High performance liquid chromatography with an electrochemical detector in the cathodic mode as a tool for the determination of p-nitrophenol and assay of acid phosphatase in urine samples.
Utilizing a commercially available helium-purging device and PEEK tubes for all tubing, especially for connection between the mobile phase and pump, high performance liquid chromatography with an electrochemical detector (ECD/HPLC) at the cathodic mode is a simple and precise method for the determination of p-nitrophenol (NP). Studies with cyclic and hydrodynamic voltammetry indicated that 25% aqueous MeOH containing 0.1% (v/v) CF(3)CO(2)H and -0.8 V vs. Ag/AgCl are the best mobile phase and detection potential for cathodic ECD/HPLC. With the present system, the limits of detection and determination were 0.2 and 0.25 microM, respectively, and up to 50 microM, a linear calibration curve was afforded. Within-day precisions for the analysis of 5 and 50 microM NP were 0.8 and 0.7% (n=6), respectively, and between-day precisions (n=6) for these samples were 3.5 and 2.2%, respectively. Compared with the commonly used Bessey-Lowry-Brock method, cathodic ECD/HPLC was useful for the assay of acid phosphatase in urine samples with p-nitrophenyl phosphate disodium salt as a substrate.
Yamauchi
Yuji
Y
Graduate School of Pharmaceutical Sciences, Osaka University, Japan.
Ido
Megumi
M
Ohta
Merime
M
Maeda
Hatsuo
H
D000135:Acid Phosphatase / Q000652:urine*; D002851:Chromatography, High Pressure Liquid / Q000379:methods / Q000592:standards; D004563:Electrochemistry; D004566:Electrodes / Q000592:standards; D006801:Humans; D009596:Nitrophenols / Q000652:urine*
D009596:Nitrophenols; D000135:Acid Phosphatase; C024836:4-nitrophenol