculty,
State University of Makassar
B.
Tools
and Materials
1. Tool
a. Centrifuge
and centrifuge tubes
b. 10
test tubes
c. Pipette
d. Small
Funnel
e. Test
tube rack
f. Lights
spiritus
g. pH
Paper / pH meter
2. Material
a. Sprout
of Green Bean
b. Starch
solution
c. Fehling
Solution A and B
d. Dilute
HCl (10%)
e. 1%
NaOH solution
f. Aquades
C.
Work
Procedure
1. Took
sprout of green bean and put it on the 10 tubes.Entered sprouts extract
obtained into the tube I, checked and recorded the pH. Furthermore, the liquid
into 3 tiny tubes, labeled a, b, c. After 10 minutes, added or Fehling solution
A and B into a tube. After 15 minutes, added the same subtances in the tube c.
Recorded the color. Then heated all of the tubes in the time.
2. In
the second tube added drops of NaOH solution, pH checked and recorded. Further
treatment such as no. 1.
3. In
the third tube added 1 drop of dilute HCl, checked and recorded the pH. Further
treatment such as no. 1.
4. In
the IV tube added Fehling A and B and note the color.
5. Compared
the color that occurs on the tubes, created a table and concluded.
CHAPTER
IV
RESULT AND
DISCUSSION
A.
Result
1.
Observation
Table
|
Nu.
Tube
|
pH
|
Change
|
|
|
Before
Heating
|
After Heating
|
||
|
I
|
6
|
a.White
|
a.White greenly
|
|
b.White
|
b.Yellowish green
|
||
|
c.White
|
c.Yellow greenness
|
||
|
II
|
12
|
a.Yellowish green
|
a.Golden yellow
|
|
b.Yellowish green
|
b.Old green
|
||
|
c.Yellowish green
|
c.Brown yellow
|
||
|
III
|
1
|
a.White
|
a.Transparent
|
|
b.White
|
b.White Blue
|
||
|
c.White
|
c.White
|
||
|
IV
|
|
White
|
Yellowish green
|
2.
Observation
Tubes
a) Before
Heated
 |
|||
 |
|||
 |
 |
b) After
Heated
 |
|||||||
 |
|||||||
 |
 |
||||||
B.
Discussion
In the 1st tube
was added with sprouts
extracts
and the pH of it is 6, after that it is added again with amilum and fehling A
and B after 5 minutes the color is white. Actually this tube is divided to be 3
tube and those are given same material but the different is the time when enter
the material. But they have same color so we just take one tube to be observer.
This tube is warmed three times. For the first warm, after 5 minutes the color
change from white to be white greenly.
And then it was warmed again and after 10 minutes the color become white to
yellowish green, like that also the color for the third warm. It change from
white to yellow greenness. The change of the color happened because the
lateness have sour and alkali characteristic.
We also have
done the same with the first tube, in the 2nd tube is added also
with sprouts extracts, amilum and fehling A and B. but this tube was given also
with NaOH solution lateness. And when the pH is measured, it had pH =12, and
the color was yellowish green. This tube was warmed three times too, for the first
warm after 5 minutes the color change from yellowish green be golden yellow,
for the second warm the color be old green, and the third warm the color yellow
brown. This happened because the lateness had sour and alkali characteristic.
And it same meaning if there was enzim
that gave influence to the amilum.
For the 3rd
tube is same with the first and the second tube, it was added with sprouts
extracts, amilum and fehling A and B, the different was this tube HCl lateness.
The color is white and its pH is 1. This tube was given three times warm also.
When the first warm after 5 minutes the color was change from white be tansparent, then for the second
warm after 5 minutes the color be white blue, the third warm the color is still
white. So, there was enzyme that gave influence for amilum, so that the amilum
be change.
In the 4th
tube was added also with sprouts extracts, amilum and fehling A and B. The
color was white, after 5 minutes and warmed until the practicum had finished
the color become yellowish green. This happen because there was not enzyme that
gave influence for the amilum to be glucose.
CHAPTER
V
CONCLUSION AND SUGGESTION
A.
Conclussion
Based
on experiment, we have known that pH has an influences for enzymes activities. Enzyme will
work on optimal pH, such as enzyme amylase work in pH between 4,5 – 4,7. The
change of color is mean that the enzyme is working although the color is not
comfort with the theory and the enzyme doesn’t work optimal.
B.
Suggestion
1.
Laboratory should prepare well the tools
which will be used in experiment.
2. In
doing an experiment we must be careful when use the tools to avoid the accident
which probably will happen. We must observe the object carefully and seriously
so that we can find a good result.
3. The
assistant should give command so we can miss the mistake while doing the
experiment.
.
BIBLIOGRAPHY
Accessed on December 8th 2012
Sandhyarani
Ningthoujam . 2011. pH Effect on
Enzymes. http://www.buzzle.com.
Accessed on December 8th 2012
Biggs, at. el. 2008. Biology.United States of America:
Glencoe.
Campbell, at. el. 2009. Biology. San Francisco: Benjamin
Commings.
Starr, at. el. 2011. Biology. Canada: Cengange
Team
Lecturer. 2012. Basic Biology Guide Book. Makassar: Biology
Departement Faculty of Mathematic and Science, State University of Makassar.
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