New high-performance liquid chromatography method for the determination of (R)-warfarin and (S)-warfarin using chiral separation on a glycopeptide-based stationary phase.

Warfarin is a well-known anticoagulant agent that occurs in two enantiomers, (R)-(+)-warfarin and (S)-(-)-warfarin. A new liquid chromatography method for the determination of both enantiomers was developed, validated and applied in in vitro studies with the aim of evaluating the accumulation of (R)-warfarin and (S)-warfarin in the hepatoma HepG2 cell line. OptiMEM cell cultivation medium samples and cellular lysates were purified using Waters Oasis MAX extraction cartridges. The chiral separation of warfarin and the internal standard p-chlorowarfarin enantiomers was performed on an Astec Chirobiotic V2 column at a flow rate of 1.2mL/min. The mobile phase was composed of 31% acetonitrile, 5% of methanol and 64% of ammonium acetate buffer (10mmol/L, pH 4.1). The enantiomers were quantified using a fluorescence detector (lambda(excit)=320nm, lambda(emiss)=415nm). The limit of detection was found to be 0.121micromol/L of (S)-warfarin and 0.109micromol/L of (R)-warfarin. The range of applicability and linearity was estimated from 0.25 to 100micromol/L. The precision ranged from 1.3% to 12.2% of the relative standard deviation, and the accuracy reached acceptable values from 95.5% to 108.4%. The new bioanalytical method confirmed the same accumulation of (R)-warfarin and (S)-warfarin in the hepatoma HepG2 cell line.

Tissue and plasma concentrations of antibiotic during cardiac surgery with cardiopulmonary bypass--microdialysis study.

AIM: Surgical-site infections are very serious complications of cardiac operations. Use of cardiopulmonary bypass (CPB) is associated with profound physiological changes, which affects the pharmacokinetic behaviour of prophylactic antibiotics. The aim of this study was to monitor tissue concentrations of cefuroxime in peripheral tissue (skeletal muscle) during cardiac surgery using CPB by means of a microdialysis. METHODS: Eleven adult patients operated on using CPB were included in the study. Cefuroxime was the prophylactic antibiotic and study drug given. Microdialysis was performed by probe CMA 60 inserted into the patient's left deltoid muscle. Samples of dialysates were collected at intervals: before CPB, each 30 minutes of CPB and at the end of CPB. Samples of blood were collected at intervals: incision, start of CPB, each 30 minutes of CPB, at the end of CPB and at the end of surgery. RESULTS: The mean (+/- S.D.) concentrations of cefuroxime in peripheral tissue were 105.4+/-41.1, 81.7+/-32.8, 74.6+/-26.0, 70.4+/-34.7, 60.5+/-27.2, 138.0+/-42.6 (mg l(-1)). Total plasma concentrations of cefuroxime were 154.4+/-41.6, 73.3+/-20.7, 67.1+/-20.4, 59.2+/-21.0, 49.0+/-16.4, 110.9+/-33.6 (mg l(-1)) and concentrations of free plasma fraction were 110.7+/-37.1, 62.2+/-18.8, 58.9+/-18.6, 48.4+/-16.6, 41.7+/-15.6, 97.6+/-28.6 (mg l(-1)). The plasma and tissue concentrations exceed throughout the operation time the minimum inhibitory concentration for most common suspected pathogens in cardiac surgery. CONCLUSIONS: Results show that CPB can modify the time course of cefuroxime tissue and plasma concentrations. Microdialysis is suitable for antibiotic tissue measurement in cardiac surgery.