Hand in to your discussion leader a typed lab report. You and your group members can submit identical cover pages and data tables, but your introduction and discussion must be written by you, in your own words. (1) Cover page: including the title of the experiment (in this case use “Comparing cell contents of plants from different environments” ), your name, the date, your discussion leader’s name, and the number of your discussion section. Also include the names of all your partners and their discussion section TAs. (2) Introduction: State your hypothesis about whether the concentration of solutes inside the cells of plants from different environments should be the same or different. Explain briefly why your hypothesis makes sense to you. State the prediction you generated from this hypothesis and describe (in general terms) how you tested it. You don’t need to detail the methods (because they are already in the lab guide), but you do need to define an isotonic point (especially what you consider to be its relationship to the cell’s internal solute concentration) and explain how you used isotonic points to test your prediction. (3) Results: On a separate page, summarize your data from tables 3 (onion cells) and 4 (three other kinds of cells) into one table, clearly labeled. (4) Discussion: In about 2 pages, explain what an isotonic point is and compare the isotonic points of your specimens with each other and with sea water. Does there appear to be a relationship between isotonic points and environment in which the plants are found? If not, then what cellular mechanisms (salt pumps, impermeable cell membranes, or others?) might these plant cells be using to maintain a stable internal environment despite widely differing external environments? Include some of the analysis and discussion questions raised on the previous page.

P.s. The first document is a model of the report (Pg 26-43 in the lab manual). Just write as it

Those pictures are the results of the report

The last document – the lab manual Page 44-60 are for this report we are working on, just ignore other pages.

All the information is in the lab manual.

Effects of environmental factors on the rate of enzyme catalyzed reactions

Guangyu Qi & Xiaoxi Yue

Professor Sahil Wadhwa

22th February 2019

· Introduction

In this report, we will mainly talk about what affect the rate of enzyme catalyzed reactions by discussing two experiments, that are experiment-effect of temperature on enzyme-catalyzed reactions and experiment-effect of PH on enzyme-catalyzed reactions. Why is enzyme so important that deserve us to do experiments on it? In living organisms, there is a class of substances that promote metabolism and promote the smooth progress of all life-related chemical reaction, which is enzyme. There is a very large family of enzymes. There are about 2,000 kinds of enzymes known at present, and there are more than 700 kinds in the human body. They are spread all over the human mouth, gastrointestinal tract, pancreas, liver, muscles and skin. In a word, we can’t live without the using of enzymes. Therefore, it’s necessary for us to figure out what factors affect such important enzymes, and how our enzymes affected by those factors.

Moreover, we will use a machine called Spectrophotometer (Spec 20) in two experiments. The reason why we use Spec 20 to compare the rate of the reaction under different conditions is that the Spec 20 is a device that measures how “dark” a liquid is, as more and more (clear) catechol is converted to (brown) benzoquinone, less and less light will be able to pass through the test tube, and the percentage of light absorbed will increase. Therefore, Spec 20 is the best choice for these two experiments.

Hypothesis:

As a substance produced by a living organism and acts as a catalyst to bring about a specific biochemical reaction. In the experiment-effect of temperature on enzyme-catalyzed reactions, we hypothesized that enzymes are most efficient at appropriate temperature like 30°C and 40°C. For example, in our daily lives, when our body have high fever or low fever, we will feel extreme discomfort; thus, we hypothesized high temperature or low temperature will affect the efficiency of enzymes.

In the experiment-effect of PH on enzyme-catalyzed reactions, we hypothesized that the rate of an enzyme catalyzed reactions is greatest at pH values that are 7. For instance, the most indispensable and needed substance in our body is water, which is typical neutral liquid in nature; thus, we hypothesized acid and base condition will affect the efficiency of enzymes.

· Material and Method

Effect of temperature on enzyme-catalyzed reactions

Materials

1. 6 test tubes with a test tube rack,

2. a wax pencil,

3. a pipette,

4. 5 different water bathes (Ice 0, room temperature, 30,40,60) and

5. a spectrophotometer.

6. potato extract,

7. water

8. catechol.

Method

1. We use a wax pencil to mark the tubes with numbers 1 through 5 and “X” and leave the last tube as “blank”.

2. We measure 1 ml of potato extract (a rich source of the enzyme catechol and 4 ml of water into each of the 5 tubes with a pipette.

3. To make a “blank,” we put 1 ml potato extract and 6 ml of water into the sixth tube. And we cover all 6 tubes with parafilm, invert to mix, and stand the tubes in rack.

4. We separate 5 tubes into different 5 water bathes and take 5 minutes to make sure the temperature of the solution inside our test tubes have reached the temperature of its water bath.

5. We add 2 ml of catechol solution to each of the 5 sample tubes simultaneously in the same sequence, so that reaction times in the 5 samples will be comparable. We remove the tube from the water bath, remove the parafilm, add the catechol, put the parafilm back on, and invert tube to mix the contents. Return each tube to its bath for 5 minutes.

6. We use the blank to recalibrate the Spec 20.

7. Exactly 5 minutes after adding the catechol, remove each sample tube from its water bath, dry it, insert the tube into the sample holder of the Spec 20, and measure absorbance. Quickly repeat for the other 4 tubes, one at a time, in numerical order. Record these values.

Effects of pH on enzyme-catalyzed reactions

Materials

1. 6 test tubes with a test tube rack

2. a wax pencil

3. a pipette

4. spec 20

5. potato extract

6. water

7. catechol

8. distilled water

9. 5 different buffer solutions (PH 3, PH 5, PH 7, PH 9, PH 11).

Method

1. We use a wax pencil to mark the tubes with numbers 1 through 5 and “X” and leave the last tube as “blank”.

2. We measure 1 ml potato extract and 4 ml buffer for pH 3 into No.1 tube. Measure 1 ml potato extract and 4 ml buffer for pH 5 into No.2 tube. Measure 1 ml potato extract and 4 ml buffer for pH 7 into No.3 tube. Measure 1 ml potato extract and 4 ml buffer for pH 9 into No.4 tube. Measure 1 ml potato extract and 4 ml buffer for pH 11 into No.5 tube. Measure 1 ml potato extract and 4 ml buffer for pH 7, plus 2 ml distilled water into “Blank” tube.

3. Cover each tube with parafilm and invert to mix. Stand all 6 tubes in the test tube rack.

4. Add 2 ml of catechol to the 5 sample tubes, put the parafilm back on, and again invert the tube to mix the contents.

5. Use our “blank” to calibrate the Spec 20.

6. Allow the browning reaction to proceed for exactly 5 minutes. Then insert the sample tubes, one at a time in numerical order, into the Spec 20 and record the absorbances. Note any color changes in the test tubes.

· Results

1th Experiment-Effect of temperature on enzyme-catalyzed reactions

Through the first experiment-effect of temperature on enzyme-catalyzed reactions, we get temp (°C), absorbance and color changes of whole 5 sample with the use and compare of blank sample. No.1 sample is in 0 °C, the absorbance after whole experiment ends is 2.34 and it change to yellow, not original white. No.2 sample is in room temperature; the absorbance is 3.23 and it change to brown which is dark than yellow a little. No.3 sample is in 30°C, the absorbance is 3.48 and it change to dark-brown than normal brown a little. No.4 sample is in 40°C, the absorbance is 3.86 and it change to the brownest which near to color-black. No.5 sample is in 60°C, the absorbance is 1.51 and it change to obviously pink.

Table: Effect of temperature on extent of browning

Graph: Effect of temperature on browning rate

2nd Experiment-Effect of PH on enzyme-catalyzed reactions

Through the second experiment-effects of pH on enzyme-catalyzed reactions, we get PH and absorbance of whole 5 sample with the use and compare of blank sample. No.1 sample with PH 3 and the absorbance is 1.76. No.2 sample with PH 5 and the absorbance is 1.08. No.3 sample with PH 7 and the absorbance is 2.98. No.4 sample with PH 9 and the absorbance is 0.51. No.5 sample with PH 11 and the absorbance is 0.98.

Table: Effect of PH on extent of browning

Graph: Effect of PH on the browning rate

· Discussion and Conclusion

To draw the results of temperature experiment, the curve is like a “mountain”, it goes up from 0°C and reaches the peak at 40°C and down to bottom at 60°C. As we can see, the best temperature for the rate of reaction is 40°C with 3.86 absorbance. In contrast, the lowest rate of reaction is 1.51 absorbance at 60°C. Obviously, our hypothesis was accepted because the rate of an enzyme catalyzed reactions is greatest at temperature that are 40°C. For a reaction to occur, the two reactant molecules must “bump into” each other; however, in this experiment, when the temperature at 0°C, the cold condition slow down the speed of molecules and enzymes, so that it directly decreases encounters between substrate and enzymes. Therefore, 2.34 absorbance at 0°C is not high like high temperature, and enzyme that under cold condition won’t highly efficiency like enzymes under higher temperature. Moreover, high temperature does not necessarily increase the efficiency of the enzyme. Just like enzymes at 1.51 absorbance at 60°C, it not efficiency like enzymes at 3.86 absorbance at 40°C.Why? Because enzymes are efficient in a narrow rang of temperature. It true that higher temperature makes enzymes fast and efficient, but once the temperature higher than a certain temperature, the enzymes become denatured. It means enzyme lose its shape and even not reactive like enzymes under low temperature condition.

Moreover, the PH experiment pattern just like the letter “W” in graph, it reaches the peak at PH 7. As we can see, the greatest rate of reaction at PH 7 which is 2.98 absorbance. The lowest rate of reaction is 0.51at PH 9. Our hypothesis was accepted because the rate of an enzyme catalyzed reactions is greatest at Ph values that are PH 7. To explain the change of the absorbance at low and high PH values, the best environment for enzymes’ reaction is neutral condition, since acid and basic environment will make enzymes become denatured. It means enzymes will lose their original shape when they are under acid and basic condition. As a result, 1.76-1.08 absorbance under acid condition and 0.51-0.98 absorbance under basic condition are not high as 2.98 absorbance under neutral condition which is PH 7. Moreover, some people know that the greatest rate of reaction for few types of enzymes are not PH 7(neutral condition), and their optimal PH is even around PH 2 which is true. Actually, in human body, typical enzymes are still highly efficient at PH 7 environment. However, some parts of human body may have a acid environment, like stomach; thus, enzymes that optimal PH is 2 would be found in acid environment like stomach.

Fortunately, earth is our home, and earth make us have nice environment to live. Some people may not live in optimal temperature rang; thus, once their enzymes not work in optimal range for a long time, they will start ill. Different like human being, there are few kinds of animals that live in extreme weather are able to properly adjust their enzymes’ efficiency, like penguin, white bear, deep-sea fish and dolphin. Actually, study what factors affect the enzyme is only a small step for natural science, there are lots of things that unknown wait for human to explore and study.

Absorbance

0°C Room temp 30°C 40°C 60°C 2.34 3.23 3.48 3.86 1.51

Temperature of water bath (°C)

Absorbance

Absorbance

3 5 7 9 11 1.76 1.08 2.98 0.51 0.98

PH

Absorbance

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