Investigating Enzymatic Reactions

• The enzyme amylase catalyses the breakdown of starch to maltose. It's easy to detect starch using iodine solution - if starch is present, the iodine solution will change from browny-orange to blue-black.
• To investigate how pH affects amylase activity, the first two steps would be:

  1. Put a drop of iodine solution into every well of a spotting tile.
  2. Place a Bunsen burner on a heat-proof mat, and a tripod and gauze over the top. Burn a beaker of water on top of the tripod and heat the water until it is 35 degrees C (use a thermometer to measure the temperature). Try to keep the temperature of the water constant throughout the experiment.
• When investigating how pH affects amylase activity, once you have heated the water to 35 degrees C, the next three steps would be:

  1. Use a syringe to add 1cm^3 of amylase solution and 1cm^3 of a buffer solution with a pH of 5 to a boiling tube. Using test tube holders, put the tube into the beaker of water and wait for five minutes.
  2. Next, use a different syringe to add 5cm^3 of a starch solution to the boiling tube.
  3. Immediately mix the contents of the boiling tube and start a stop clock.
• When investigating how pH affects amylase activity, once you've started the stop clock, the last two steps would be to:

  1. Use continuous sampling to record how long it takes for the amylase to break down all the starch. To do this, use a dropping pipette to take a fresh sample from the boiling tube every 30 seconds and put a drop into a well. When the iodine solution remains browny-orange, starch is no longer present.
  2. Repeat the whole experiment with buffer solutions of different pH values to see how pH affects the time taken for the starch to be broken down.
• When investigating the effect of pH on enzyme activity, remember to control any variables each time (e.g. concentration and volume of amylase solution) to make it a fair test.
• It's often useful to calculate the rate of reaction after an experiment. Rate is a measure of how much something changes over time.
• If an experiment measures how much something changes over time, you calculate the rate of reaction by dividing the amount that it has changed by the time taken.
• The enzyme catalase, catalyses the breakdown of hydrogen peroxide into water and oxygen. During an investigation into the activity of catalase, 24cm^3 of oxygen was released in 50 seconds. If you calculate the rate of reaction in cm^3, the answer would be 0.48cm^3/s because the amount of product formed was 24cm^3 and the time it took was 50s so 23/50 = 0.48cm^3/s.