Used as a chromogenic substrate in enzymatic assays, particularly for measuring the activity of proteolytic enzymes such as chymotrypsin and other serine proteases. Upon enzymatic hydrolysis, it releases 4-nitroaniline, which produces a yellow-colored solution that can be measured spectrophotometrically at around 405 nm. This property makes it valuable in biochemical research and diagnostic applications where real-time monitoring of enzyme activity is required. It is also employed in kinetic studies to determine enzyme specificity, inhibition, and catalytic efficiency.