Investigation of the acid/base behaviour of the opium alkaloid thebaine in LC-ESI-MS mobile phase by NMR spectroscopy

As part of a research program to establish an analytical method for the simultaneous detection of the five major opium alkaloids in poppy seeds by liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) it was discovered that the inclusion of thebaine produced two peaks for the same compound. This was in contrast to the effective simultaneous detection, by LC-ESI-MS, of morphine, codeine, papaverine and noscapine. The presence of these two peaks for thebaine was investigated using nuclear magnetic resonance (NMR) spectroscopy with deuterated solvents to emulate the mobile phase conditions experienced. It was found that the presence of 80%, or higher ratios of, water caused two epimeric forms of thebaine to be formed; this explained the presence of two peaks on the chromatogram. In contrast, when a lower water content was used with 1% acetic acid, one stable form of thebaine could be analyzed and resulted in a single peak visible in the subsequent chromatography.

本研究作为建立液相色谱-电喷雾电离-质谱法（LC-ESI-MS）同时检测罂粟籽中5种主要鸦片生物碱的分析方法的科研计划的一部分，实验中发现，在检测体系中加入蒂巴因时，该化合物会出现两个色谱峰。这与利用LC-ESI-MS对吗啡、可待因、罂粟碱及那可丁实现的高效同时检测结果形成显著差异。针对蒂巴因出现双峰的现象，本研究采用氘代溶剂模拟色谱流动相的实际条件，借助核磁共振（NMR）光谱法开展了系统探究。结果显示，当体系中水的占比达到80%或更高时，蒂巴因会形成两种差向异构体，这一发现成功解释了色谱图中出现双峰的原因。与之相反，当使用含水量更低且添加1%乙酸的体系时，蒂巴因可保持单一稳定构型，经后续色谱分析后仅呈现单个色谱峰。