Equipment List

• Powdered milk suspension
• Trypsin solution (0.5%)
• Distilled water
• Hydrochloric acid (0.1M)
• 5 cm³ syringes
• Flat-bottomed tubes
• Water bath
• Timer

Method (Variable: Temperature)

1. Prepare Controls:

• Set up two flat-bottomed tubes with 5 cm³ of milk suspension each.
• In one tube, add 5 cm³ of distilled water to show the absence of enzyme activity.
• In the other, add 5 cm³ of hydrochloric acid to indicate the colour of a fully hydrolysed sample.

2. Set Up Experiment:

• Place 5 cm³ of milk in each of three test tubes and put them in a water bath at 10°C for 5 minutes to equilibrate.

3. Add Enzyme and Measure Reaction Time:

• Add 5 cm³ of trypsin to each test tube simultaneously and start the timer immediately.
• Record the time taken for each milk sample to completely hydrolyse and turn colourless.

4. Repeat at Different Temperatures:

• Repeat steps 2-3 at temperatures of 20°C, 30°C, 40°C, and 50°C.

5. Calculate Mean and Rate of Reaction:

• Find the mean time taken for hydrolysis at each temperature, then calculate the rate of reaction.

Conclusion

• Casein, a protein in milk, turns the milk colourless when broken down. Trypsin is a protease enzyme that hydrolyses casein.
• As the temperature increases from 10°C, kinetic energy increases, leading to more frequent enzyme-substrate complexes and a faster reaction rate.
• Beyond the enzyme's optimum temperature, bonds in the tertiary structure break, changing the enzyme's active site shape. This loss of complementarity prevents the enzyme from binding to the substrate effectively, reducing the reaction rate.