Purple non-sulfur bacteria (PNSB) are well known for converting short-chain organic acids to H2, however, a decrease in pH caused by metabolic acids production limited H-2 production during the photo-fermentation from glucose. Here we address why volatile fatty acids (VFA) excreted as fermentation products cannot be further degraded by R. sphaeroides that readily use them. We found that the photo-fermentation with pH controlled at 6.9 +/- 0.1 resulted in a 90% increase of H-2 yield and a 107.6% increase in volume H-2 production relative to the pH-uncontrolled culture. Comparative fermentations on glucose at pH 5.8 and pH 7.1 using culture medium supplemented with 50% spent fermentation broth demonstrated that low pH alone is not the limiting factor and compounds present in the supernatants along with pH decrease were the most inhibitory to H-2 production. The impact of byproducts VFA on phototrophic H-2 production was dependent on both the pH and VFA concentrations; even 7 mM VFA addition totally inhibited H-2 production from glucose at pH 5.4. H-2 production with pH control for the Ahup strain was not discernibly different from the parent strain, which are all significantly higher than high-performance strains by metabolic engineering. These results demonstrate that pH dependent VFA inhibition can be turned into a driving force for enhanced H-2 production from glucose by pH regulation. (C) 2020 Hydrogen Energy Publications LLC. Published by Elsevier Ltd. All rights reserved.