1.9 Required practical 1

Question 1

What is the reasoning behind the method used in the practical

Answer 1

• The rate of an enzyme-controlled reaction is influenced by different factors: temperature, pH, concentration of the substrate and enzyme.
• The effect of each of these can be determined by changing a single variable and measuring its effect on the ROR.
• It is important to keep other variables constant so they don’t influence the results.

Question 2

Describe the method of required practical 1 (with named variable as temperature)

Answer 2

1. Make 2 control samples:
   - Take two flat bottomed tubes and add 5cm3 of milk suspension.
   - Add 5cm3 of distilled water to one of the tubes (this control will indicate the absence of enzyme activity).
   - Add 5cm3 of HCl to the other tube (this control indicates the colour of a completely hydrolysed sample).
2. Take 3 test tubes and measure 5cm3 of milk into each. Place in water bath at 10 degrees for 5 minutes to equilibriate.
3. Add 5cm3 of trypsin to each test tube simultaneously and start the timer immediately.
4. Record how long it takes for the milk samples to completely hydrolyse and become colourless.
5. Repeat steps 2-3 at 20, 30, 40 and 50 degrees.
6. Find the mean time for the milk to be hydrolysed at each temperature and use this to work out the ROR.

Question 3

What is the formula for calculating rate of reaction

Answer 3

Rate of reaction = 1/mean time

Question 4

What are 4 hazards in this practical and what are their risks

Answer 4

1. Broken glass - cuts from sharp object.
2. HCl - May cause harm/irritation to eyes or in cuts.
3. Hot liquids - Scalding.
4. Enzymes - allergies/ is an irritant.

Question 5

What are safety precautions of broken glass

Answer 5

• Take care when handling glass objects.
• Keep away from edge of desk.

Question 6

What are safety precautions for HCl

Answer 6

• Wear eye protection and gloves.
• Avoid contact with skin.

Question 7

What are safety precautions for hot liquids

Answer 7

• Handle with care.
• Use tongs to remove boiling tubes from water bath.
• Wear eye protection.
• Keep away from the edge of the desk.

Question 8

What are safety precautions for enzymes

Answer 8

• Avoid contact with skin/eyes.
• Wear eye protection and gloves.

Question 9

What should the graph look like for this practical

Answer 9

Plot a graph for the rate of reaction against temperature (rate of reaction on y axis, temperature on x axis).

Question 10

What is the conclusion of the experiment (explaining the results obtained)

Answer 10

• Milk contains a protein called casein which causes the milk to turn colourless when broken down.
• Trypsin is a protease enzyme which hydrolyses the casein protein.

Question 11

Why is the rate of reaction higher at higher temperatures

Answer 11

• As the temperature increases, the kinetic energy increases.
• This causes more enzyme-substrate complexes to form which increases the rate of reaction up to the optimum temperature.

Question 12

Why does the rate of reaction decrease at temperatures higher than the optimum temperature

Answer 12

• At temperatures beyond the optimum, bonds in the enzyme tertiary structure break which changes the shape of the active site.
• This means the enzyme is no longer complementary to the substrate (enzymes denature) so less and less enzyme-substrate complexes can form.

Question 13

What is the independent variable in this practical

Answer 13

The temperature

Question 14

What is the dependent variable in this practical

Answer 14

The rate of reaction (found through recording the time).

Question 15

What are some control variables in the practical (name at least 3)

Answer 15

• Volume of trypsin used
• Volume of milk used
• Control test tubes: milk suspension and distilled water, milk suspension and hydrochloric acid).
• Using same person to observe the colour change (decide when the solution has become colourless).