The present study reports a validated automated solid-phase extraction (SPE) protocol associated to GC-MS/MS assay to quantify 5 drugs (caffeine, cyamemazine, meprobamate, morphine and 6-monoacetylmorphine (6-MAM)) in 11 biological matrices. Assessment of the protocol to be used as a starter set for the development of further analytical assays is demonstrated.

Analyses are performed on 200±5mg of tissue samples for lung, kidney, liver, muscle and bone marrow, 500±10mg for adipose tissue and brain, and 200μl for biological fluids (bile, blood, urine and vitreous humor). Automated SPE was performed using a Gx-271 ASPEC (Gilson Inc.) on Strata™ X-C (Phenomenex) SPE columns. The molecules were eluted in 2 different eluates, each analyzed by GCMS/ MS. A full validation was performed on BM, the most complex matrix, and partial validation on the other 10 matrices according to the validation protocol proposed by Cartiser et al. (J. Chrom. B, 2011). The sample preparation was assessed in blood, BM and lung for the analysis of 12 further compounds exhibiting varied physicochemical properties: (alimemazine, alprazolam, amitryptiline, citalopram, cocaine, diazepam, levomepromazine, nordazepam, tramadol, venlafaxine, pentobarbital, and phenobarbital).

Extraction recovery range between 5.3% for morphine in brain to 78.4% for caffeine in bile. The limits of quantification were 12.5ng/ml(ng/g) for 6-MAM, morphine and cyamemazine, 100ng/ml (ng/g) for meprobamate and 50ng/ml(ng/g) for caffeine. Precision and accuracy at the LOQ were respectively less than 20% and between 80% and 120% except for morphine in muscle (121.2%), meprobamate in kidney (138.6%), and cyamemazine in kidney (148.6%). None of the analytes demonstrated precision greater than 15% or accuracy outside of the 85–115% range in BM. Validation on the other matrices showed accuracy of 80–120% for all analytes, except for cyamemazine in kidney where accuracy was 135% at the high QC level. Precision was <15% for all parameters, except for 6-MAM and morphine in blood where it was slightly greater than 15% at high QC level (15.2% and 15.1%, respectively). The automated extraction protocol applied without modifications allows detection of the 12 further compounds at therapeutic concentrations in blood and in the alternative matrices.

An automated SPE protocol associated to GC-MS/MS analysis was developed for quantification of caffeine, cyamemazine, meprobamate, morphine and 6-MAM in 11 biological matrices. The performance of this assay allows application in forensic toxicology or pharmacokinetic studies. The extraction protocol can be used for targeted analysis of various basic drugs and, with optimization, for other drugs or screening procedures in complex matrices. It could thus be an interesting starting set for further development for laboratories seeking to optimize their activity and confronted to analysis of new compounds or new matrices.