High-throughput determination of valproate in human samples by modified QuEChERS extraction and GC-MS/MS.

A new high-throughput method was developed for analysis of valproate in human plasma samples by QuEChERS extraction and gas chromatography-tandem mass spectrometry (GC-MS/MS). Plasma samples (0.2 ml) spiked with valproate and secobarbital-d5 (internal standard) were diluted with 1.3 ml of distilled water. Acetonitrile (1 ml) was added followed by 0.4 g MgSO4 and 0.1 g NaOAC. After a centrifugation step (2000 g for 10 min), 1 ml of the supernatant was transferred to a dispersive-solid phase extraction (dSPE) tube containing 150 mg MgSO4 and 50 mg C18. This mixture was vortexed and centrifuged at 3000 g for 5 min, and then the upper layer was evaporated to dryness under a stream of nitrogen. The residue was dissolved in 40 µl ethyl acetate, and a 1-µl aliquot was injected into the GC-MS/MS. The GC separation of the compounds was achieved on a fused-silica capillary column Rxi-5Sil MS (30 m × 0.25 mm i.d.; 0.25-µm film thickness) and detected by MS/MS operating in electron ionization ion source mode. The regression equations showed excellent linearity (r > 0.9997) from 50 to 5000 ng/ml for plasma, with limit of detection of 10 ng/ml. The extraction efficiency of valproate for plasma ranged between 71.2%-103.5%. The coefficient of variation was <18.5%. The method was successfully applied to actual analyses of an autopsy case. This method can be useful for simple and reliable measurements of valproate in clinical and toxicological analyses; it can be integrated in screening and simultaneous determination methods for multiple drugs and poisons in the further studies.

Rapid simultaneous capillary electrophoretic determination of (R)- and (S)-secobarbital from serum and prediction of hydroxypropyl-gamma-cyclodextrin-secobarbital stereoselective interaction using molecular mechanics simulation.

Secobarbital is a hypnotic sedative that is used in the treatment of preoperative anxiety and to manage elevated intracranial pressures and cerebral ischemia due to neurosurgical procedures. Stereoselective resolution is first predicted using hydroxypropyl-gamma-cyclodextrin (HPGCD)-(R)- and (S)-secobarbital transient complex energy calculations using SYBYL 6.01 molecular modeling software. Separation is accomplished as predicted using 40 mM HPGCD in 50 mM phosphate buffer (pH 9.0), resulting in a rapid, sensitive method for the determination of (S)- and (R)-enantiomers of secobarbital from serum using solid-phase extraction, capillary electrophoresis (CE), and ultraviolet detection. The method involves extraction of both enantiomers and the internal standard, aprobarbital, from serum using C18 Bond-Elut solid-phase cartridges. The CE system consists of fused-silica capillary (52 cm x 75-microns i.d.) maintained at a run voltage of 15 kV with detection performed at a wavelength of 254 nm. The detection and quantitation limits from serum for (S)- and (R)-secobarbital are 1 microgram/mL. Linear calibration curves from 1 to 60 micrograms of both (S)- and (R)-secobarbital show a coefficient of determination of more than 0.999. The precision and accuracy of the method, calculated as relative standard deviation (%) and percent error, are 0.35-4.48% and 0.71-8.67%, respectively, for (R)-secobarbital and 0.39-4.08% and 2.16-3.70%, respectively, for (S)-secobarbital.