EXPERIMENT2

UNIVERSITI TEKNOLOGI MARA
BIO 411
CELL BIOLOGY
PRACTICE 3
REDOX -REACTION/CATALASE ACTIVITY
GROUP MEMBERS: Syazani Zahier Bin Zakaria (2014292204)
Nur Fatin Syamimi Binti Ahmad Tarmize (2014236664)
Zuliakha Binti Zolkafali(2014279698)
Nur Izzati Binti Azizan(2014284578)
GROUP:ASB1Cb
DATE OF SUBMISSION :12 November 2014
INSTRUCTOR: Miss Ellia Kartini Mujar

OBJECTIVES
1. To demonstrate reduction-oxidation reaction on living tissue.
2. To compare enzyme catalase on plant and animal tissues.
3. To measure the rate on enzyme reaction.
4. To measure enzyme reaction.
INTRODUCTION
In this lab activity, we explore on the enzyme that is found in the cells of many living tissues. The enzyme is catalase. Catalase functions in speeding up reaction in our cells that break down hydrogen peroxide, a toxic substance into a harmless substance which is water and oxygen. Hydrogen peroxide (H2O2) is the by-product of many normal cellular respirations mainly in our body for it to function properly. However, H2O2 is a toxic substance that can be harmful for the body once it’s not processed properly. If the cell cannot break down H2O2 into a harmless substance such as water and hydrogen, we as the multicellular organism will be poisoned to death.
For experiment 3 the experiment is designed to measure the rate of reaction by measuring the amount of substrate remaining after some period of reaction time. H2O2 is the substrate that will convert to H2O2+ O2 over time. Hence, this will give us indirect measure of the substrate converted to product and the rate of reaction. The remaining substrate from the reaction is equal to the initial amount of the substrate minus the amount converted to product by the enzymatic action.
MATERIALS
3% hydrogen peroxide
Sulphuric acid (1M)
Potassium permanganate solution
Soaked beans,potatoes,apple and chicken meat
Razor blades
Lime juice
Test tubes and racks
Ice,beakers
Watch plates
Burettes
Water bath
Conical flask
METHODS
Experiment 1:Observing redox reaction
1. Three small pieces of potato and apple were cut and placed pair by pair into the 3 watch plates.
2. The first watch plate was left exposed to air, the second one was added with water while the third one was added with the lime juice.
3. After 45 minutes, the observation was compared and recorded for all the slices in the 3 watch glass. Experiment 2 :Comparing catalase activity in living tissues
1. 2mL of hydrogen peroxide solution was placed into four clean test tubes.
2. Slices of potato,apple,chicken meat and 3 pieces of beans were prepared.
3. Each of the test tubes were then added with the prepared slices and the 3 beans respectively for test tubes 1, 2, 3 and 4.
4. The reaction that occurs once the ingredients react with the hydrogen peroxide was observed and graded based on the amount of bubbles produced, from the scale 0-5 with 0 as no reaction while 5 as very fast reaction.
5. The temperature of the test tubes was also felt using hands and was recorded.
Experiment 3 :Effect of Temperature on Catalase Activity
1. 4 test tubes were added with 3mL of hydrogen peroxide. The the test tubes was placed into different beaker to obtain different temperature of the hydrogen peroxide solution.
2. Test tube 1-4°C,Test tube 2-24°C,Test tube 3-34°C,Test tube 4-44°C respectively.
3. The test tubes were left for 20 minutes to obtain the temperatures.
4. After 20 minutes, each test tube was added with 6 pieces of beans. The reaction was left to occur for about 1 minute.
5. Then, 3.5 mL of sulphuric acid was added into each test tube to stop the reaction immediately.
6. The content of the test tubes were poured into conical flask and each one of the test tubes content was titrated with KMnO4 until the mixture turns pink.
7. The amount of KMnO4 used was observed and recorded.
RESULTS
TEST TUBE
1
2
3

NO. OF SLICED ITEM
2
2
2

CHANGE OF COLOUR
Brown
Brown
Brown

TIMING
45 minutes
45 minutes
45 minutes

RESULT
Changed to brown early into the timing
Change to brown in 15 minutes from the timing
Change to brown 40 minutes from the timing

Experiment 1:Observing redox reaction
Test tube
Type of cell in hydrogen peroxide (H2O2)
Grade of reaction (0-5)
Presence of heat due to the reaction
1
Small piece of potato
1
No
2
Small piece of apple
2
No
3
Small chicken piece
4
No
4
3 pieces of soaked peas
5
No
Experiment 2 :Comparing catalase activity in living tissues

Experiment 3 :Effect of Temperature on Catalase Activity

DISCUSSION
From this experiment 1, in the first test tube, the color of potato and apple is early turn to brown because of the exposure to air that causes the oxidation. The oxidation thus turns the slices of item (potatoes and apples) to turn brownish. Next, the potato and apple that put together with water oxidized slightly slower compared to the ones exposed to air. This is because the water consists of oxygen and it can cause oxidation yet provide some layer of protection from oxidation caused by the air. In the last test tube, the potato and apple turn into brown color later because the slices of item immersed in lime juice. The lime juice has low in pH value and the rate of the fruit to oxidize is also low.
Based on experiment two, we found that the peas produced a large amount of bubbles (scale 5) which meant that the reaction between the peas and hydrogen peroxide (H2O2) was very fast. The second cell that released such lot amount of bubbles was the chicken (animal) with the scale of four, followed by potato and the least amount of bubbles was the apple. The peas react very fast as it have higher amount of enzyme catalase among those four other cells. When cells react with the hydrogen peroxide, water and oxygen gas would be released. This shown in the equation ; 2H2O2 2H2O + O2. Thus, catalase reacts more with the small size of plant cells than the animal cell.
In experiment 3, the effect of temperature on catalase activity was studied. Enzymes are globular proteins, which are polymers of amino acids. Enzyme functions to catalyze or speed up chemical reactions. Enzyme increase reactions rates by a factor of about 1 million by lowering the activation energy ( Morgulis S. et al, 1926). All enzyme are specific to single chemical reactions. For example, in this experiment, enzyme catalase. Catalase is one of the most important catalyst. Catalase catalyses conversion of hydrogen peroxide, a potentially harmful oxiding agent to water and oxygen. Apart from that, catalase also uses hydrogen peroxide oxidise toxins including phenols, formic acid, formaldehyde and alcohols ( Ester H. C., 1950 ). Like all ezymes, catalase reactions are affected by temperature and pH. Under a low temperature, based on the experiment, at 4 °C, the rate of catalase is very low. According to collision theory, for a reaction to occur, particles must collide with sufficient energy to overcome activation energy and with correct geometric orientation. At 4 °C, the kinetic energy of the subtrate(H2O2) and catalase are slow makes the frequency of collision decrease hence decrease the number of effective collision. At temperature of 34 °C, the rate of catalase activity is the highest. This indicated that at this temperature, the enzyme reacts under the optimum condition where the kinetic energy needed for effective collision is at the best. However, at temperature of 44 °C, the enzyme starts to denatured since it is a protein. At this temperature, the structure of the protein has changed and the substrate can no longer attach to the enzyme at the correct geometric orientation ( Eyster H. C., 1950). Hence, the rate of catalase activity decreasing.Potassium permanganate ( KMnO4 ), a strong oxidizing agent used in this experiment is to determine the activity of the enzyme. The function of potassium permanganate is the same as the enzyme catalase, but potassium permanganate is a chemical catalyst, not biological catalyst (www.xula.edu, accessed on 12 Nov 2014). The amount of titrated potassium permanganate indicated the amount of hydrogen peroxide (subtrate) left in the mixture. Hence, the amount of of subtrate remaining in the mixture is inversely proportional to the rate of catalase activity (www.xula.edu, accessed on 12 Nov 2014). Sulphuric acid (H2SO4) used in this experiment is stop the reaction between the substrate and enzyme catalase. Alike temperature, sulphuric acid is a chemical which can destroy the protein, enzyme catalase by increasing the pH of the mixture. By adding sulphuric acid, the enzymatic activity can be stop ( www.xula.edu, accessed on 12 Nov 2014).
CONCLUSION
The reduction-oxidation reaction on living tissue can be demonstrated by the colour changes due to the reaction on the sliced apples and potatoes. The enzyme catalase on plant are more compred to animal tissues since the beans give out more bubbles and reaction compared to the chicken meat.Enzyme reaction is affected by temperature and pH of the enviroment. Enzyme needs optimum temperature and optimum pH to efficiently functional.
REFERENCES
http://www.xula.edu/chemistry/documents/biolab/Carroll%20lab%20Chap%208.pdf ( accessed on 12 Nov 2014)
Eyster H. C. (1950). Effect of The Temperature On Catalase Activity. The Ohio Journal of Science, v50 n6, 273-277.
Morgulis S., Beber M., Rabkin I. (1926). Studies On The Effect Of The Temperature On The Catalase Reaction Journal of Biology Chemistry, 68:521-533.

References: http://www.xula.edu/chemistry/documents/biolab/Carroll%20lab%20Chap%208.pdf ( accessed on 12 Nov 2014) Eyster H. C. (1950). Effect of The Temperature On Catalase Activity. The Ohio Journal of Science, v50 n6, 273-277. Morgulis S., Beber M., Rabkin I. (1926). Studies On The Effect Of The Temperature On The Catalase Reaction Journal of Biology Chemistry, 68:521-533.