Elena M, Audrey M, and Duaa K
Question
How do abiotic or biotic factors influence the rates of enzymatic reactions (chemical reactions that are assisted by enzymes)?
Background
Enzymes speed up chemical reactions by lowering activation energy (that is, the energy
needed for a reaction to begin). In every chemical reaction, the starting materials (the
substrate(s) in the case of enzymes) can take many different paths to forming products.
For each path, there is an intermediate or transitional product between reactants and
final products. The energy needed to start a reaction is the energy required to form that
transitional product. Enzymes make it easier for substrates to reach that transitional
state. The easier it is to reach that state, the less energy the reaction needs.
Enzymes are biological catalysts. They are large protein molecules, folded so that they
have very specifically shaped substrate binding sites. These binding sites make substrates
go into the transition state. To catalyze the reaction, several regions of the binding site
must be precisely positioned around the substrate molecules. Any change in the shape of
the overall folded enzyme molecule can change the shape of the binding site.
The optimum reaction conditions are different for each enzyme. The correct
environmental conditions, proper substrates, and, often, particular cofactors associated
with an enzyme are needed. In some instances, the optimum conditions can be deduced
fairly accurately based on the following:
- The organism from which the enzyme is derived
- The part of the organism in which the enzyme functions
- The environmental conditions in which that organism lives
Take the example of lactase, an enzyme that catabolizes (breaks down) the disaccharide sugar lactose into two monosaccharides, glucose and galactose. In humans, lactase is found mostly in the small intestine, where the pH is around 7. It would be reasonable to hypothesize that human lactase is optimally active at pH 7 and at 37°C (normal human core body temperature in degrees celsius). Free-living decomposer fungi in soil also produce lactase. However, soil pH usually is between 5 and 6.5. As could be predicted, the purified enzyme from a common soil fungus has a pH optimum of 5.5. The main enzyme for this lab, peroxidase, is found in many different forms, with optimum pHs ranging from 4 to 11 depending on the source and optimum temperatures varying from 10 to 70°C.
Purpose
In this experiment you will investigate the effect of environmental factors on the enzyme hydrogen peroxidase. This enzyme is found in all aerobic (using oxygen) cells and functions to decompose hydrogen peroxide into O2(g) and H2O. The specific environmental factors you will test (as a class) are temperature, pH, substrate concentration, and enzyme concentration. Your team will select one of these factors (variables) to test and report on.
Materials
- Digital balance (scale)
- Mortar and pestle
- Distilled water
- 3 100-liter glass or plastic beakers
- 1 mL or 5 mL syringe
- Hydrogen peroxide
- 1 Paper towel square (for filtration)
- Glass test tubes
- Test tube rack or holder
- Small plastic ruler
- Safety glasses
Depending on which environmental factor you choose to investigate, some of the following items will be needed for your experiment:
- Acid solutions with pH values between 2 and 6
- Alkaline solutions with a pH between 8 and 12
- pH test strips
- Ice
- Large plastic beaker (for ice bath)
- Hot water
- Large plastic beaker (for hot water bath)
- Thermometers
Procedure
This part will be determined by the students conducting the experiment. For an overview of the general scientific experimentation and research process, see the flow diagram below. Use the worksheet that follows to write out in detail the hypothesis you are seeking to test, the materials you will use, and the steps you will follow to conduct your experiment.
Enzyme Lab Worksheet
Hypothesis: If changing the temperature level affects how the enzymes function, then increasing the temperature of the ripgut brome grass juice and peroxide will speed up the reaction, while decreasing the temperature will slow the reaction compared to the room temperature control.
Independent Variable: The temperature
Dependent Variable: The height of the foam in cm.
Controlled Variables: Grass juice and water.
Justification of hypothesis: We think this because when the enzymes come into contact will different temperatures then the enzymes will either speed up or slow down depending on the water temperature.
Why did you choose this as your hypothesis?
We wanted to observe the overall reaction of the temperature differences that would either speed up or slow down the process of the catalyst.
Materials (Your Team’s Experiment):
- 8.62 ml grams of grass
- 30 milliliters of water
- Test tubes
- Test rack
- Paper towels
- Guacamole thing (mortar and pestle)
- Syringe
- Thermometer
- Ruler
Procedure: Our group will be mixing a certain amount of distilled water and grass to make grass juice. We will then add different temperature mixtures to see whether the reaction goes faster or slower, and what the reaction is.
Summary:
For our experiment we put the grass juice in the water and overall concluded that the temperature does affect the way the enzymes work. We know this because during our experiment we tested three different temperatures of water. This data showed that the room temperature water went the slowest in terms of how fast the bubbles appeared. While the cold slowed down our reaction time of the bubbles compared to the heated peroxide. Lastly, the fastest was the heated water which dramatically speed up the reaction time of the bubbles.
Detailed Steps:
We started our experiment by grinding up 8.62 grams of grass and in total added 30 milliliters of water to the grass. By mashing the two together in a mortar and pestle it formed a juice substance as (we referred to it as grass juice). Next we lined up 9 test tubes on a rack, having the room temperature, cold, and heated water each grouped in one section by temperature. After we put approximately 2 milliliters of grass juice and 1 milliliter of hydrogen peroxide into each test tube. As we put the hydrogen peroxide in the cold temperature test tubes we watched the reaction. At 0 degrees celsius, 30 seconds in the foam/bubbles in the test tube had grown 0.2 centimeters. After 60 seconds at 0 degrees celsius the bubbles/foam moved up 0.8 centimeters, and after 90 seconds 1 centimeter. Lastly after 120 seconds (2minutes) the foam/bubbles came up to 1.5 centimeters. Next we tried the room temperature water (24 degrees Celsius). As we out the hydrogen peroxide into the test tube we began to watch the bubbles/foam. We observed the reaction from the room temperature test tubes and it showed that after 30 seconds the foam/bubbles was at 0.3 centimeters. Next after 60 seconds it was grown 0.5 centimeters, and after 90 seconds it had moved 1 centimeter. Finally after 120 seconds (2 minutes) it had moved to 1.5 centimeters. The last test we did was with the heated water; the temperature from that was 40 degrees celsius. For our first test after dumping the peroxide in we turned on the watch and watched it move up 0.5 centimeters. Soon 60 seconds passed and we watch is go up 1 centimeter, and after 90 seconds it had gone up 2 centimeters. In closing at 120 seconds (2 minutes) it had gone up to 2.3 centimeters. After we had gotten all of our data from the experiment we washed out all of the materials we had used for the lab.
Data and Results:
(This section should include at least one graph!)
https://drive.google.com/drive/folders/1V4TXAwrsEv1NV1UG2E-fR3BU6lnqodcD
Conclusions:
Overall, the heated peroxide mixed with the grass juice created foam faster rather than the room temperature peroxide and the cold peroxide. The cold peroxide mixed with the grass juice had a relatively same reaction rate as the room temperature, but the room temperature succeeded the colder peroxide after 2 minutes, so we can conclude that the cooler the temperature the slower the reaction will be.
To start, the heated peroxide was to reach 7 cm, which was the tallest amount of foam. The heated hydrogen peroxide mixed with the grass juice had increased an average of 0.575 cm per 30 seconds. The heated hydrogen peroxide sped up the reaction, we can tell this from the fact that more foam was able to form faster over the given amount of time.
The room temperature experiment ( our control ) reached a maximum foam height of 5 cm and had an average reaction rate of 0.56 cm per 30 seconds.
The cold hydrogen peroxide reached up to 4 cm. The average rate per 30 seconds for the cooled peroxide mixed with the grass juice was 0.4 cm per 30 seconds. The cooled hydrogen peroxide slowed down the reaction, we can tell this because the average rate of reaction is slower than the room temperature and the greatest height of the reaction is lower.
In closure, our experiment data has shown that when water is heated, cold, or just at room temperature, the temperature has an affect on how the enzymes are performing their job and how fast and how much of a reaction will occur. When increasing the temperature the enzyme, reaction rate increases. When decreasing the temperature the enzyme, reaction rate will mildly decreases. We know that if we were to increase the temperature too much the enzyme will be denatured, but in this experiment we know that the enzyme was not denatured at 40 degrees celsius, but would most likely become denatured if we were to increase the temperature any more.
