This compound is primarily utilized in biochemical research as a substrate for studying the activity of proteolytic enzymes, particularly chymotrypsin and other serine proteases. Its cleavage by these enzymes releases p-nitroaniline, which can be easily detected spectrophotometrically due to its distinct yellow color. This makes it a valuable tool in enzyme kinetics studies, allowing researchers to measure enzyme activity and inhibition. Additionally, it is used in the development and optimization of assays for drug discovery, where understanding enzyme behavior is critical. Its specificity and sensitivity make it a reliable choice for high-throughput screening and mechanistic studies in enzymology.