1.10 - Core Practical - Effect of pH on Enzyme Activity Flashcards
What is the focus of the practical experiment?
The effect of pH on the rate of activity of a particular enzyme.
What enzyme is being used in the experiment?
Amylase.
What does amylase break down?
Carbohydrates such as starch into simple sugars such as maltose.
What indicator is used to check for the presence of starch?
Iodine, which turns dark orange.
What color indicates the presence of starch when using iodine?
Blue-black.
What is the purpose of the experiment regarding amylase?
To estimate its optimal pH.
What materials are required for the experiment?
1% Amylase solution, 1% Starch solution, Iodine solution, Buffer solutions of different pH values (e.g. pH 4, 6, 7, 8), Test tubes and rack, Dropping pipette, Spotting tile (tray with wells), Stopwatch, Measuring cylinders / syringes, Beaker with water, Bunsen burner, gauze, tripod (for water bath setup), Thermometer.
What is the first step in the method?
Add iodine solution into each well of the spotting tile – 1 drop per well.
How do you set up the water bath?
Heat a beaker containing ~50ml of water for 3 minutes above a Bunsen burner.
What should be added to the labeled test tube?
2 cm³ of amylase solution, 2 cm³ of starch solution, 1 cm³ of the buffer solution (for desired pH).
What is done after placing the test tube in the water bath?
Start the stopwatch.
What should be done after 10 seconds?
Use a pipette to take a small sample from the test tube and add it to one of the wells containing iodine.
What indicates the presence of starch?
If starch is present, the well turns blue-black.
What should you do until the iodine no longer changes color?
Repeat sampling every 10 seconds, adding a drop to a new well each time.
What does it mean when the iodine remains orange?
This means all the starch has been broken down.
What should be recorded during the experiment?
Record the time it took for the reaction to complete (when no blue-black colour appears).
What should you do after completing the first trial?
Repeat steps 1–7 using buffer solutions of different pH values.
What should be plotted on the graph?
X-axis: pH of buffer solution; Y-axis: Time taken for starch to be completely digested.
What is the aim of the experiment?
To investigate how pH affects the rate of starch breakdown by the enzyme amylase.
Why do we use a Bunsen Burner and water beaker?
We use this equipment to keep the solution at a relatively constant temperature throughout the reaction (temperature is a control variable in this experiment).
What results do we expect to see?
The optimal pH of amylase will be at whichever pH the reaction completes in the shortest time. This should be somewhere around pH 7.0.
