title: Comparison of capillary electrophoresis-mass spectrometry and hydrophilic interaction chromatography-mass spectrometry for anionic metabolic profiling of urine.
abstract: In order to assess the utility of a recently developed capillary electrophoresis-mass spectrometry (CE-MS) method for the study of anionic metabolites in urine, a comparison was made with hydrophilic interaction chromatography-MS (HILIC-MS) using negative electrospray ionization. After optimization of the HILIC conditions, a gradient employing 10mM ammonium acetate (pH 6.8) in acetonitrile-water (5 min 90% acetonitrile followed by 90%-50% acetonitrile in 10 min) was selected, providing baseline separation of five representative anionic test metabolites. Relative standard deviations (RSDs) for HILIC retention times and peak areas were below 0.2% and 7.7%, respectively, and detection limits were in the range 0.04-2.21 μM. Metabolites in rat urine could also be analysed in a reproducible way with retention time and peak area RSDs below 0.6% and 13.6%, respectively. The CE-MS and HILIC-MS methods were compared in terms of reproducibility, sensitivity, selectivity and coverage of the anionic urinary metabolome. In general, peak area RSDs were similar whereas HILIC-MS yielded better retention-time repeatability and up to 80 times lower detection limits (expressed in injected concentration) for test metabolites as compared to CE-MS. Rat urine analysis by HILIC-MS provided detection of 1360 molecular features compared to 347 molecular features revealed with CE-MS. Of these, a number of 144 molecular features were found with both HILIC-MS and CE-MS, which showed on average 10 times higher peak areas in HILIC-MS. The HILIC retention and CE migration times of the common features were clearly not correlated. The HILIC and CE behavior of the test metabolites and 16 putatively identified common features were evaluated involving their physicochemical properties, indicating a markedly different separation selectivity, and thus significant degree of orthogonality of HILIC and CE.
authors.affiliation: Biomolecular Analysis, Department of Pharmaceutical Sciences, Utrecht University, P.O. Box 80082, 3508 TB Utrecht, The Netherlands; Research group Analysis Techniques in Life Sciences, Avans Hogeschool, P.O. Box 90116, 4800 RA Breda, The Netherlands. Electronic address: M.G.M.Kok@uu.nl.
authors.forename: Miranda G M
authors.initials: MG
authors.lastname: Kok
authors.affiliation: Biomolecular Analysis, Department of Pharmaceutical Sciences, Utrecht University, P.O. Box 80082, 3508 TB Utrecht, The Netherlands; AIMSS Division of BioAnalytical Chemistry, Department of Chemistry and Pharmaceutical Sciences, VU University, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands.
authors.forename: Govert W
authors.initials: GW
authors.lastname: Somsen
authors.affiliation: Biomolecular Analysis, Department of Pharmaceutical Sciences, Utrecht University, P.O. Box 80082, 3508 TB Utrecht, The Netherlands; Research group Analysis Techniques in Life Sciences, Avans Hogeschool, P.O. Box 90116, 4800 RA Breda, The Netherlands.
authors.forename: Gerhardus J
authors.initials: GJ
authors.lastname: de Jong
mesh_terms: D000085:Acetates; D000097:Acetonitriles; D000818:Animals; D000838:Anions / Q000652:urine*; D019075:Electrophoresis, Capillary / Q000295:instrumentation* / Q000379:methods / Q000592:standards; D057927:Hydrophobic and Hydrophilic Interactions; D057230:Limit of Detection; D013058:Mass Spectrometry / Q000295:instrumentation* / Q000379:methods / Q000592:standards; D055442:Metabolome / Q000502:physiology; D051381:Rats; D012015:Reference Standards; D021241:Spectrometry, Mass, Electrospray Ionization / Q000295:instrumentation / Q000379:methods; D016482:Urinalysis / Q000295:instrumentation* / Q000379:methods / Q000592:standards; D014867:Water
chemical_list: D000085:Acetates; D000097:Acetonitriles; D000838:Anions; D014867:Water; C018824:ammonium acetate; C032159:acetonitrile
keywords: Anionic metabolic profiling; Capillary electrophoresis; Hydrophilic interaction chromatography; Mass spectrometry; Urine