Separation of Cucurbit[7]uril Complexation Protomer by Ion Mobility Spectrometry-Mass Spectrometry

Abstract

Protonation of molecules can influence properties such as functions or binding affinity in the physiological environment. Among them, the characteristics of molecules with multiple protonation sites can vary depending on the protonation site. However, distinguishing protomers of small molecules is not easy because they often have insignificant structural differences, and protons are labile. In this study, the protomers of small molecules with multiple protonation sites were separated by introducing cucurbit[7]uril (CB[7]). The small molecule used in this study is imipramine, which contains two nitrogen atoms. Since it binds differently to cucurbituril according to the protonation site, it was possible to separate the protomers of the complexes and confirm the protonation sites using ion mobility spectrometry-mass spectrometry (IMS-MS). Singly and doubly protonated CB[7]-imipramine complexes were observed, and the protonation sites of each protomer were assigned. We confirmed that protomers in doubly protonated complexes are distinguishable in IMS. Through this research, we demonstrate the use of cucurbituril's host property to separate protomers using IMS-MS. Additionally, we expand the application of IMS-MS by showing that it can be used as a tool to confirm protomers of host-guest complexes.