When you test the effect of pH on enzyme activity, you should vary the pH. However, you can do this in good or bad ways. Keep in mind what extra factors may confound the effects of varying pH. Otherwise, the results obtained may not be due to the change in pH, but some other factor. Knowing how to properly vary pH and what factors confound an experiment’s pH will help you get good results and understand why your results may not be exactly what you expected.

When testing the effect of pH on enzyme activity, vary only the pH while holding other factors constant. These other factors include enzyme concentration, substrate concentration and temperature. The factors that remain constant are called the control variables. Control variables allow you to conclude that the results on enzyme activity obtained in your experiment are due to the varying of pH, the independent variable. Knowing what factors not to change in an experiment is just as important as knowing what factor to vary, otherwise it will be difficult to conclude whether the results are actually due to the one thing that was tested.

The pH of a solution can be changed by dissolving different amounts of an acid or a base in water. One way to test the effect of pH on enzyme activity is to gradually add drops of a strong acid or a strong base into the solution that contains the enzyme, and then observe the point at which enzyme activity slows down or stops. An acid is defined as a compound that donates a hydrogen ion, called a proton (H+), and a base is defined as a compound that donates a hydroxide ion (-OH). Different acids and bases have different numbers of protons or hydroxide ions to give away. Not all protons or hydroxide ions are immediately donated when an acid or base to added to a solution, but the number of donated protons or hydroxide ions changes the pH at different rates. Thus, it is a good idea to vary the pH in an enzyme experiment using only one type of acid or one type of base. Otherwise, other variables are unintentionally added.

Some laboratory experiments that study enzyme activity involve grinding up fresh tissue to release the enzymes from the cells and then adding substrate to measure enzyme activity. Fresh tissue contains blood. Due to the presence of enzymes in blood that change the carbon dioxide gas that is dissolved in the blood into carbonic acid, the tissue itself may effect pH. Thus, in experiments that involve enzyme activity in fresh tissue, it is helpful to wash off the blood in a beaker of cold water before grinding the tissue. This will minimize the unintended change of pH due to the tissue, so that the purposeful change in pH can be studied.

As discussed above, enzyme concentration is a control factor that should not be varied when testing the effect of pH on enzyme activity. However, experimental procedures still inherently vary enzyme concentration in subtle ways. If one is using a pure solution of enzymes, then keeping enzyme concentration constant. However, in experiments in which the enzyme is from fresh tissue, such as chunks of potato, pieces of plants, or pieces of liver, the size of the chunks changes the amount of enzyme in each test tube. Thus, it will be helpful to cut the pieces of tissue as uniformly as possible. This is another example of how knowing what not to change, and why it’s hard not to avoid change completely, helps interpret the results of varying a factor such as pH.