Rapid Determination of 13 Sedative Substances and Their Metabolites in Blood by LC-MS/MS

Objective To establish an analysis method for simultaneous determination of 13 sedative substances and their metabolites in blood by liquid-liquid extraction and liquid chromatography-tandem mass spectrometry （LC-MS/MS） technology and to apply the method to actual cases. Methods The samples were extracted with ethyl acetate after an internal standard was added. The extract was condensed until it was nearly dry and then its residues were dissolved with methanol, filtered through 0.22 μm filter and finally determined. The 13 sedative substances and their metabolites were separated through the C₁₈ chromatographic column, then gradient elution was performed on them with methanol and 20 mmol/L ammonium formate （containing 0.1% formic acid） solution. After that, they were determined in the electrospray positive ion mode and quantified by internal standard method. Results The 13 sedative substances and their metabolites in blood showed good linearity in the range of 5-200 μg/L with correlation coefficients ranging from 0.990 3 to 0.999 8. The detection limits were 0.1-1.0 μg/L. Recovery rates of sedative substances were in the range of 71.2%-93.4% when solutions with concentrations of 10, 50 and 200 μg/L were added. The deviations of intra-day and inter-day relative standard deviations （RSD） were not more than 8.6%. Accuracies （bias） were within ±9.8%. Conclusion This method is rapid, simple, effective and sensitive, and can be applied to analysis of 13 sedative substances and their metabolites in blood in forensic toxicology.