A 4-nitrophenyl β-glycoside of lactose (Galβ1-4Glc), in which the aglycone is linked through the reducing-end glucose. Used as a chromogenic substrate for β-galactosidases and lactases (LacZ-type and mammalian intestinal lactase / lactase-phlorizin hydrolase); enzymatic cleavage of the terminal β1-4 galactosidic bond, or of the aglycone-proximal glucosidic bond by β-glucosidases, ultimately releases 4-nitrophenol. 4-Nitrophenyl (pNP) glycosides are chromogenic enzyme substrates: cleavage of the glycosidic bond by the target enzyme releases free 4-nitrophenol, which is yellow under basic conditions (absorbance maximum ~405 nm), enabling straightforward spectrophotometric assays of enzyme activity in solution and in high-throughput screening formats.