Used as a biochemical reagent in enzyme assays, particularly for studying glutamine-dependent enzymes. It serves as a chromogenic substrate in the detection and quantification of amidases and glutaminases, where enzymatic hydrolysis releases 4-nitroaniline, producing a yellow color measurable spectrophotometrically. This property makes it valuable in diagnostic applications and biochemical research for monitoring enzyme activity and inhibition. Also employed in the development of analytical methods for microbial enzyme profiling and in high-throughput screening of enzyme inhibitors.