Investigate the progress of enzyme-catalysed reactions by measuring rates of formation of products using catalase and rates of disappearance of substrate using amylase

Investigation: Progress of Enzyme-Catalysed Reactions

Reactions Under Investigation:

1. Catalase-Catalysed Reaction:
   • Reaction: 2H₂O₂ (Hydrogen Peroxide) → 2H₂O (Water) + O₂ (Oxygen)
   • Measured: Rate of Formation of Product (O₂)
2. Amylase-Catalysed Reaction:
   • Reaction: Starch (or Amylose) → Maltose (and eventually Glucose)
   • Measured: Rate of Disappearance of Substrate (Starch)

Experimental Design:

Catalase-Catalysed Reaction (O₂ Formation)

• Materials:
  • Catalase solution
  • Hydrogen peroxide (H₂O₂)
  • Gas syringe or oxygen sensor
• Procedure:
  1. Prepare several identical reactions with catalase and H₂O₂.
  2. Initiate each reaction at timed intervals (e.g., every 2 minutes).
  3. Measure the volume of O₂ produced (using a gas syringe) or the concentration of O₂ (using an oxygen sensor) at fixed time points (e.g., every 1 minute) after initiating each reaction.
  4. Plot the volume/concentration of O₂ vs. time for each reaction start time to observe the rate of product formation over time.
• Data Analysis:
  • Calculate the initial rate of reaction (rate of O₂ formation) for each reaction using the linear portion of the plot.
  • Compare the initial rates among the reactions to assess the consistency of the catalase activity.

Amylase-Catalysed Reaction (Starch Disappearance)

• Materials:
  • Amylase solution
  • Starch solution
  • Iodine solution (for starch detection)
• Procedure:
  1. Prepare several identical reactions with amylase and starch.
  2. Initiate each reaction at timed intervals (e.g., every 2 minutes).
  3. At fixed time points (e.g., every 1 minute) after initiating each reaction, withdraw a sample and add iodine solution.
  4. Measure the absorbance (using a spectrophotometer) or color intensity (visually or with a colorimeter) of the resulting solution, which correlates with the remaining starch concentration.
  5. Plot the absorbance/color intensity (proportional to starch concentration) vs. time for each reaction start time to observe the rate of substrate disappearance over time.
• Data Analysis:
  • Calculate the initial rate of reaction (rate of starch disappearance) for each reaction using the linear portion of the plot.
  • Compare the initial rates among the reactions to evaluate the consistency of the amylase activity.

Expected Outcomes and Interpretations:

• Catalase Reaction:
  • Plot: Volume/Concentration of O₂ vs. Time will show a rapid increase in O₂ production initially, followed by a plateau as H₂O₂ is depleted.
  • Initial Rate Analysis: Consistent initial rates among reactions indicate reliable catalase activity.
• Amylase Reaction:
  • Plot: Absorbance/Color Intensity (proportional to Starch Concentration) vs. Time will decrease over time, indicating starch breakdown.
  • Initial Rate Analysis: Similar initial rates across reactions suggest consistent amylase activity.

Factors Influencing Rates (Variables to Consider for Future Investigations):

• Enzyme Concentration: How does varying the amount of catalase or amylase affect the reaction rates?
• Substrate Concentration: What impact does changing the initial H₂O₂ or starch concentration have on the reaction rates?
• Temperature and pH: How do variations in temperature and pH affect the activity of catalase and amylase, and consequently, the reaction rates?