4-Nitrophenol in 4-nitrophenyl phosphate, a substrate for alkaline phosphatase, as measured by paired-ion high-performance liquid chromatography.
We used paired-ion high-performance liquid chromatography to determine the 4-nitrophenol content of 4-nitrophenyl phosphate, a substrate for alkaline phosphatase analysis. This was done on a reversed-phase column with a mobile phase of methanol/water, 45/55 by vol, containing 3 ml of tetrabutylammonium phosphate reagent per 200 ml of solvent. At a flow rate of 1 ml/min, 4-nitrophenol was eluted at 9 min and monitored at 404 nm; 4-nitrophenyl phosphate was eluted at 5 min and could be monitored at 311 nm. Samples of 4-nitrophenyl phosphate obtained from several sources contained 0.3 to 7.8 mole of 4-nitrophenol per mole of 4-nitrophenyl phosphate.
D009597:4-Nitrophenylphosphatase / Q000032:analysis; D000469:Alkaline Phosphatase; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D009596:Nitrophenols / Q000032:analysis*; D009943:Organophosphorus Compounds / Q000032:analysis*; D013053:Spectrophotometry / Q000379:methods
D009596:Nitrophenols; D009943:Organophosphorus Compounds; D000469:Alkaline Phosphatase; D009597:4-Nitrophenylphosphatase