Pharmaceutical Analysis, Institute of Pharmacy, Freie Universität Berlin, Königin-Luise-Straße 2+4, 14195 Berlin, Germany.
Pharmaceutical and Medicinal Chemistry (Computer-Aided Drug Design), Institute of Pharmacy, Freie Universität Berlin, Königin-Luise-Straße 2+4, 14195 Berlin, Germany.
Molecules. 2023 Oct 21;28(20):7206. doi: 10.3390/molecules28207206.
The aim of this study was to develop and optimize a chiral HPLC-MS/MS method for quantitative analysis of ()-/()-salbutamol and ()-/()-salbutamol-4'--sulfate in human urine to allow for bioanalytical quantitation of the targeted analytes and investigations of stereoselectivity in the sulfonation pathway of human phase Ⅱ metabolism. For analytical method development, a systematic screening of columns and mobile phases to develop a separation via enantiomerically selective high performance liquid chromatography was performed. Electrospray ionization settings were optimized via multiple-step screening and a full factorial design-of-experiment. Both approaches were performed matrix-assisted and the predicted values were compared. The full factorial design was superior in terms of prediction power and knowledge generation. Performing a longitudinal excretion study in one healthy volunteer allowed for the calculation of excretion rates for all four targeted analytes. For this proof-of-concept, either racemic salbutamol or enantiopure levosalbutamol was administered perorally or via inhalation, respectively. A strong preference for sulfonation of ()-salbutamol for inhalation and peroral application was found in experiments. In previous studies phenol sulfotransferase 1A3 was described to be mainly responsible for salbutamol sulfonation in humans. Thus, in vitro and in silico investigations of the stereoselectivity of sulfotransferase 1A3 complemented the study and confirmed these findings.
本研究旨在开发和优化一种手性 HPLC-MS/MS 方法，用于定量分析人尿中的 ()-/()-沙丁胺醇和 ()-/()-沙丁胺醇-4'--硫酸盐，以实现对目标分析物的生物分析定量，并研究人Ⅱ相代谢中磺化途径的立体选择性。为了进行分析方法开发，系统地筛选了柱和流动相，以通过对映体选择性高效液相色谱法进行分离。通过多步筛选和完全析因实验设计优化了电喷雾电离条件。这两种方法都进行了基质辅助，并且比较了预测值。全因子设计在预测能力和知识生成方面具有优势。在一名健康志愿者中进行纵向排泄研究，允许计算所有四个目标分析物的排泄率。为此概念验证，分别通过口服或吸入给予外消旋沙丁胺醇或对映纯左沙丁胺醇。在实验中发现，吸入和口服应用时，（）-沙丁胺醇的磺化强烈偏向于磺化。在先前的研究中，酚磺基转移酶 1A3 被描述为人类沙丁胺醇磺化的主要负责酶。因此，磺基转移酶 1A3 的体外和计算立体选择性研究补充了这项研究，并证实了这些发现。
