Effect of pH on Amylase- Practical

• Iodine solution


  A chemical that turns dark blue-black in the presence of starch, allowing detection of whether amylase has broken down starch
• Using iodine solution to determine rate of amylase reaction

  1. Add iodine solution to sample

  2. Observe color change

  3. No color change = starch digested

  4. Blue-black color = starch present
• Required practical investigation

  • Investigate impact of pH on activity of amylase enzyme
  • Use continuous monitoring method
  • Control temperature using water bath
• Independent variable


  The pH, which is changed between trials
• Dependent variable


  The rate of reaction of the amylase enzyme
• Control variables

  • Concentration and volume of starch solution and amylase solution
  • Temperature of reaction
• Procedure for investigation

  1. Prepare tubes with starch solution and pH buffers

  2. Prepare spotting tile with iodine solution

  3. Place tubes and spotting tile in water bath at 37°C

  4. Add amylase to first tube, start timer

  5. Every 30 seconds, remove sample from tube and add to iodine on spotting tile

  6. Note time when iodine no longer changes color (all starch digested)

  7. Repeat investigation at different pH values
• Optimum pH for amylase is likely near pH 7 (for salivary amylase) or more alkaline (for intestinal amylase)
• Extreme pH values (e.g. pH 2, pH 12) are likely to denature the amylase enzyme
• To improve the investigation, test more pH values within a narrow range around the suspected optimum pH