Determination of gamma-hydroxybutyric acid in dried blood spots using a simple GC-MS method with direct on spot derivatization

The objective of this study was the development of an accurate and sensitive method for the determination of gamma-hydroxybutyric acid (GHB) in dried whole blood samples using a GC-MS method. The complete procedure was optimized, with special attention on the sample pretreatment, and validated. Therefore, dried blood spots of only 50 mu l were prepared and, after the addition of internal standard GHB-d6, directly derivatized using 100 mu l of a freshly prepared mixture of trifluoroacetic acid anhydride and heptafluorobutanol (2:1). The derivatized extract was injected into a gas chromatograph coupled to a mass spectrometer (GC-MS), operating in the electron impact mode, with a total run time of 12.3 min. Method validation included the evaluation of linearity, precision, accuracy, sensitivity, selectivity, and stability. A weighting factor of 1/x (2) was chosen and acceptable intra-batch precision, inter-batch precision, and accuracy were seen. The linear calibration curve ranged from 2 to 100 mu g/ml, with a limit of detection of 1 mu g/ml. Our procedure, utilizing the novel approach of direct on spot derivatization followed by analysis with GC-MS, proved to be reliable, fast, and applicable in routine toxicology.