Stable isotope labeling derivatization and magnetic dispersive solid phase extraction coupled with UHPLC-MS/MS for the measurement of brain neurotransmitters in post-stroke depression rats administrated with gastrodin

Abstract

Synchronous measurement of brain neurotransmitters and blood drug metabolism by dual sites in vivo microdialysis can provide very important information for several neurological disorders such as post-stroke depression (PSD). In this work, d₀/d₃-6-N-methyl-rhodamine 6G-N-hydroxysuccinimidyl formate (d₀/d₃-MRSF) were firstly designed, synthesized and used as stable isotope labeling derivatization reagents for the determination of multiple neurotransmitters by ultra-high performance liquid chromatography-tandem mass spectrometry. The light d₀-MRSF was used to label neurotransmitters in real samples and the heavy d₃-MRSF was used to label standards, which served as internal standards for quantification to minimize matrix effect. The d₀/d₃-MRSF-neurotransmitter derivatives were easily adsorbed onto the surface of Fe₃O₄@graphene oxide. Magnetic dispersive solid phase extraction followed with stable isotope labeling derivatization was developed for efficient sample pretreatment. The permanent positively charged moiety of d₀/d₃-MRSF significantly enhanced the ionization efficiency of neurotransmitters. The comparison results indicated that the synthesized MRSF derivatization method can bring better detection sensitivity than the commercial dansyl chloride method about tens to hundreds of times on the same instrumentation. Specific and regular product ions of m/z 413.3 and 416.3, m/z 429.4 and 432.4 were observed for d₀-and d₃-MRSF labeled neurotransmitters, respectively. The limits of detection were in the range of 2.0-40.0 pM. The accuracies ranged from 92.3% to 108.2% with the intra- and inter-day precisions in the range of 2.2-12.8%. Synchronous and dynamic determination of concentrations changes of brain neurotransmitters and blood gastrodin of PSD rats was achieved. The results indicated that gastrodin had analogous regulating effect of brain neurotransmitter, which like the clinical medicine fluoxetine of PSD. Taken together, the developed method was demonstrated to be promising for sensitive, accurate and simultaneous determination of multiple neurotransmitters in trace biofluids.

Keywords: In vivo microdialysis; Magnetic dispersive solid phase extraction; Neurotransmitters; Post-stroke depression; Stable isotope labeling derivatization; Triple quadrupole mass spectrometry.