title: Sample preparation optimization in fecal metabolic profiling.
abstract: Metabolomic analysis of feces can provide useful insight on the metabolic status, the health/disease state of the human/animal and the symbiosis with the gut microbiome. As a result, recently there is increased interest on the application of holistic analysis of feces for biomarker discovery. For metabolomics applications, the sample preparation process used prior to the analysis of fecal samples is of high importance, as it greatly affects the obtained metabolic profile, especially since feces, as matrix are diversifying in their physicochemical characteristics and molecular content. However there is still little information in the literature and lack of a universal approach on sample treatment for fecal metabolic profiling. The scope of the present work was to study the conditions for sample preparation of rat feces with the ultimate goal of the acquisition of comprehensive metabolic profiles either untargeted by NMR spectroscopy and GC-MS or targeted by HILIC-MS/MS. A fecal sample pooled from male and female Wistar rats was extracted under various conditions by modifying the pH value, the nature of the organic solvent and the sample weight to solvent volume ratio. It was found that the 1/2 (wf/vs) ratio provided the highest number of metabolites under neutral and basic conditions in both untargeted profiling techniques. Concerning LC-MS profiles, neutral acetonitrile and propanol provided higher signals and wide metabolite coverage, though extraction efficiency is metabolite dependent.
authors.affiliation: Department of Chemistry, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Olga
authors.initials: O
authors.lastname: Deda
authors.affiliation: Department of Chemistry, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Anastasia Chrysovalantou
authors.initials: AC
authors.lastname: Chatziioannou
authors.affiliation: Department of Chemistry, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Stella
authors.initials: S
authors.lastname: Fasoula
authors.affiliation: Department of Chemistry, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Dimitris
authors.initials: D
authors.lastname: Palachanis
authors.affiliation: Laboratory of Forensic Medicine and Toxicology, School of Medicine, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Νicolaos
authors.initials: Ν
authors.lastname: Raikos
authors.affiliation: Department of Chemistry, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.
authors.forename: Georgios A
authors.initials: GA
authors.lastname: Theodoridis
authors.affiliation: Laboratory of Forensic Medicine and Toxicology, School of Medicine, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece. Electronic address: gkikae@auth.gr.
authors.forename: Helen G
authors.initials: HG
authors.lastname: Gika
mesh_terms: D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods; D005243:Feces / Q000737:chemistry*; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008297:Male; D055442:Metabolome; D055432:Metabolomics / Q000379:methods*; D051381:Rats; D017208:Rats, Wistar; D013048:Specimen Handling / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods
keywords: Extraction optimization; Feces; Metabolic profiling; Metabolomics; Sample preparation