Ph Concentration Lab Report

The experimental samples for the pH concentration experiment were put together by using a 10ml-graduated cylinder to obtain 4ml of each pH buffer to insert into cuvettes, a micropipette was then used to obtain 0.5ml of catechol and 0.5ml of the catechol oxidase. The pH buffer was made first to avoid any denaturation of the catechol oxidase. Our positive control for this experiment was pH 7 because that is the pH level of most cell membranes in the cytoplasm (Whitson, 2016.) Our negative controls varied for each pH buffer, but all “blank” cuvettes contained 0.5 ml of catechol oxidase and 4.5ml of the pH buffer. We prepared the negative control for each pH buffer before preparing the experimental sample because each cuvette had to be mixed for exactly 3 minutes as instructed in the lab manual (Scott et al, 2016) and we tried avoid denaturing the enzyme. The benzoquinone had a greater absorption at acidic pH levels because the acidic solution denatures catechol oxidase, causing it to lose its secondary and
We used 0.5ml of the 5mM catechol solution and 4ml of pH 6 buffers. The “blank” or negative control in this experiment contained 0.5ml of catechol oxidase and 4.5ml of pH 6 buffers. It was decided that the positive control was the sample inside the cuvette that was inserted into the spectrophotometer after 3 minutes of mixing. We followed instructions from the lab manual on the previous experiment (Scott et al, 2016.) The more time the pH and the catechol solution were mixed the more absorption it had. The acidity of the pH denatured the enzyme by breaking down hydrogen bonds in the alpha helix and beta pleated sheets; therefore the catechol oxidase loses its secondary and tertiary structure. When the enzyme loses its structure, it also loses its active site. Without an active site, the catechol (substrate) is not able to bind and benzoquinone is not made as efficiently as it