Laboratory Exercise # 7

Influence of pH and Temperature on Enzyme Activity

WORKSHEET

Name: Almira Carla S. Prologo Date: 7-19-19

Year and Section: BSMT2-C Group No: 7

OBSERVATIONS
Table 1. Influence of pH

A. Without liver puree

Flask No. pH Gas Remarks
Formation
1 3 0 No bubbles
2 6 1 Few bubbles
3 7 1 Few bubbles
4 10 0 No bubbles
5 11 0 No bubbles

B. With liver puree

Flask No. Gas Formation Remarks

1 0 No bubbles
2 1 Few bubbles
3 3 Many bubbles
4 0 No bubbles
5 0 No bubbles

Table 2. Influence of Temperature

Flask No. Temperature Gas Formation Remarks

1 10 degree 1 There was only few bubbles formation in
celsius the solution
2 70 degree 0 No formation of bubbles was seen or
Celsius there is no gas formation
3 23degree 3 There was a gas formation in the
celsius solution and bubbles formed.
DISCUSSION:

EFFECT ON THE PH ON ENZYME ACTIVITY

 Explain the relationship of pH and temperature to the enzymatic reaction based on your observations.

The liver uses specialized enzymes to help it break

down toxic substances and make them safer for the body to
process. But an enzyme, just like the chemical reactions it
modifies, needs certain conditions to do its work. So, some
environments can make a liver enzyme effective, whereas
others can prevent it from working at all. Catalase is an
enzyme in the liver that breaks down harmful hydrogen
peroxide into oxygen and water. When this reaction occurs,
oxygen gas bubbles escape and create foam (Science
Buddies, 2012). Based on our activity the as a drop of
hydrogen peroxide was poured on the liver , it formed
bubbles indicating that catalase in liver breaks down the
Figure 1: shows the reaction of
hydrogen peroxide which is why there is a gas formation
liver with hydrogen peroxide.
present ( see figure 1)

Figure 2: shows flasks labelled from 1 to 5, with solutions on each flask:

Flask 1: 10 drops of H20 + 5ml HCL
Flask 2: 10 drops of HCL + 5ml H2O
Flask 3: 5.5 ml H20
Flask 4: 10 drops of NaOH + 5ml H20
Flask 5: 10 drops of H20 + 5ml NaOH
OBTAINING PH

WITHOU LIVER PUREE

Flask 1 Flask 2

Figure 3: shows the pH of flask 1 (pH = 3) Figure 4: shows the pH of flask 2 (pH = 6)

Flask 3 Flask 4

Figure 5: shows the pH of flask 3 (pH = 7) Figure 6: shows the pH of flask 4 (pH = 10)

Flask 5

Figure 7: shows the pH of flask 5 (pH = 11)

Adding of Hydrogen peroxide

Figure 8: shows no bubble Figure 9: shows few bubble Figure 10: shows few bubble
formation as hydrogen formation as hydrogen formation as hydrogen
peroxide was added peroxide was added peroxide was added

Figure 11: shows no bubble Figure 12: shows no bubble

formation as hydrogen formation as hydrogen
peroxide was added peroxide was added

WITH LIVER PUREE

Figure 13: shows no bubble Figure 14: shows few bubble Figure 14: shows many
formation as hydrogen formation as hydrogen bubble formation as hydrogen
peroxide was added to the peroxide was added to the peroxide was added to the
flask with liver puree flask with liver puree flask with liver puree
 Figure 15: shows no bubble Figure 16: shows no bubble
formation as hydrogen formation as hydrogen
peroxide was added to the peroxide was added to the
flask with liver puree flask with liver puree

According to Science and Evolution Blog (n.d.), every enzyme shows maximum activity at an optimum
pH. Their activity is slow above or below the optimum pH. Enzymes have active sites where the substrates
bind. These active sites are damaged or in other words their shape is changed by changing the pH. Substrates
no longer fit the active site and the reaction does not occur. A pH of about 7 is the optimum and as the pH
moves further away from the optimum pH the enzyme activity starts to slow down. Based on our
observations, on the test without liver puree the flask 1-5 have a pH of 3,6,7,10,11 correspondingly. There was
a gas formation on the bubbles formation on the flask with pH 6 and 7, same as the flask with liver puree. This
is because the enzyme activity is most active at pH 7 that is why reaction is visibly seen in flask with pH 7,
however in the flask 2 with pH 6 there is still enzymatic reaction happen, this might be due to the other factors
that cause the reaction of enzymes to happen. The flask 1, 4 and 5 with pH of 3, 10, and 11, there was no gas
formation happen or enzymatic reaction happen this is because at pH lower or higher than 7 enzymes are
denatured.

EFFECT OF TEMPERATURE ON ENZYME ACTIVITY

Figure 17: shows the three flask Figure 18: shows the flask a subject to
labelled accordingly cold water bath at 10 degree Celsius
 Figure 19: shows the flask B subject Figure 20: shows the flask C subject
to hot water bath at 70 degree to ambient temperature
Celsius

Figure 11: shows the solutions of

each flask added with liver puree

According to Science and Evolution Blog (n.d.), enzyme shows highest activity at a specific temperature
which is called the optimum temperature. The rate of reaction above or below this temperature is slower.
Increasing the temperature causes the frequency of collisions to increase as the molecules move faster when
the heat is supplied. For a 10 degree Celsius rise in the temperature, the activity of enzyme doubles until the
optimum temperature is reached. Increasing further temperature damages the active sites and changes their
shapes. Substrates can no longer fit into the active sites. At temperatures higher than 60 degrees Celsius
enzymes are basically completely denatured. Based on our activity, at 10 degree Celsius there was a few
bubbles formed or gas formation where the solution becomes turbid color, this means that in this
temperature the rate of reaction is increasing until it reached the optimum temperature. While in the 70
degree Celsius temperature, there was no bubbles formed or gas formation happened and there is a formed
substance in the mixture because greater than 60 degree causes the enzymes denaturation. In the ambient
temperature, which is 23 degree Celsius, the solution turned turbid in color an there was many bubbles
formed this means that the enzymes have reached the optimum pH or the enzymes is almost at the optimum
pH.
GUIDE QUESTIONS:

1. At what pH is the enzyme catalase most active? Do you have any idea why is this so?

Based on the Libre text article (2016), the single most important property of enzymes is the ability to
increase the rates of reactions occurring in living organisms, a property known as catalytic activity. Because
most enzymes are proteins, their activity is affected by factors that disrupt protein structure, as well as by
factors that affect catalysts in general. Factors that disrupt protein structure include temperature and pH;
factors that affect catalysts in general include reactant or substrate concentration and catalyst or enzyme
concentration. At the optimum pH is the enzyme catalase most active. This is because enzymes are affected by
changes in pH. The most favorable pH value - the point where the enzyme is most active - is known as the
optimum pH. Extremely high or low pH values generally result in complete loss of activity for most enzymes.
PH is also a factor in the stability of enzymes. As with activity, for each enzyme there is also a region of pH
optimal stability (Worthington Biochemical Corporation, 2019).

2. What is the effect of increasing temperature on enzyme activity? What about the increasing pH?

Temperature affects the reaction rate of enzymes, as do pH, substrate concentration and enzyme
concentration. At low temperatures, enzymes have low activity. As the temperature rises the rate of reaction
increases, usually 2-fold for every 10 degree Celsius rise. According to Brilliant Biology Student article (n.d.),
the activity peaks at a specific temperature unique to the enzyme. This is known as the optimum temperature -
the temperature at which an enzyme is maximally active. Beyond the optimum temperature the activity of the
enzyme decreases. At extreme temperatures, the enzymes are denatured and activity ceases.

CONCLUSIONS:

Biochemical reactions are necessary for growth, repairing damaged tissues, and obtaining energy and they
take place in all living organisms’ bodies. These reactions are called ‘metabolism’ and they happen all the time
in living organisms. If they stop working, this leads to the death of the organism. Enzymes are biological
catalysts made up of large protein molecules. They speed up the chemical reactions inside the cell. The enzyme
is made up of a combination of amino acids which for a chain of polypeptides between each other.

REFERENCES:

Brilliant Biology Student article (n.d.).Effects of Temperature on Enzyme Action. Retrieved from http://
brilliantbiologystudent.weebly.com/effect-of-temperature.html on July 27 2019

Science Buddies. (2012).The Liver: Helping Enzymes Help You. Retrieved from https://www.scientific
american.com/article/bring-science-home-liver-helping-enzymes/ on July 27 2019

Science and Evolution Blog. (n.d.).Effect of Temperature and pH on enzyme activity. Retrieved from
https://www.drupalbycity.com/effect-of-temperature-and-ph-on-enzyme-activity/ on July 27 2019

Worthington Biochemical Corporation, (2019).Introduction to Enzymes. Retrieved from http://www.worthing

ton-biochem.com/introbiochem/effectspH.html on July 27 2019