Ortho-Nitrophenyl-β-galactoside

ortho-Nitrophenyl-β-galactoside (ONPG) is a colorimetric and spectrophotometric substrate for detection of beta-galactosidase activity. This compound is normally colorless. However if β-galactosidase is present, it hydrolyzes the ONPG molecule into galactose and ortho-nitrophenol. The latter compound has a yellow coloration that can be used to check for enzyme activity by means of a colorimetric assay.

Beta-galactosidase is required for lactose utilization, so the intensity of the color produced can be used as a measure of the rate that Beta-galactosidase can hydrolyze lactose.

Though ONPG mimics lactose and is hydrolysed by Beta-galactosidase, it is unable to act as an inducer for the lac operon. Without another lactose analog that can act as an inducer, such as IPTG, Beta-galactosidase will not be transcribed and ONPG will not be hydrolysed.

O-Nitrophenyl-β-D-galactopyranosid Ortho-nitrophenyl-β-galactoside