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 μl were prepared and, after the addition of internal standard GHB-d6, directly derivatized using 100 μl 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 μg/ml, with a limit of detection of 1 μg/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.