BIO 141L Grossmont Effect of Temperature on Lactase Activity Lab Report

1. In this activity, you will study the effect of pH, temperature, and lactose concentration on lactase activity. By the end of this activity, you should be able to: a. design experiments to study the effects of pH, temperature, and substrate concentration on enzyme activity. b. analyze data related to enzyme activity. c. identify the components of the scientific method. d. identify components of the substrate-enzyme-product system being studied. e. summarize and communicate your lab data. • To prepare for the lab, watch the following video and think about enzymes, substrates, products, maximum rate of reaction, and optimal conditions. https://www.youtube.com/watch?v=D2j2KGwJXJc • To complete the lab activity, review the Zoom link for lab posted on Canvas by Wednesday of this week and go to the following website (https://sites.google.com/site/biologydarkow/lactaseenzyme-simulation The simulation is found on page 4 of the site or you can use the simulate tab. • You do not submit this document on Canvas. This document includes the instructions you need to complete the lab. You will submit the Lactase Enzyme Data document where you will report your results. Experiment I: The effect of pH on lactase activity. Click on the Simulate tab. The standard condition settings are: lactose for 500 mg/dL , pH at 7.0 and temperature at 250C, and 5 mmol lactase enzyme solution. You will vary one of these conditions in each experiment. In this Experiment I, you are varying pH. The condition you are varying is the independent variable for that experiment. The outcome you measure for that experiment is the dependent variable. The dependent variable in Experiment I is glucose produced and is a measure of the rate of the enzyme activity. The hypothesis of an experiment is a tentative proposed relationship of the independent and dependent variable of the experiment. The hypothesis of Experiment I is pH does not affect the amount of glucose produced in a reaction catalyzed by lactase. Experiment I has two parts: I. determining the interval of interest and II. determining the optimum pH for lactase activity. I. Determining the interval of interest. Before you can begin your experiment, you must determine the interval of pH which will give you the most valuable, relevant data. Start by running a simulation with the standard initial lactose, temperature and lactase enzyme solution. Run simulations at a pH of 7. Record the glucose concentration (mg/dL) at 4 (you do not need to record the data for 7 and 11 minutes in this part) and repeat for a total of 5 runs. Repeat the simulations at a pH of 3, 5, 9, and 11. Make sure to have 5 runs for each pH setting. Record the glucose concentration at 4 minutes for each of the runs. Record all your data in the table. pH Run 1 (mg/dL) Run 2 (mg/dL) Run 3 (mg/dL) Run 4 (mg/dL) Run 5 (mg/dL) Mean (mg/dL) Range Range (High-Low) 3 5 7 9 11 Using the above data to produce a line graph (See the Zoom recording for this lab). Use the line graph to help you determine the interval of interest. This will be covered in the Zoom lab. II. Use the data and graph to determine the pH interval which will give you the most valuable, relevant data for determining the optimum pH. Divide this interval into ten (10) equally spaced intervals (for example if you determine the interval between pH 7 and 11 is where the peak enzyme rate is occurring, you will collect data at 7.4, 7.8, 8.2, 8.6 …. 10.2, 10.6 and 11). Adapt the data collection table to record the results of these simulations at 4 minutes for five (5) runs for each pH level. Run the simulation and record the data. pH Run 1 (mg/dL) Run 2 (mg/dL) Run 3 (mg/dL) Run 4 (mg/dL) Run 5 (mg/dL) Mean (mg/dL) Range Range (High-Low) Graph the data and determine the optimum pH (where maximum lactase enzyme activity is likely located). Experiment II: The effect of temperature on lactase activity. You will now use Experiment I as a model to design an experiment to test the effect of temperature on lactase activity. Experiment II has two parts: I. determining the interval of interest for the effect of temperature on lactase activity and II. determining the optimum temperature for lactase activity. First determine the independent variable of the temperature experiment. Next determine the dependent variable of the temperature experiment. Identify the variables you will standardize in the temperature experiment. Write out the hypothesis of your experiment. Record this information in the Lactase Data Sheet. I. Determine the interval of interest for temperature. Choose a wide range of temperatures. You can try a few temperatures to see those conditions result in any glucose production and then work between the temperatures with low lactase activity. Determine these temperatures and record them in the table below. Use the simulation website to determine the glucose produced (rate of lactase activity) at 4 minutes for each of these temperatures. Temp 0 C Run 1 (mg/dL) Run 2 (mg/dL) Run 3 (mg/dL) Run 4 (mg/dL) Run 5 (mg/dL) Mean (mg/dL) Range Range (High-Low) After you complete the table, use the data to determine the inflection point based on your results. The interval of interest is the range of temperatures which will give you the most valuable, relevant data for determining the optimum temperature in the second part of the experiment. Copy and paste the above data table into the data document you will submit on Canvas. II. Use the temperature interval of interest to design the second part of the experiment. Divide this interval into equally spaced intervals. Use the table below to record the results of these five simulations at 4 minutes for each temperature. Run the simulation and record the data. Temp 0 C Run 1 (mg/dL) Run 2 (mg/dL) Run 3 (mg/dL) Run 4 (mg/dL) Run 5 (mg/dL) Mean (mg/dL) Range Range (High-Low) Use the data from the table to determine the optimum temperature (where maximum lactase enzyme activity is likely located). Copy and paste the above data table into the data document you will submit on Canvas. Experiment III: The effect of lactose concentration on lactase activity. This experiment is NOT part of the data sheet you will submit on Canvas. It is part of the quiz you will complete this week. For this experiment, use Experiment I and Experiment II as models for studying the effect of lactose concentration on lactase activity. Note that you will need to develop your own data collection tables to record data. Design an experiment to test the effect of lactose concentration on lactase activity. What is the independent variable of this experiment? What is the dependent variable of this experiment? Which variables will you standardize? What is the hypothesis of this experiment? Describe your experiments. What is the optimum lactose concentration for maximum lactase activity? Experiment IV: Studying specificity using lactase activity. This experiment is part of the data sheet you will submit on Canvas (question 5) and it is part of the quiz you will complete this week. Think about the following question: Are you able to design an experiment to study specificity in lactase using the Lactase Enzyme Simulation? Yes or No Explain specificity and the experiment you would design to study this concept in lactase using the simulation or how you would need to modify the simulation to test this concept. You don’t need to do this experiment, just explain its design. You do not submit this document on Canvas. This document includes the instructions you need to complete the lab, but the data you submit is the completed Lactase Enzyme Data document.

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