Introduction and Objectives The integrity of the gut microbiota is directly related to host health, and one way to maintain the diversity of the gut microbiota is to promote the exchange of compounds between donor and recipient bacteria, a phenomenon called cross-feeding. In this way, the discovery of new compounds that are exchanged could help keep us healthy. Quinones are key molecules in the energy-producing bacterial respiratory chains and have been proposed to be cross-fed. The aim of this study was to test whether quinone cross-feeding actually exists and to assess its prevalence in the human gut microbiota. Material and Methods Menaquinone (MK) is the predominant quinone in bacteria, and we performed a bioinformatic annotation with HMM profiles of its biosynthetic pathway in more than 1500 bacterial species from the Unified Human Gastrointestinal Genome (UHGG) datase. Phylogenetic trees with maximum likelihood were obtained to predict transmissions of MK pathways previously detected. Biochemical analysis of MK precursors and potential MK donor species revealed quinone synthesis in recipient bacteria. Heterologous complementation experiments confirmed the functionality of partial MK pathways found in recipient species. Finally, we developed a novel method based on high performance liquid chromatography-mass spectrometry (HPLC-MS) to quantify the diversity of quinones present in gut microbiota samples. Results, Discussion and Conclusion Bioinformatic annotations revealed partial MK pathways in bacterial species belonging to all major phyla present in the gut microbiota. The functionality of the genes constituting the partial MK pathways in recipient bacteria was validated experimentally, and cross-feeding of quinone precursors was confirmed. Our phylogenetic analysis revealed that the first part of the MK biosynthesis pathway was repeatedly lost in several bacterial phyla. Overall, our data show that the biosynthesis of quinones in several species depends on cross-feeding from other bacteria, or also possibly from the host. This suggests that this phenomenon is likely to contribute to the equilibrium of the gut microbiota, considering the physiological importance of quinones in bacterial bioenergetics. Our quinone quantification method will now allow us to confirm quinone cross-feeding in vivo, and to evaluate the quinone content in dysbiotic gut microbiota.