What is the role of the buffer used in the tubes during the amylase activity assay?

• A. to provide additional substrate
• B. to denature the enzyme
• C. to increase temperature
• D. to maintain pH at 7.0

Answer: (D)

Enzyme activity is highly sensitive to pH, so the buffer’s job is to keep the reaction environment at a constant, optimal pH. In this amylase assay, maintaining pH around 7.0 ensures the enzyme’s active site residues stay correctly charged and the substrate binds effectively. If the pH drifted, the enzyme could slow down or denature, and the results would reflect pH changes rather than true enzyme activity. The buffer does not add substrate, nor does it denature the enzyme or raise the temperature—that temperature control comes from the incubator or water bath. So keeping pH at 7.0 makes the measured activity accurate and reproducible.