最近,我们的研究小组已经表明,芬太尼及其许多类似物在电喷雾电离过程中形成原位异构体(\"原位体\").这些不同的原聚体可以使用离子迁移谱来解析,并使用迁移对齐的串联质谱片段进行注释。然而,它们的形成以及实验变量对它们的相对比率的贡献程度仍然知之甚少。在本研究中,我们系统地研究了常见色谱溶剂(水,甲醇,和乙腈)和pH值取决于先前观察到的23种芬太尼类似物的前药的比例。有趣的是,这些比率(N-哌啶质子化与仲胺/O=质子化)对于许多类似物显著降低(例如,去丙酰基邻位,元-,和对甲基芬太尼),对其他人来说显著增加(例如,顺式-异芬太尼),当溶剂条件从100%有机溶剂(甲醇或乙腈)变为100%水时,其余的保持相对恒定。有趣的是,pH值对该比例也有显著影响,在许多情况下导致切换比率的变化。最后,将条件增加到pH≥4.0也促使邻-和对-甲基乙酰芬太尼出现新的迁移率峰,所有以前的研究只显示一个单一的分布。因为这些比率有望定性地用于鉴定这些(和新兴的)芬太尼类似物,了解各种条件(即,流动相选择和/或色谱梯度)影响它们的比率对于开发先进的离子迁移和质谱方法以鉴定芬太尼类似物至关重要。
Recently, our group has shown that fentanyl and many of its analogues form prototropic isomers (\"protomers\") during electrospray ionization. These different protomers can be resolved using ion mobility spectrometry and annotated using mobility-aligned tandem mass spectrometry fragmentation. However, their formation and the extent to which experimental variables contribute to their relative ratio remain poorly understood. In the present study, we systematically investigated the effects of mixtures of common chromatographic solvents (water, methanol, and acetonitrile) and pH on the ratio of previously observed protomers for 23 fentanyl analogues. Interestingly, these ratios (N-piperidine protonation vs. secondary amine/O = protonation) decreased significantly for many analogues (e.g., despropionyl ortho-, meta-, and para-methyl fentanyl), increased significantly for others (e.g., cis-isofentanyl), and remained relatively constant for the others as solvent conditions changed from 100% organic solvent (methanol or acetonitrile) to 100% water. Interestingly, pH also had significant effects on this ratio, causing the change in ratio to switch in many cases. Lastly, increasing conditions to pH ≥ 4.0 also prompted the appearance of new mobility peaks for ortho- and para-methyl acetyl fentanyl, where all previous studies had only showed one single distribution. Because these ratios have promise to be used qualitatively for identification of these (and emerging) fentanyl analogues, understanding how various conditions (i.e., mobile phase selection and/or chromatographic gradient) affect their ratios is critically important to the development of advanced ion mobility and mass spectrometry methodologies to identify fentanyl analogues.