EXPERIMENT 2: ENZYMES ❖ Proenzyme pepsinogen is released by

MC-2 LAB the chief cells in the stomach wall. For

it to be activated it reacts with
hydrochloric acid of the gastric juice and
OBJECTIVES of the Experiment
becomes pepsin.
● Demonstrate the catalytic action of
enzymes through different organic SAMPLES Observations
specimen.
A1 = Distilled water No obrservable
● Recognize the specificity of enzymes to (rate 3) changes
its substrate.
● Identify the products of each reaction A2 = 1% Pepsin Cloudiness can be
through different color tests. solution (rate 2) seen
● Describe the role of enzyme in digestive Boiled egg formed
A3 = 1% Pepsin with
processes. some hair like
0.4% HCl (rate 1)
precipitates
ENZYMES
❖ Enzymes are substances that act as a For theoretical
catalyst for biochemical reactions by results, test tubes
finding a pathway which has lower with distilled water
activation energy. and pepsin only
❖ Each enzyme has an active site where should have no
the chemical reactions occur. The digestion. Digestion
reactant of the said chemical reaction is will only occur in test
called substrate. tube of pepsin with
❖ CHARACTERISTICS HCl
➢ They are not altered or
consumed during reaction.
➢ Reusable Enzymes show
NOTES: Distilled water does not contain an
specificity to the reaction they
enzyme, so no digestion was seen. In test
control.
tube 2, cloudiness can be seen which may
➢ Enzymes are sensitive to their
indicate precipitation but no hair like.
environment so they can be
controlled by adjusting the
temperature, the pH or the AMYLASE
substrate concentration. ❖ Salivary Amylase
(Ptyalin)
PEPSIN ❖ Present in saliva
❖ an enzyme that with optimum pH
hydrolyzes of 6.7-7.0
protein to ❖ An enzyme that
smaller hydrolyzes
peptides to α-1,4-glycosidic
amino acids, bonds in starch
cleaving peptide producing
bonds between oligosaccharides,
hydrophobic and aromatic amino acids. maltose or glucose
❖ It is active in acidic environment, depending on the time spent in the
having an optimum pH of 1.5-2.0 mouth of the food.
 ❖ Iodine Test ❖ Potato catalase - enzyme found in
➢ Iodine reacts with starch potato, optimum pH = 9.0
forming a blue-black solution
RESULTS:
RESULTS

STARCH SAMPLE +
SAMPLES
IODINE
B1 = 1% Starch Blue-black color
without saliva (positive for I2 test)
Brown to Purple
B2 = 1% Starch with
color
Saliva
(negative for I2 test)
SAMPLES Observations
C1 = Potato with acetic acid
Very small
(pH = 2.4) (rate 3)
Experimental and C2 = Potato with sodium
theoretical results carbonate (pH = 11.6) (rate Medium
coincide with each 2)
other.
Large volume of
C3 = Potato only (rate 1)
bubbles

Erratum: Sodium carbonate (pH=12) was used

in the experiment NOT sodium bicarbonate
(pH=9)
NOTES: The test tube with saliva shows
digestion, there is breakdown of starch.
Theoretical and experimental results are the
While test tube without saliva showed same.
presence of starch, meaning starch is not
digested.
NOTES: Catalase denatures are pH 2.4 and
11.6 that’s why bubbles are lesser compared
CATALASE to the last graduated cylinder.
❖ An enzyme
that protects
organisms EXPERIMENT 4: FACTORS
from the AFFECTING ENZYME ACTIVITY
effects of MC-2 LAB
hydrogen
peroxide. Objectives of the Experiment
❖ H2O2 is a
powerful ❖ Describe the effect of the factors
oxidizing agent and is potentially presented in the experiment on
damaging to cells. enzyme’s activity.
❖ It breaks down hydrogen peroxide into ❖ Relate the factors to human enzymatic
water and oxygen gas. metabolic activities.
➢ 2H2O2 → 2H2O + O2
ENZYMATIC ACTION EXPEREMENTAL RESULTS
❖ Catalysis of enzymes can increase the Theoretical and experimental results coincide.
reaction rate many million times faster
than without a catalyst.
❖ Their action on the substrate can be NOTES: Hot
temperature
controlled by adjusting the temperature,
denatures enzymes
pH, or substrate or enzyme’s while cold
concentration. temperatures not
❖ Further, since they are proteins, they are necessarily denature
also affected by agents that causes the enzyme but can
them to undergo denaturation. This lower its kinetic
experiment shows the effect of these energy and can slow
down the rate.
factors on enzymatic reactions.

EFFECT OF TEMPERATURE
EFFECT OF PH
❖ The effect of pH depends on the
❖ Temperature
enzyme’s optimum pH (pH where
increases
enzyme is most active). But extreme
enzyme activity
pH levels will cause denaturation.
up to its
❖ If pH slightly different from the enzyme’s
optimum
optimum value, there will be small
temperature.
changes in the charges of the enzyme
❖ Optimum
and its substrate’s molecule.
temperature is
❖ This change in ionization will affect the
the temperature
binding of the substrate with the active
at which
site.
enzymatic
❖ Salivary amylase has an optimum pH
reaction occur
of 6.8
fastest.
❖ For most enzymes, the optimum
temperature is about 40°C.

RESULTS

THEORETICAL RESULTS

HEIGHT OF
SAMPLES
BUBBLES
A1 = Potato in room Highest amount of
temp bubbles
A2 = Potato in cold Medium amount of
temp bubbles
RESULTS
A3 = Potato in hot Experimental results of acid and base solution
No bubbles
temp did not match the theoretical results
 ❖ Increasing enzyme
THEORETICAL RESULTS
concentration will
SAMPLES Observations increase rate of
reaction but when
B1 = Starch with
acetic acid (pH = Bluish solution enzyme is no
2.4) longer available, it
reaches a
B2 = Starch with maximum point
sodium carbonate Bluish solution
(bottom image).
(pH = 11.6)
B3 = Starch in pH Yellow coloration of
buffer 7 solution RESULTS

EXPERIMENTAL RESULTS
THEORETICAL RESULTS

HEIGHT OF
SAMPLES
BUBBLES
C1 = 5 grams potato + 5ml
Highest
H2O2
C2 = 3 grams potato + 5ml
Medium
H2O2
C3 = 1 gram potato + 5ml
Low
H2O2

Theoretical and experimental results coincide.

Experimental Results
Possible error: Buffers should be used instead
acid since acid hydrolyzes starch and also
sodium carbonate will react with iodine forming
iodide ion solution which is colorless. B4 is an
evidence that acids have reactions with starch.
B4 is not heated with saliva.

EFFECT OF ENZYMES CONCENTRATION

❖ When enzyme
concentration
is continuously
increasing, as
shown in the
Est 27-6 = 19-4ml = 13-2ml =
top image, the 21ml 15ml 11ml
reaction rate
will continue to
increase.
EFFECT OF SUBSTRATE CONCENTRATION Experimental Results

❖ The rate of reaction increases as

substrate concentration increases (at
constant enzyme concentration).

18 -12 = 6ml 15 – 8 = 7ml 18 -12 = 6ml

❖ The increase in velocity is proportional

to the substrate concentration but when
enzyme is no longer available, V max is
reached – saturation point.

RESULTS

THEORETICAL RESULTS

HEIGHT OF
SAMPLES
BUBBLES
D1 = 3 grams potato + 10ml
Equal
H2O2
D2 = 3 grams potato + 5ml
Equal
H2O2
D3 = 3 gram potato + 2ml
Equal
H2O2

Theoretical and experimental results coincide.