Fermentation Of Amino Acids By Anaerobic Microbiomes In The Absence Of Methanogenesis

Anaerobic digestion is commonly used to recover beneficial products from organic wastes. While combustible biogas is the most common product of anaerobic digestion, volatile fatty acids (VFAs) are an emerging alternative and can be used to produce chemicals and fuels that are otherwise derived from fossil fuels. This work investigates the fermentation of amino acids widely present in organic waste streams using anaerobic microbiomes. Individual and complex mixtures of amino acids were assessed, and the influence of operational parameters were evaluated. First, batch experiments were performed to assess the degradation of the 20 most common amino acids while methanogenesis is inhibited. Each amino acid was tested individually for degradability and 15 showed significant degradation (>50% consumed) after 14 days of incubation. Butyric acid was the most common VFA produced throughout the batch experiments. Following the batch experiments, fermentation of mixed amino acids was tested in a flow-through bioreactor. In the first 96-day phase, the pH of the bioreactors was not controlled, and it remained stable at near neutral pH. In a second 96-day phase, the pH was controlled at 5.5. VFA production was greater when the pH was controlled at acidic levels. Similar to batch experiments with single amino acids, butyric acid was the most abundant VFA produced in both phases, followed by isovaleric acid when the pH was controlled at 5.5. Analysis of the microbial community with 16S rRNA gene amplicon sequencing showed that bioreactors were dominated by species within the Arcobacter genus. While Caproiciproducens species were present and are known to produce hexanoic acid, no appreciable amounts of hexanoic acid were generated by the bioreactors. To further assess the microbial communities in the bioreactors degrading amino acids, metagenomics and metatranscriptomics were used to generate draft genomes of abundant organisms and to determine which genes were highly transcribed. These analyses revealed that the different pH phases varied at the species level but genomic potential between the microbiomes remained similar. An unclassified Comamonas species became enriched at pH 5.5 and its gene expression suggests it has a major role in fermenting amino acids directly to butyric acid via reverse b-oxidation. In summary, this research suggests that amino acids are a suitable substrate for butyric acid production, but more work is needed to increase production of other beneficial products from protein-rich waste streams.