[Enzyme Digestion LC-MS/MS Detection of Etomidate, Ketamine, Methamphe‑ tamine and Codeine in Hair].

Abstract

Objectives: To establish a liquid chromatography-tandem mass spectrometry (LC-MS/MS)method based on enzymatic digestion for the rapid qualitative and quantitative detection of drugs in hair.

Methods: The enzymatic digestion parameters for etomidate, ketamine, methamphetamine, and codeine, including dithiothreitol (DTT) concentration, proteinase K concentration, incubation temperature and digestion time were optimized using controlled-variable experiments. After digestion, hair samples were filtered through a 0.22 μm membrane and separated on a C18 column with a gradient elution using 0.1% formic acid in water and 0.1% formic acid in acetonitrile and analyzed using electrospray ionization (ESI) in multiple reaction monitoring (MRM) mode. After validation, the method was applied to 155 drug-related hair samples and compared with the cryogenic wet-grinding method.

Results: Etomidate, ketamine, methamphetamine, and codeine in hair samples all showed good linearity with their respective linear ranges (R²>0.998 0), with limits of detection (LOD) of 0.005-0.010 ng/mg and limits of quantification (LOQ) of 0.02-0.04 ng/mg. Matrix effects ranged from -12.09% to 8.23%, while the relative standard deviations of intra-day precision and inter-day precision were 2.56%-10.37% and 5.50%-15.11%, respectively. Average recoveries (88.97%-104.05%) met analytical requirements. Results from drug-related hair samples suggested good consistency between enzymatic digestion and cryogenic grinding pretreatment method, while enzymatic digestion demonstrated higher extraction amounts for codeine than cryogenic grinding pretreatment method.

Conclusions: The established method is simple to operate, highly sensitive, and reproducible, making it suitable for high-throughput hair drug analysis, providing an efficient and reliable analytical method for forensic identification.