Acidogenic fermentation of tequila vinasse: Impact of operational pH on metabolic profile and microbial communities

The objective of this study was to determine how operational pH influences metabolite production and bacterial community composition during the acidogenic fermentation of tequila vinasse (TV). Batch assays were conducted at pH 6.0-8.0, followed by a single sequencing batch reactor (SBR) operated sequentially at pH 6.5-4.5. In batch assays, pH 6.0 resulted in the highest apparent soluble COD conversion degree (53%), butyrate production (4.8 g/L; 75% selectivity), and hydrogen yield (60 mL H 2 /g COD added ). At pH 7.0 and 8.0, hydrogen was not detected, and the predominant net products shifted toward ethanol and acetate, respectively. During SBR operation, hydrogen production increased from 0.2 L H 2 /L TV at pH 6.5 to 1.86-2.00 L H 2 /L TV at pH 5.5-4.5. The highest apparent conversion degree occurred at pH 5.5 (58%), whereas hydrogen production was maximized at pH 5.0. Butyrate remained the main product, while acidification reached 71-76% under the most acidic conditions. Bacterial community composition shifted markedly with the operational conditions. Particularly, facultative genera predominated under neutral-to-alkaline batch conditions, whereas lactic acid bacteria became increasingly dominant across the sequential acidic SBR periods and Clostridium remained present throughout operation. These results show that operational pH can orient TV fermentation toward distinct hydrogen- and SCCA-rich profiles.