Establishment of in vitro metabolism of fentanyl by human liver microsomes in Chinese population / 中华麻醉学杂志

ABSTRACT

Objective To establish in vitro metabolism of fentanyl by human liver microsomes in Chinese population.Methods Thirty patients undergoing elective operation on liver were enrolled in the study.Normal liver specimens were obtained during removal of liver and gall for preparation of liver microsomes (by calcium precipitation) which were used for establishment of the liver microsomal incubation system for fentanyl.Fentanyl served as the metabolic substrate in the incubation reaction.The concentration of fentanyl in the incubation medium was detected at 0,5,10,15,20 and 30 min of incubation using HPLC-UV.Sufentanil served as the interior label element.The n-hexane-ethanol absolute was used to extract the sample.The chromatographic column used in this method was Grace C18 (4.6 mm × 250.0 mm,5 μm).The mobile phase was methyl cyanide-KH2PO4 buffer solution with the flow rate of 1.0 ml/min,detection wavelength of 205 nm and sample size of 20 μl.Linear regression analysis was performed by using the least-squares method.The specimens of the blank incubation system with the final concentration of fentanyl 0.6,2.4 and 10.0 μg/ml were obtained to determine the recovery,precision and stability.The metabolic rate of fentanyl in human hepatic microsomes was calculated.Results Fentanyl and the interior label element sufentanil were separated completely,and the retention time were 5.730 and 9.336 min,respectively.Endogenous matrix of microsomes did not interfere with the analysis.Regression equation was C =0.945 8A-0.140 4,R2 =0.999 2.C was the concentration of fentanyl,and A was the peak area ratio of fentanyl versus sufentanil.The recovery of incubation system with low,medium and high concentrations of fentanyl was 85％-115％,and relative standard deviation (RSD) was less than 10％.The RSD of intra-and inter-day precision and stability was less than 10％.The method was proved to meet the requirements of biological sample analysis.The metabolic rate of fentanyl was (1.6 + 0.8) nmol/min per milligram protein in human hepatic microsomes of 30 cases.Conclusion The in vitro metabolism of fentanyl by human liver microsomes is convenient,and the detectability is high,so it can be used for the research on the in vitro metabolism of fentanyl in Chinese population.