Impact of pretreatment severity and nutrient parameters on microbial PHB production from shrub willow hydrolysate using recombinant Escherichia coli

Developing biodegradable bioplastics from renewable feedstocks is critical to reduce dependence on fossil-based plastics. This study assessed the feasibility of producing polyhydroxybutyrate (PHB) from a fast-growing woody crop, Shrub willow (Salix spp.), and examined the effects of pretreatment severity and nutrient parameters on PHB production. Importantly, the study systematically evaluated different nitrogen sources and the carbon-to-nitrogen (C/N) ratio, key factors influencing PHB accumulation. Biomass was pretreated using hot water at 160–220 °C, followed by disk milling. The highest glucose yield during subsequent hydrolysis was observed at 200 °C; however, the hydrolysate obtained from 180 °C supported the highest PHB titer of 1.63 g/L. Among the various organic and inorganic nitrogen sources investigated, yeast extract improved the cell growth by nearly twofold (6.33 g/L) at a C/N ratio of 20 compared to the control. Investigation of a broader range of C/N ratios identified 10 as the most favorable, producing 9.1 g/L cell dry weight, 44.7% PHB inclusion, and a PHB titer of 4.06 g/L. These findings display the feasibility of producing PHB from shrub willow and identify key parameters influencing PHB yields.