Emerging Microbial Lipid Shifts in a Multistage Simulated Gut Inoculated with Human Fecal Microbiota

Abstract Food residues that bypass human digestion are further digested by gut microbes, leading to the production of diverse metabolites, including lipids. To investigate how lipids in our intestine are affected during this transition, we used a colon simulator with four distinct vessels (V1–V4) that mimic the proximal to distal part of the human colon. In total, 44 samples were collected from the colon simulator vessels (V1–V4). We observed dynamic shifts in a diverse array of microbially linked lipid molecules in the simulated intestinal chyme, including bile acids and N-acyl amides with short- and odd-chain lipids. Histamine-linked N-acyl lipids (histamine-C5) increased from the proximal to the distal colon vessels (pH 5.5–7.0), whereas putrescine-linked ones (putrescine-C19:2), initially abundant in the media, decreased across the colon vessels. We uncovered dynamic associations between in vitro-derived short-chain N-acyl lipids (C4:0, C5, C6, and C7 conjugates) and major lipid species such as cholesterol esters, phosphatidylethanolamines, ceramides, and sphingomyelins. To determine the broader relevance of these findings, we applied a reverse metabolomics approach and examined N-acyl lipid profiles in human small intestine and fecal samples from public data sets. Our results validate the colon simulator as a dynamic model for studying microbially transformed metabolites and suggest its potential utility as a platform for discovering novel microbial metabolites with relevance to human and animal health.