LAB REPORT: CATALASE EXPERIMENT
Chamodi Basnayake
PURPOSE: The activity of an enzyme is largely affected by environmental factor (like how the
secondary, tertiary and quaternary structures of proteins are affected). Enzymes work their best at
an optimal condition as they favor the most active shape for the enzyme molecule. The reason this
experiment was conducted was so the optimum pH of the potato can be found. By finding the
optimum pH, the pH in which the enzymes within the potato work the best is found.
HYPOTHESIS:
Substrate concentration- The rate of chemical reactions increases if the substrate concentration
increases. Although, it’s initially responsive, at a certain concentration level of substrate, the rate of
reactions reaches a maximum and the rate wouldn’t get any higher no matter how much we
increase the concentration (the saturation point). By adding substrate, we increase the molecular
colliding. But once every enzyme is occupied, the rate won’t change.
Enzyme concentration- increasing enzyme concentration will increase the rate of reaction because
more enzymes will be colliding with substrate molecules. But it will only have an affect up to the
certain concentration (saturation point) where adding more enzymes wouldn’t make the rate any
higher because there aren’t enough substrate molecules. Too much enzymes and too less substrate
is no good.
Temperature – The optimum temperature is when the enzymes of the potato wok the best in a
certain temperature. Up to a certain point, the rate of enzyme reaction increases because the
substrate collides with the enzyme more when temperature is increased (they move a lot). But after
that certain temperature the reaction rate drops because temperature disrupts the bonds and
therefore the active sites denaturing it. I think potatoes grow in a little bit cold environment. So the
optimum temperature would be 23- 27 degrees Celsius.
PH - Optimum pH is when the enzymes in the potato work the best at a certain pH. Just like how the
temperature works, as the pH gets higher, the rate of enzyme reaction increases (enzyme
renatures)but once it goes beyond the optimal pH, the rate starts to drop. If we graph it, it should
look something like a bell shape. When it’s beyond the optimum point, the pH starts to denatures. I
think the optimum pH is between 6 – 8 pH, more likely 7 because potatoes taste more basic than
acidic and it grows in soil, it can’t be of any extremes.
EQUIPMENT:
* 3 potatoes
* 30% Hydrogen peroxide
* Sodium hydroxide
* Hydrochloric acid
* Filter paper
* Hole puncher
* Tweezers
* Timer
* Spatula
* Tap water
* pH probe
* blender
* 4 beakers
* Knife
PROCEDURE:
1. Cut the three potatoes into 3 pieces each with a knife
2. Measure and prepare 200mls of water on a beaker labeled
3. Blend the potatoes with 200mls of water in a blender
4. Pour the potato juice into a beaker while filtering it, making an enzyme solution of catalase
5. Scoop out the frost on top of the catalase with a spatula
6. Divide the catalase into two different beakers, and label one acid, another one base
7. Get another beaker, label it Hydrogen Peroxide, and pour in 20mls of Hydrogen Peroxide in the
beaker
8. Re-add the water in the labeled beaker
9. Use the pH probe to measure the pH of normal catalase solution, and then record it
10. Hole punch the filter paper with a hole puncher
11. Pick up one of the filter paper with tweezers and submerge it in the normal catalase solution and
time it for 6 seconds with a timer
12. Put the filter paper out and place the paper at the bottom of the beaker with the tweezers
13. Measure the amount of time it takes for the filter paper to float up to the surface of the solution
with a timer
14. Record the time it takes for the filter paper to float up to the surface of the solution
15. Repeat the procedures 11 to 14 three times
16. Wash the probe in the water prepared
17. Add in 1 drop of hydrochloric acid on the acid beaker
18. Measure the pH of the catalase, and drop acid until the pH becomes somewhere near pH 3-4,
keep on measuring with the probe after adding every drop
19. Record the pH
20. Repeat the procedures 11 to 14 three times
21. Add in another drop of hydrochloric acid on the acid beaker
22. Measure the pH of the catalase, and drop more acid until the pH becomes somewhere near pH1,
measuring with the probe after every drop
23. Record the pH
24. Repeat the procedures 11 to 14 three times
25. Wash the probe in the water prepared, and rinses it until the pH of the probe comes back to
neutral
26. Add in 1 drop of sodium hydroxide on the base beaker
27. Measure the pH of the catalase, and drop sodium hydroxide until the pH becomes somewhere
near pH 9-10, measuring with the probe after every drop
28. Record the pH
29. Repeat the procedures 11 to 14 three times
30. Add in another drop of sodium hydroxide on the base beaker
31. Measure the pH of the catalase, and drop more acid until the pH becomes somewhere near
pH14, measuring with the probe after every drop
32. Record the pH
33. Repeat the procedures 11 to 14 three times
34. Graph the results
RESULTS:
*Infinity = 1 minute and up
pH
First trial (sec)
1.7
4
6
9.7
12.83
Infinity
Infinity
4.41
Infinity
Infinity
Second trial
(sec)
Infinity
Infinity
5.9
Infinity
Infinity
Third trial (sec)
Average (sec)
Infinity
Infinity
4.00
Infinity
Infinity
Infinity
Infinity
4.77
Infinity
Infinity
ANALYSIS:
Uncertainty of Volume of the 100ml beaker
Substance / Equipment
Hydrogen Peroxide
pH probe
Uncertainty
± 5% for 100ml measured
Mathematically calculated for 20ml measured
 ± 1ml
± 0.01 pH
(for every pH measurement)
I think we chose pH values that are too much towards the extremes of the pH spectrum, which
denatured the enzymes completely, landing us with all results as infinity except for the optimum
temperature. Another big weakness was the timing. We used a stop watch also and we most
certainly weren’t accurate. The uncertainty of time can be the human reaction time which is 0.215 s
later than the original time. More error is added because sometimes we stopped the time before the
whole disc reaches the top because some of us thought it has fully surfaced while some of us
thought it hasn’t, so there was some confusion. To measure 20ml of hydrogen peroxide we used a
100ml beaker which was a huge error since measuring such a small value in a big beaker will
increase our error because for every 1 ml we measure wrong, the volume change is huge because
big beakers have a big surface area. During the measurement of pH using the probe there was a
problem. When we measured the pH of the same filtrate at different times, it came out as different
values at different times and we think that’s because we didn’t stir the filtrate enough after adding
acid or base into to it, so sometimes the pH of one area of the filtrate was taken. The froth could’ve
also affected how fast the filter paper disc emerges from the hydrogen peroxide solution. Another
thing is, after we submerge our disc in different beakers with different pH, we place that at the
bottom of the hydrogen peroxide solution. Each time we do this, we use the same hydrogen
peroxide solution and each time we do this, some of the filtrate absorbed by the disc will be left
behind in the hydrogen peroxide solution changing its pH too. So it would’ve affected how fast our
disc reached the surface too. The first time we attempted to make our base filtrate we ended up
adding too much that it reached the very end of the pH scale. So we were forced to make another
solution with a less basic solution and needed more filtrate. We didn’t have enough, only a little and
I think the filtrate we used had some starch in it too because we had to use the very last bit of
filtrate left and that would’ve made a big difference.
CONCLUSION:
The objective of the experiment was to find the optimum pH where potato enzymes will work the
best and the fastest. The pH of the filtrate of the potatoes had a pH of 6 and that is the optimum pH,
since that’s the natural pH and naturally they maintain the optimum pH conditions so everything can
work every well inside the potato. As it becomes too acidic or too basic, the enzymes start to change
shape, which gets us back to how environment changes secondary structure of proteins, which
change tertiary, which changes quaternary structure. So as the enzyme slowly denature, the active
site doesn’t fit the substrate that well, not an induced fit, so it works badly. As the pH is close to the
optimum pH, the enzymes re-nature and the substrate becomes a much more induced fit to the
active site and things work faster. Explanation of our experiment:
To find the optimum pH of the enzyme, which is catalase, we find how fast reactants disappear and
how fast the product appears (We wanted to break H2O2 to H2+O2.) We timed how fast the filtrate
disc came to the surface on the hydrogen peroxide solution. That’s how we measured how fast the
product appears because when hydrogen peroxide goes through chemical reaction with catalase,
oxygen was a product, which later bubbled up to the surface. By timing how fast the disc comes up,
we measure how fast oxygen is made in the reaction. The shorter time, the better catalase works.
When we got results like infinity, it shows that the enzyme has denatured so much that they don’t fit
the substrate anymore. According to our results, the optimum point is pH 6 which was the original
pH of the potato filtrate.
I hypothesized that the optimum pH for potato would be somewhere around 6-8 pH. My hypothesis
was somewhat correct since the optimum pH turned out to be 6 pH. I was expecting a pH 7 or any
value that is base than acid, so I guess I was correct.