Exploring media optimization and extraction methods to enhance poly-β-hydroxybutyrate (PHB) yield in Synechocystis pevaleikii: Scale-up from shake flask to 10 L photobioreactor.

The surge in demand for sustainable alternatives to petroleum-based plastics has redirected attention towards microbial biopolymers, such as Poly-β-hydroxybutyrate (PHB). This study identified the cyanobacterium Synechocystis pevaleikii as an unexplored potential organism for PHB production. Cultivation was performed in BG-11 media with varying exogenous acetate concentrations (0.08-2.46 g/L). The highest chlorophyll-a content (9.12 ± 0.44 μg/mL), biomass yield (600 ± 17 mg/L), and PHB accumulation (3.01 % w/w) were achieved with 1.64 g/L acetate. Scaling up to a Recirculating Media Photobioreactor of 10 L capacity using optimized media resulted in a chlorophyll-a content of 7.67 ± 0.10 μg/mL, while the biomass yield was enhanced to 950 ± 50 mg/L and PHB accumulation of 2.28 % (w/w). For biopolymer recovery, Soxhlet extraction was optimized across varying durations (1-10h). The 5 h Soxhlet extraction was identified as optimal, yielding polyhydroxyalkanoates (PHA) 13.4 ± 0.6 % and PHB of 2.8 ± 0.13 % with maximum medium chain length polymer (C16-C20). Polymer purification using methanol precipitation yielded lower PHA content (0.507 ± 0.02 % w/w), highlighting limited efficiency. FTIR analysis confirmed the presence of consistent functional groups of PHA across extraction durations. GC-MS revealed that the polymer mainly consisted of esters and hydrocarbons, especially alkanes and alkenes. DSC revealed that the PHA was semicrystalline with a crystallinity of 54.8 %, a glass transition temperature of 5 °C, and a melting temperature of 175 °C. These findings underscore the scalability of S. pevaleikii for biopolymer production using a low-cost nitrate phosphate carbonate (NPC) media supplemented with acetate. Furthermore, a 5 h Soxhlet extraction provides an effective method for biopolymer recovery in reactor-scale applications.