Determination of gamma-hydroxybutyric acid in biofluids using a one-step procedure with “in-vial” derivatization and headspace-trap gas chromatography–mass spectrometry

A headspace-trap gas chromatography–mass spectrometry (HS-trap GC–MS) method was developed to determine GHB, a low molecular weight compound and drug of abuse, in various biological fluids. Combining this relatively novel and fully automated headspace technique with “in-vial” methylation of GHB allowed for a straightforward approach. One single method could be used for all biofluids (urine, plasma, serum, whole blood or lyzed blood), requiring only 100 μl of sample. Moreover, our approach involves mere addition of all reagents and sample into one vial. Following optimization of headspace conditions and trap settings, validation was performed. Although sample preparation only consists of the addition of salt and derivatization reagents directly to a 100 μl-sample in a HS-vial, adequate method sensitivity and selectivity was obtained. Calibration curves ranged from 5 to 150 μg/ml GHB for urine, from 2 to 150 μg/ml for plasma, and from 3.5 to 200 μg/ml for whole blood. Acceptable precision and accuracy (<13% bias and imprecision) were seen for all quality controls (QCʹs) (LLOQ-level, low, medium, high), including for the supplementary serum- and lyzed blood-based QCʹs, using calibration curves prepared in plasma or whole blood, respectively. Incurred sample reanalysis demonstrated assay reproducibility, while cross-validation with another GC–MS method demonstrated that our method is a valuable alternative for GHB determination in toxicological samples, with the advantage of requiring only 100 μl and minimal hands-on time, as sample preparation is easy and injection automated.