Simultaneous determination of phenylglyoxylic acid, mandelic acid, styrene glycol and hippuric acid in primary culture of rat hepatocytes incubate by high-performance liquid chromatography.
A simple HPLC method for the simultaneous determination of phenylglyoxylic acid (PGA), mandelic acid (MA), styrene glycol (SG) and hippuric acid (HA) in cell culture medium was developed. Analysis was performed on a C(18) column with a mobile phase composed of methanol-potassium dihydrogen phosphate (pH 2.5; 10 mM; 10:90, v/v) at 220 nm. The flow-rate of mobile phase was set at 0.5 mL/min. The mean absolute recoveries of PGA, MA, SG and HA were 95.9, 98.4, 98.0 and 97.1%, respectively. The inter-day and intra-day precisions, determined at three concentration levels, were less than 10% of RSD. The limits of quantification for PGA, MA, SG and HA were 13.2, 13.1, 14.5 and 11.2 microM with RSD less than 20%. The limits of detection for PGA, MA, SG and HA were 4.6, 4.6, 5.1 and 3.9 microM, respectively. The method was successfully applied to study the stereoselective metabolism of SG and MA in primary culture of rat hepatocytes. The results show that there is stereoselective metabolism for both of MA and SG in primary culture of rat hepatocytes. The extent of biotransformation from S-MA to PGA is significantly greater than that from the R enantiomer and the main metabolites are PGA and HA for S-SG and R-SG, respectively.
Wang
Jin-Zhao
JZ
Department of Pharmaceutical Analysis and Drug Metabolism, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, Zhejiang 310031, People's Republic of China.
Lu
Xin-Yan
XY
Zhao
Na-Ping
NP
Cheng
Yi-Yu
YY
Zeng
Su
S
D000818:Animals; D002138:Calibration; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003470:Culture Media; D005026:Ethylene Glycols / Q000032:analysis*; D006038:Glyoxylates / Q000032:analysis*; D022781:Hepatocytes / Q000737:chemistry*; D006626:Hippurates / Q000032:analysis*; D008297:Male; D008333:Mandelic Acids / Q000032:analysis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet
D003470:Culture Media; D005026:Ethylene Glycols; D006038:Glyoxylates; D006626:Hippurates; D008333:Mandelic Acids; C012482:phenylglyoxylic acid; C031474:styrene glycol; C037938:mandelic acid; C030514:hippuric acid