Gas chromatographic assay for free and total plasma levels of thiopental.

A rapid gas chromatographic assay for the determination of free and total plasma thiopental is described. Free thiopental was obtained by ultrafiltration through Amicon Centroflo membrane cones. Gas chromatographic assay utilized secobarbital as an internal standard and employed on-column methylation of the barbiturates to improve peak resolution. In 73 blood samples from 22 patients total thiopental concentrations ranged from 4.2 to 134 mug/ml plasma, with a mean of 28 mug/ml. Free thiopental values ranged from 8.6 to 22.7 per cent of total, with a mean of 13.7 per cent free thiopental and a standard deviation of 3.2 per cent. At a total thiopental level of 10 mug/ml, unbound thiopental averaged 10.7 per cent with ultrafiltration, compared with 11.5 per cent with equilibrium dialysis. Assays of thiopental by gas chromatography and 14C scintillation counting gave similar results. There were progressive increases in the percentages of thiopental that were unbound when thiopental was added to plasma, purified crystalline albumin (4.5 g/l), and normal serum albumin (5 g/l), and a solution of purified protein fractions (5 g/l). Differences in protein binding determined by this method and previously reported methods are discussed.