High-throughput identification of gut microbiome-dependent metabolites.

A significant hurdle that has limited progress in microbiome science has been identifying and studying the diverse set of metabolites produced by gut microbes. Gut microbial metabolism produces thousands of difficult-to-identify metabolites, which present a challenge to study their roles in host biology. In recent years, mass spectrometry-based metabolomics has become one of the core technologies for identifying small metabolites. However, metabolomics expertise, ranging from sample preparation to instrument use and data analysis, is often lacking in academic labs. Most targeted metabolomics methods provide high levels of sensitivity and quantification, while they are limited to a panel of predefined molecules that may not be informative to microbiome-focused studies. Here we have developed a gut microbe-focused and wide-spectrum metabolomic protocol using liquid chromatography-mass spectrometry and bioinformatic analysis. This protocol enables users to carry out experiments from sample collection to data analysis, only requiring access to a liquid chromatography-mass spectrometry instrument, which is often available at local core facilities. By applying this protocol to samples containing human gut microbial metabolites, spanning from culture supernatant to human biospecimens, our approach enables high-confidence identification of >800 metabolites that can serve as candidate mediators of microbe-host interactions. We expect this protocol will lower the barrier to tracking gut bacterial metabolism in vitro and in mammalian hosts, propelling hypothesis-driven mechanistic studies and accelerating our understanding of the gut microbiome at the chemical level.

Duke Scholars

Author Shuo Han Biochemistry