Accepted: U. Mons

Introduction

Saponins are amphiphilic molecules of pharmaceutical interest. Saponins correspond to the combination of a hydrophilic sugar-containing glycone linked to a hydrophobic triterpenoid/steroid aglycone. Due to their huge diversity, their structural characterization remains challenging and the development of efficient methods is a perpetual task. Structure elucidation of saponins by mass spectrometry is usually performed using LC-MS(MS). Recently, we introduced ion-mobility mass spectrometry for saponin characterization. Differences in CCS between isomeric saponins are often too small to allow the gas phase discrimination using ion-mobility on standard instruments. However, the successful gas phase separation between stereoisomeric and regioisomeric saponin ions has been achieved by using cyclic-ion-mobility thanks to the flexible ion manipulation experiments (ion selection by IM , pre/post-IM ion storage and fragmentation).

Methods

We selected the Horse Chestnuts saponins that are a mixture of triterpenoid saponins. These molecules are based on a protoaescigenin substituted by different side chains. The Escin family counts several molecules that are mostly distinguished based on the non-glycosidic side chains. Amongst them, tiglic acid and angelic acid residues are of prime interest for the present study since they are only differentiated based on the cis-trans configuration of the C=C bond. Saponin isomers were extracted from the HC seeds using a standard protocol developed in our lab and the corresponding ions, generated upon electrospray ionization, were submitted to ion mobility on two different instruments; a Waters Synapt G2-Si mass spectrometer and a cyclic-ion-mobility enabled quadrupole time-of-flight (Q-cIM-oaToF) mass spectrometer.

Preliminary Data

The HC saponin extract is particularly challenging from an analytical point of view since regioisomeric and stereoisomeric saponins are present. In the present communication, we tested ion mobility experiments together with liquid chromatography separation for the structural characterization of stereoisomeric and regioisomeric saponins. We considered all the different types of ions that can be generated upon electrospray ionization of a given molecule, namely the common [M+H]+, [M+Na]+ and [M-H]-. We also used more exotic ionic species such as [2M-2H]2- ions to distinguish isomeric saponins. On Waters Synapt G2-Si mass spectrometer, saponins presenting isomeric side chains, such as tiglic and angelic acid residues, are hardly distinguished by recording the Arrival Time Distributions (ATD) due to the lack of ion mobility resolution. The slight differences in ATD are only detected when the saponin isomers are first separated using liquid chromatography. This points to a need for higher ion mobility resolution to tackle small structural differences for both configurational and positional isomers. We then submitted the crude extract, without any liquid chromatography separation, to ion mobility experiments on the cyclic IM enabled instrument. By associating mass selection, collision-induced dissociation and multi-pass ion mobility in a defined sequence of ion manipulation, we successfully distinguished steroisomeric saponins with a baseline separation. As a promising perspective, we detected and characterized six new HC saponins by associating liquid chromatography, ion mobility and high resolution mass spectrometry data.

Novel Aspect
Cyclic-ion-mobility setup provides higher mobility resolution, IMn (fragmentation/spectrum clean-up/ion manipulation capabilities), useful for stereoisomeric and regioisomeric discrimination, such as saponins.
Options:

A post-doc is presenting author on this abstract? Yes
A graduate student is presenting author on this abstract? No
An undergraduate student is presenting author on this abstract? No

Poster:

Ion Mobility: Applications

Submitting Author:

Julien De Winter
University of Mons
Mons,
julien.dewinter@umons.ac.be