Determination of nerve agent biomarkers in human urine by a natural hydrophobic deep eutectic solvent-parallel...

Alkyl methyl phosphonic acids (AMPAs) are the major metabolites of organophosphorus nerve agents. A method based on the use of natural hydrophobic deep eutectic solvents as supported liquid membrane in parallel artificial liquid microextraction (PALME) combined with LC-MS/MS analysis was developed and applied to their extraction from urine samples. PALME is a miniaturized liquid-phase extraction method performed in a multiwell plate format where the aqueous sample and the aqueous acceptor phase are separated by a flat membrane impregnated with an organic solvent. In this study, we investigated the possibility of replacing the harmful conventional organic solvent by an emerging green solvent, a coumarin/thymol-based deep eutectic solvent, in ordered to raise the greenness of the sample preparation method. Linear response was obtained in an interval of 0.5, 5 or 10-100 ng/ml depending on the AMPAs with a determination coefficients (R²s) ranging from 0.9751 to 0.9989 for their determination in not treated urine samples. Enrichment factors (EFs) up to 12.65 were obtained, and repeatability was within 8.90-16.28% RSD (n = 12). The limit of quantifications (LOQs: S/N ≥ 10) of the whole analytical procedure were in the range from 0.04 to 5.35 ng/ml. In addition to its good sensitivity, the presented method permitted the treatment of 192 samples in 120 min (equivalent to 37.5 s/sample), which places it as one of the most powerful preparation technique for biomonitoring of civilian or military people exposed to nerve agents in case of public health emergency. Indeed, the developed procedure combined sensitivity, high-throughput, greenness, simplicity and practicality for the determination of five acidic polar AMPAs in urine samples.