Browning in apples: Exploring the biochemical basis of an easily-observable phenotype

Many fruits and vegetables undergo browning when they are cut and the tissue is exposed to the air. This is due to the activity of the enzyme polyphenol oxidase (PPO, EC 1.14.18.1) with endogenous substrates. In this laboratory experiment, students prepare slices of different varieties of apples and assess the rate of browning. They make a simple extract of the apple tissue and measure the activity of PPO using 3,4-dihydroxy-l-phenylalanine (l-DOPA) as substrate. They determine the protein concentration of the extract with the Bradford Coomassie Blue reagent and calculate the specific activity of PPO. Finally, the students measure the total concentration of the potential substrates for PPO with the Folin-Ciocalteau phenol reagent using a gallic acid standard curve. By comparing the tendency of the apples to turn brown, the specific activity of PPO, and the concentration of potential substrates, they can assess the biochemical basis of the browning phenotype. This experiment can be done as a series of weekly laboratory exercises, as an intensive 1-week laboratory project, or as the basis of an extended student research investigation. © 2017 by The International Union of Biochemistry and Molecular Biology, 46(1):76-82, 2018.