Process optimisation for polyhydroxyalkanoates production with tailored hydroxyvalerate content from agri-food residues using mixed microbial cultures

BACKGROUND: Polyhydroxyalkanoates (PHAs) are promising bio-based and biodegradable polymers that can contribute to the transition towards a circular bioeconomy, particularly when produced from renewable feedstocks. Nevertheless, the use of mixed microbial cultures (MMC) and real agri-food residues poses challenges related to substrate variability, process stability and control of polymer composition. In this study, a three-stage process was investigated to produce poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) from agri-food residues. The process consisted of: (i) acidogenic fermentation of cattle slurry and corn residues to generate a carboxylic acid (CA)-rich stream; (ii) microbial selection under feast-famine conditions to enrich PHA-accumulating microorganisms; and (iii) PHA accumulation in a fed-batch reactor. Two operational setups, differing in the pretreatment type, were compared to evaluate possible effects on PHA production: setup 1 used centrifuged fermentate, whereas setup 2 included an additional 0.22-µm filtration step. RESULTS: , mean PHA titre of 1.70 ± 0.44 g/L and intracellular PHA values up to 57.0% w/w. The hydroxyvalerate (HV) monomer content showed temporal variability, ranging from 14.0-32.0% w/w in setup 1 and from 13.0-35.0% w/w in setup 2, with values generally within or close to the target range required to improve polymer flexibility and processability. Microbial community analysis revealed a marked reduction in genus richness following feedstock filtration and the progressive dominance of specialised PHA-storing genera, indicating effective selection pressure under feast-famine operation and supporting the observed improvements in polymer accumulation. CONCLUSIONS: Overall, the results indicate that MMC-based PHBV production from agri-food residues can achieve polymer contents comparable to those reported for similar systems using complex substrates, while highlighting the importance of feedstock pretreatment and process control. The study provides insights into the integration of acidogenic fermentation with MMC-based PHA production, and identifies operational aspects that require further optimisation to enhance process robustness and product consistency.