Effect of Temperature on Enzyme Activity | Experiment

Rationale:

Enzymes are proteins that greatly speed up the rate of reaction and lower the activation energy required for the reaction to occur (Evans, B., Ladgies, P., McKenzie, J. and Spencely, M. 2004). Since they are proteins, enzymes can be denatured by an increase in heat. Though limited in rage of effectiveness, an increase in heat causes an increase in molecular motion, the reactants begin to move around more quickly and become more likely to collide into each other, causing a reaction (Kuipers, K., Leillor, P., Sharpe, P., Bloomfield, C. and Silvester, H. 2019). The temperature that causes the most activity is called the optimum temperature (Bliss, C., Fesuk, S. and Jacobs, J.). If the temperature is decreased and increased, the efficiency of the enzyme decreases and there is a decrease in reaction rate. This is because the temperature has the ability to change the structure of enzymes and make them become denatured or inactivated (Borger, P., Grant, P., Munro, L. and Wright, J. 2019).

The enzyme rennin if found in the gastric juice of young mammals and has the ability to react with milk proteins. It can turn the protein caseinogen found in milk, into clumps of insoluble curds of casein. This process brings out the proteins of the milk so that they are more readily available to the digestive system. Rennin performs best at the body temperature of 37^oC.

Junket is a dessert that is based off this enzymatic reaction and is composed of the commercially available substance rennet.

Research question:

Does increasing the amount of junket solution applied, produce more defined results in showing that the milk at a higher temperature will allow the rennin to perform more efficiently because the heightened temperature will cause the enzymes to increase their activity?

Original experiment:

The methodology used was Experiment 7.2A: The effects of temperature on the rate of reaction of an enzyme, which was composed and developed by Oxford University Press Australia and New Zealand.

The original experiment had milk set to three different temperatures; Less than 15^oC, 37^oC and 60^oC. Each had a set time of 10 minutes to allow the milk to properly adjust in temperature. 100mL of junket solution was made up with the concentration of one junket tablet. 6 drops of the junket solution were then applied to the three different temperature beakers of milk. Each sample was left for 10 minutes in the correct temperature to allow the reaction to take place and then were taken out and the results were recorded. Each sample was examined and its qualitative properties were recorded on whether they were liquid, solid or a state in between.

Modifications to the experiment:

To ensure that accurate and reliable data was collected, the methodology was much the same as the original with only a single change made. The change was instead of applying 6 drops of junket solution to the milk samples, 9 drops would be applied. This change to the original experiment was made in the pursuit to produce more defined results in showing how the enzyme rennin performs in correlation to temperature. In order to determine whether a higher temperature will allow the rennin to perform more efficiently, more precise results were needed. To produce these needed results, the amount of junket solution applied to the milk was increased with the expectation that some of the milk samples would set and prove the hypothesis that a higher temperature will allow the rennin to perform more efficiently.

Safety considerations:

Safety and some of the risks were taken into consideration by completing a full risk assessment

(see Appendix #1)

Processed Data:

The data recorded was collected as qualitative data. Each sample was put into a table and given a number to represent their state and degree of setting:

1. liquid
2. lumpy liquid
3. Very softy set, does not maintain shape when gently wobbled
4. Maintains shape when gently wobbled
5. Maintains shape when vigorously wobbled

The milk samples were poured from the test tubes into beakers and the degree of setting was recorded. The first experiment where 6 drops of junket solution are applied was repeated three times and the results are shown in the table below:

Every sample had no change in state and all remained liquids. The experiment that was modified to have 9 drops of junket solution applied was also repeated three times. The results are shown in the table below:

The concept that increasing the number of drops of junket solution applied would provide more definitive and defined results was in no way supported by this data. By increasing the number of drops, instead of a more defined result, the results were much more obscure and vague making is even more difficult to evaluate whether rennin performs more efficiently in higher temperatures.

By increasing the amount of junket solution applied the results were in no way improved, thus meaning the best data collected to make an evaluation on the correlation of temperature and the efficiency of enzymes is the data from the original experiment.

Going off the data collected from the original experiment, the evaluation can be made that the optimum temperature for rennin is not higher temperatures such as 60^oC but is closer to 37^oC. The data showed that cold temperatures cause the rennin enzymes to inactivate as the samples that were <15^oC showed no degrees of setting. The data also showed that higher temperature causes the enzymes to denature as the samples that were 60^oC showed inconsistent degrees of setting. Finally, it was found that normal or warm temperature is the optimum temperature for enzymes as the most degrees of setting occurred at this temperature.

Drawing these conclusions from the results from the original experiment is limited by the fact that the experiment did not provide definitive results. With the range of setting being so finite that the samples that did set, only had minuscule lumps in them. This makes the evaluation that warm temperature is the optimum temperature for rennin an imprecise and uncertain.

Evaluation:

The modified experiment possessed a main problem that caused limitations to the experiment as a whole which in turn produced results that were limited in their communication of enzyme efficiency.

The main source of limitation was found in the lack of range in the number of drops of junket solution applied to the milk samples. The application of only 6 and 9 drops created a very limited array of tests and provided even more so results. Instead of only changing the number of drops to a set number, the number should have been changed to an array to provide more extensive samples and results.

Sources of error:

Both the original and the modified experiment exhibited a number of sources of error that could have changed the reliability and validity of the results collected.

One source of error was found in the use of the water baths and the management of their temperature. Each water bath was set to a temperature and was required to maintain that temperature throughout the whole experiment. It was found that by placing such a large number of milk samples into the water baths, they fluctuated in temperature. This could have caused an indifference in the data collected due to the changes in temperature and them not being what they were meant to be consistently set too.

Another source of error was found in the creation and application of the junket solution. Once the junket tablet was placed into the beaker of 100mL of water, it never fully dissolved. It was found that the junket only partially dissolved and the rest created a cloudy mixture till it settled at the bottom of the beaker. This is a large source of error as the concentration of the junket solution applied to the milk samples would have been inconsistent and produced inconsistent results.

Suggested improvements & extensions:

The experiment had a number of problems and areas that could be modified to produce more reliable and commutative results.

The main change that should take place is the issue raised in the evaluation concerning the number of drops and the lack of array. An effective way to improve the experiment would be instead of changing the number of drops of junket solution from 6 to 9, the number should be changed to 2,3,4,5,6,7,8,9 and 10 drops.

Another change that could be made is the order of which the milk samples are placed into the water bath. Instead of placing every milk sample into the water baths at the same time, they should be placed one after the other allowing 10 minutes each. This would lower the fluctuation in temperature produced by there presence in the mixture and would produce more consistent results.

Conclusion:

After modifying the original experiment, it was found that it did not produce more defined results, thus making an evaluation of enzyme efficiency in correlation to temperature more limited and inaccurate. It was found that by increasing the number of drops applied, the results were less defined and were less effective in communicating whether or not enzymes perform better at higher temperatures. Going of the data from the original experiment the evaluation can be made that the optimum temperature for rennin is not higher temperatures such as 60^oC but is closer to 37^oC or warm temperatures.

Appendix #1

References:

• Evans, B., Ladgies, P., McKenzie, J. and Spencely, M. (2004). Pearson Biology. 1st edn. Melbourne, Victoria: Malcom Parsons.
• Kuipers, K., Leillor, P., Sharpe,P., Bloomfield, C. and Silvester, H. (2019) Chemistry for Queensland Units 1 & 2. 1st edn. Australia: Oxford University Press.
• Bliss, C., Fesuk, S. and Jacobs, J. (2019). Pearson Biology 11 Queensland Student Book. 1st edn. Melbourne, victoria: Alicia Brown
• Borger, P., Grant, P., Munro, L. and Wright, J. (2019). Nelson Science Biology units 1 & 2. 1st edn. Australia: Rachel Ford