The Effects of Temperature on Enzymatic Activity

Introduction

What types of environmental factors may affect enzymatic activity? Why? Several factors known to affect enzymatic activity are temperature, pH, and substrate (the reactant molecule manipulated by the enzyme) concentration. In a typical chemical reaction, increasing temperature causes the substrates to become more energetic and hence more likely to bump into each other in solution. However, changes in temperature can cause an enzyme to denature, which changes the three-dimensional structure of the enzyme molecule.

Safety Precautions

• Goggles should be worn at all times while in laboratory
• No open toe shoes worn in laboratory
• Measure fluids carefully using graduated cylinders to avoid breakage and spillage.
• Be careful not to touch solutions of concentrated acids and bases directly.
• Take precautions when using a hot plate and touching hot glassware.
• Inform your teacher immediately of any broken glassware as it could cause injuries.
• Clean up any spilled fluids to prevent other people from slipping.

Materials

• Graduated cylinder
• Beakers
• Water
• Milk
• Lactase
• Test tubes
• Stirring rod
• Hot plates
• Ice bath
• Thermometers
• Glucose test strips
• Timer

For this activity, you will work in pairs.

Procedure

Step 1: Prepare a large beaker of boiling tap water on a hot plate, and another at 37 degrees C (human body temperature).  Prepare a third beaker with room temperature tap water, and make an ice bath in a fourth beaker.  Place thermometers in each.  Prepare four identical test tubes, each containing 2 mL of milk. Place one tube of milk in each of the four baths.  Place a small beaker next to each large one, on the table.  Measure out 1 ml of lactase solution into each small beaker.

Step 2:  Create a data table to enter your results for each of these test tubes over time.  Place each temperature in a separate row, and five columns corresponding to glucose levels to be measured at 3 minute intervals up to 15 min.   In each cell, record the elapsed time on the stopwatch (to start at the addition of lactase to the milk samples) at which each test strip should have its color analyzed.  These color assessments should each be made 6 minutes after each strip is dipped in the milk.

Step 3:   Discuss with your partner, who will be responsible for which roles in the experiment.

Step 4: Start the stopwatch, and immediately add 1 mL of the lactase enzyme solution to each of the four test tubes in the baths.

Step 5:  At 3 minutes, pour the milk from the test tubes each into their small beakers, dip the test strip, and pour the milk back into the test tube, which should remain in the water bath.  Set the test strip on the table adjacent.

Monitor the temperatures of each of these locations, both before and after the experiment using the thermometers.  Adjust the hot plates as needed.

Step 6: Repeat dipping the test strips into the milk at 6, 9, 12, and 15 minutes, returning the milk to the test tube in the bath each time.

Step 6:   In your lab notebook, describe your results, reporting the ranking of temperatures (in speed of glucose production by lactase from milk).  Did any temperatures have notably less glucose production than the others?  Can you speculate why?  Can you speculate why the fastest temperature, was the fastest?