Emprego da eletroextração multifásica para extração de antidepressivos tricíclicos em fluido oral e determinação por UHPLC-MS/MS e PS-MS

Abstract

Tricyclic antidepressants (TCAs) are substances used for the treatment of anxiety disorders and depression. And despite its effectiveness, have many side effects. Due to this and also to narrow therapeutic window of these drugs it is necessary to its determination in biological samples. For this reason, a multiphase multiwell plate for electroextraction was employed to extract the tricyclic antidepressants: doxepin, imipramine, amitriptyline, nortriptyline, and clomipramine from oral fluid. Afterwards the extract was analyzed by ultra-high liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) and also by paper spray mass spectrometry analysis. The effect of electric field in the extraction was evaluated since it is a crucial factor to achieve good recovery of TCAs. The analytes were extracted from donor solution consisted of a mixture of a sample and acetonitrile 1:1 (v/v), across the free liquid membrane made of 1-octanol, arriving in the acceptor phase. This last one was constituted of chromatographic paper embedded in acetic acid 400 mmol L-1 with a triangular tip suitable to PS-MS technique. Univariate studies were carried out to optimize the parameters: desorption of analytes from acceptor phase, chromatography paper and electrolyte types used in the acceptor phase, the organic solvent used as organic filter, presence of carrier on the organic filter and organic solvent type used in the donor phase. A multivariate optimization was performed to evaluate the extraction time, applied electric potential difference and the solvent percentage in the donor phase. After optimization, extraction efficiency was about 55% with 10 minutes of extraction at 300 V and 50% (v/v) of acetonitrile in the donor phase. The optimized EE-UHPLC-MS/MS method was validated in the range 2 – 12 ng mL-1 to TCAs, with LD was 0,13 to 0,25 and LQ was 0,44 to 0,84. The precision and accuracy were assessed at three concentration levels (2, 6 and 10 ng mL) with relative standard deviations under 26% and recovery higher than 75%. The method validated was applied to real fortified samples with low variation among different sample sources. The method was selective and unaffected by matrix effect. A new electroextraction configuration has been tested in an attempt to reduce variation sources on the paper position of the acceptor phase and to easily and directly couple electroextraction to paper spray mass spectrometry. Using this system, the extraction efficiency was about 35%. And the relative standard deviations were lower than 20% in the analysis by PS-MS.