The comprehended knowledge of the metabolic profile of the fecal matter has been recognized as an important point for understanding metabolic changes in the human systemic metabolism and it can provide precious information about host-gut microbiota interactions. However, few analytical strategies have been addressed for a broad analysis of metabolites with different chemical properties to better understand the chemical space of fecal samples. Here we report a systematic pipeline to achieve comprehensive coverage of the fecal metabolome, from high polar to nonpolar metabolites, using dog fecal samples as a proof-of-concept. This pipeline comprises a monophasic (ACN/H2O) and a biphasic extraction (methyl tert-butyl ether (MTBE)/MeOH/H2O) of the sample, followed by three liquid chromatography-high resolution tandem mass spectrometry (LC-HRMS/MS) methods using HILIC-amide, RP-C18 and CSH-C18 columns, and a switch polarity acquisition mode in the electrospray ion source. This approach allowed the annotation of 376 metabolites from 70 different chemical classes. The chemical space analysis by molecular networking and the pathway analysis revealed the complexity of the fecal sample and the importance of combined methods to better understand biochemical pathways. This pipeline can be used as a valuable tool to comprehend the relationship between host-gut microbiota metabolites and the influence of diet, medication, or environmental changes.
A diferenciação taxonômica de espécies tem sido cada vez mais suportada por análises de amostras fecais e abordagens moleculares. A partir da análise semiquantitativa de ácidos biliares fecais de cinco mustelídeos neotropicais acessamos a diversidade destes compostos, avaliando seu potencial na discriminação taxonômica de espécies. Amostras fecais coletadas de espécimes cativos foram analisadas por cromatografia líquida acoplada a espectrômetro de massas de alta performance. Diferenças significativas em quatro ácidos biliares foram detectadas entre as espécies analisadas. Os ácidos biliares são compostos endógenos fundamentais em vias bioquímicas, assim detectar diferenças interespecíficas pode ser relevante para o conhecimento da fisiologia, estado de saúde e respostas metabólicas dos mustelídeos às mudanças ambientais.
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