CHEMISTRY LABORATORY 528

Activity No. 8
Factors affecting enzyme activity

Aries Jay Bernal Reyes

MST – Chemistry

I- Introduction:
Enzymes are such efficient catalysts that they generally speed up reactions by a factor
of 10 – 1026 over the uncatalyzed rate of reaction. However the rates of enzyme controlled
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reactions are affected by a number of different factors. In most cases the effect of a factor on
the rate of the reaction is due to its influence on the active site.
At low substrate concentrations, each enzyme molecule is reacting with fewer substrate
molecules than is suggested by its turnover number. As the substrate concentration is
increased, each enzyme is able to locate and react with more substrate molecules and the
observed enzyme activity increases. However, once the turnover number is reached, the
addition of more substrate does not further increase the rate.
Under normal biological conditions the substrate is present in a large excess (there is
much more substrate than enzyme). As long as this condition is maintained, the addition of
more enzyme results in a proportional increase in rate.
Enzyme activity is influenced by pH with each enzyme having an optimum pH. The
optimum pH is the pH at which the activity of a particular enzyme is at a maximum. The
optimum pH's for enzymes found in the body are matched to the pH of the biological systems
in which they operate. For example, pepsin, a digestive enzyme, has an optimum pH of 1.5,
the pH of the stomach. At pH's far from the optimum pH, enzyme activity can be reduced to
nearly zero. If the reduction in activity is reversible the enzyme has been denatured. If it is
not reversible, the enzyme has been digested.
Just as each enzyme has an optimum pH, each has an optimum temperature as well.
Most human enzymes have an optimum temperature about that of body temperature (98.6 oF)
and are denatured or digested at extreme temperatures.
Some enzymes are capable of catalytic activity by themselves. Others require the
presence of an additional substance called a cofactor to induce this behavior. If the cofactor is
an organic compound, it is called a coenzyme. If it is a metal ion, it is called a metal ion
 activator. If a required cofactor is not present, the catalytic activity of the enzyme is
dramatically reduced.
Inhibitors are substances that reduce the rate of enzyme activity, usually by binding
with the enzyme and interfering with the formation of the enzyme-substrate complex. While
some heavy metals act as metal ion activators, they can also act as enzyme inhibitors.
II- Objectives:
a. To be able to determine the factors affecting enzyme activities .
b. To be able to compare the substances using the factors affecting enzyme
activities.
III- Data, Results & Discussion:
Table I: Effect of temperature
Time Test tube 1 – Cooked Test tube 2 – Cooked Test tube 3 – Cooked
Interval starch at 40oC starch at 60oC starch at 10oC
5 min. Light yellow Dark green Golden yellow
10 Light yellow Dark green Golden yellow
15 Light yellow Dark green Golden yellow
20 Light yellow Dark green Golden yellow
25 Golden yellow Dark green Golden yellow
30 Golden yellow Dark green Golden yellow
35 Golden yellow Dark green Golden yellow
40 Golden yellow Dark green Golden yellow
45 Golden yellow Dark green Golden yellow
50 Golden yellow Dark green Golden yellow
55 Golden yellow Dark green Golden yellow
60 Dark yellow Dark green Dark yellow
Discussion:
In this experiment, 3 test tubes were used to contain cooked starch with
saliva. The first test tube was heated at 40 oC, the second was heated at 60 oC and the
third one was cooled at 10 oC. Here we can see that as the temperature rises the
enzymes works faster until such time that it can no longer breakdown the molecules.
We can see that temperature affects the activity of enzymes.

Table II: Effect of pH

 Sample Reagents Added Results

Coagulated egg 5ml 2% pepsin + 0.4% HCl It almost completely dissolve

white the egg white

Coagulated egg 5ml 2% pepsin + 0.4%

white Na2CO3 The egg white is quite dissolve

Coagulated egg It almost completely dissolve

white 2% pancreatin + 0.4% HCl the egg white

Coagulated egg The egg white is quite dissolve

white 2% pancreatin + 0.4% Na2CO3

Discussion:
In the experiment, we can see that the coagulated egg white was dissolved in
pepsin and pancreatin with the help of HCl. This is because the enzyme activity was
influence by the pH of each enzyme with optimum pH which means that an enzyme,
at a given pH, can work best to digest the substance.

Table III: Influence of coenzymes

Sample Reagents Added Results
Glucose Red precipitate was
formed
Supernatant liquid Benedict’s reagent It takes a few seconds for
the red precipitate to form
Residue It takes a few seconds for
the red precipitate to form
Supernatant liquid + It takes a few seconds for
Residue the red precipitate to form
Discussion:
In this experiment, the yeast is mixed with white sand and water and was ground
and centrifuged. Afterwards the supernatant liquid and the residue were used to be
mix with the Benedict’s soln. It takes a few seconds to form red precipitate. This is
because some enzymes are capable of catalytic activity by themselves. Others require the
presence of an additional substance called a cofactor to induce this behavior. If the cofactor is
an organic compound, it is called a coenzyme.
IV- Conclusion:
 In this experiment, enzymes are affected by the following factors: pH,
Temperature and influence of coenzymes. An enzyme at an optimum pH works best
and breaks down substances easily. Just as enzyme has optimum pH, it has also its
optimum temperature wherein at a given temperature they work best and speed up
reaction to breakdown substance but if it reaches its maximum point, it loses it
capability to breakdown substances instead the enzyme has denatured.