Metabolic Phenotyping Using UPLC–MS and Rapid Microbore UPLC–IM–MS: Determination of the Effect of Different Dietary Regimes on the Urinary Metabolome of the Rat

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 dietspecific 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.

The Pharmacometabodynamics of Gefitinib after Intravenous Administration to Mice: A Preliminary UPLC–IM–MS Study

“…Like many others [39][40][41], we have observed that the addition of IM provides many advantages in the characterization of metabolites in untargeted metabolic phenotyping [42][43][44].…”

Recurrent Topics in Mass Spectrometry-Based Metabolomics and Lipidomics—Standardization, Coverage, and Throughput

“…In metabolomics, the sensitivity gain provided by micro-LC has been exploited to design rapid separations for metabolic phenotyping. Throughput has been optimized at the expense of chromatographic performance, providing fit for purpose platforms with enhanced but not maximized sensitivity upon miniaturization. − A systematic comparison to conventional HPLC methods in terms of LOD and LLOQ was beyond the scope of these studies. Short micro-ultraperformance (UPLC) type of separation utilizing RP materials with sub-2-μm particles (100% wettable, 1.0 × 50 mm, 1.7 μm) and separation times of 2.5 min was successfully applied in large scale phenotyping studies and recently explored in combination with ion mobility. , A rapid micro-HILIC method utilizing sub-2-μm particles (Acquity UPLC BEH-Amide, 1.0 × 50 mm, 1.7 μm) addressed the analysis of polar metabolites in rat urine in less than 3.5 min.…”

“…Throughput has been optimized at the expense of chromatographic performance, providing fit for purpose platforms with enhanced but not maximized sensitivity upon miniaturization. − A systematic comparison to conventional HPLC methods in terms of LOD and LLOQ was beyond the scope of these studies. Short micro-ultraperformance (UPLC) type of separation utilizing RP materials with sub-2-μm particles (100% wettable, 1.0 × 50 mm, 1.7 μm) and separation times of 2.5 min was successfully applied in large scale phenotyping studies and recently explored in combination with ion mobility. , A rapid micro-HILIC method utilizing sub-2-μm particles (Acquity UPLC BEH-Amide, 1.0 × 50 mm, 1.7 μm) addressed the analysis of polar metabolites in rat urine in less than 3.5 min. Comparison to conventional HILIC-MS demonstrated 4-fold reduction of analysis time, 75% reduction in solvent consumption, and 18-fold reduction of sample consumption, while providing sufficient retention of polar metabolites (e.g.…”