Metabolic Phenotyping Using UPLC\u2013MS and Rapid Microbore UPLC\u2013IM\u2013MS: Determination of the Effect of Different Dietary Regimes on the Urinary Metabolome of the Rat

Marine Letertre,Susan E Slade,Nyasha C Munjoma,Ian D Wilson,Muireann Coen,Jeremy K Nicholson,Jon Swann,Robert S Plumb

doi:10.1007/s10337-020-03900-4

Abstract

A rapid reversed-phase gradient method employing a 50 mm × 1 mm i.d., C18 microbore column, combined with ion mobility and high-resolution mass spectrometry, was applied to the metabolic phenotyping of urine samples obtained from rats receiving different diets. This method was directly compared to a “conventional” method employing a 150 × 2.1 mm i.d. column packed with the same C18 bonded phase using the same samples. Multivariate statistical analysis of the resulting data showed similar class discrimination for both microbore and conventional methods, despite the detection of fewer mass/retention time features by the former. Multivariate statistical analysis highlighted a number of ions that represented diet-specific markers in the samples. Several of these were then identified using the combination of mass, ion-mobility-derived collision cross section and retention time including N-acetylglutamate, urocanic acid, and xanthurenic acid. Kynurenic acid was tentatively identified based on mass and ion mobility data.

Highlights

Metabolic phenotyping [1–3], using methods based on liquid chromatography (LC), of the type routinely undertaken during metabonomic/metabolomic investigations faces a number of challenges connected withElectronic supplementary material The online version of this article contains supplementary material, which is available to authorized users.MA 01757, USA 4 Oncology Safety, Clinical Pharmacology and SafetySciences, R&D, AstraZeneca, Cambridge CB4 0WG, UK 5 Australian National Phenome Centre, Murdoch University, Harry Perkins Building, Perth, WA, Australia both throughput and metabolite identification
As a partial solution to some of these problems we recently introduced the concept of rapid microbore metabolic profiling (RAMMP) using reversed-phase ultra performance liquid chromatography (U(H)PLC) on a 1 mm i.d. column, coupled to mass spectrometric detection (MS) [4] and, more recently, similar rapid HILIC and RP-lipidomic methods have been described [5, 6]
As indicated in the experimental section, the urine samples obtained from these animals were analysed using both conventional UPLC–MS [14] and RAMMP–ion mobility (IM)–MS

Summary

Introduction

As a partial solution to some of these problems we recently introduced the concept of rapid microbore metabolic profiling (RAMMP) using reversed-phase ultra (high) performance liquid chromatography (U(H)PLC) on a 1 mm i.d. column, coupled to mass spectrometric detection (MS) [4] and, more recently, similar rapid HILIC and RP-lipidomic methods have been described [5, 6]. As would be expected, the peak capacity of the RAMMP method was reduced (from 150 to 50) compared to analysis on a conventional 2.1 mm i.d., and the number of measured features was decreased from ca. Notwithstanding the loss of peak capacity and reduced number of features the RAMMP methodology was still able to achieve similar levels of group discrimination by PCA,

Methods

Results

Conclusion