Real time in vivo methods are needed to better understand the interplay between diet and the gastrointestinal microbiota. Therefore, a rodent indirect calorimetry system was equipped with hydrogen (H2) and methane (CH4) sensors. H2 production was readily detected in C57BL/6J mice and followed a circadian rhythm. H2 production was increased within 12 hours after first exposure to a lowly-digestible starch diet (LDD) compared to a highly-digestible starch diet (HDD). Marked differences were observed in the faecal microbiota of animals fed the LDD and HDD diets. H2 was identified as a key variable explaining the variation in microbial communities, with specific taxa (including Bacteroides and Parasutterella) correlating with H2 production upon LDD-feeding. CH4 production was undetectable which was in line with absence of CH4 producers in the gut. We conclude that real-time in vivo monitoring of gases provides a non-invasive time-resolved system to explore the interplay between nutrition and gut microbes in a mouse model, and demonstrates potential for translation to other animal models and human studies.