Metabolic engineering of Ralstonia eutropha for enhanced poly(3-hydroxybutyrate) production from crude glycerol

A recombinant strain of Ralstonia eutropha B101 was developed with introduction of Vitreoscilla hemoglobin (VHb) gene into the wild-type R. eutropha H16 for cell growth and PHA production enhancement. The recombinant strain consumed approximately 33.35 g L^-1 of the glycerol consumed after 72 h of fermentation, significantly outperforming R. eutropha H16 (13.65 g L^-1). Higher cell dry weight (CDW) of 37.94 g L^-1, PHA content of 68.39 %, and a PHA concentration of 22.95 g L^-1 were obtained under optimal condition. The synthesized PHA was identified as poly(3-hydroxybutyrate) (P3HB), with a molecular weight (M_W) of 223 kDa, a polydispersity index (PDI) of 1.43, thermal degradation onset at 240 °C, and a crystallinity degree of 49 %. These findings demonstrate the feasibility of crude glycerol originated from industrial waste as a sustainable and cost-effective carbon source for recombinant R. eutropha B101 to produce PHA, thereby providing an eco-friendly approach in industrial-scale production of the biodegradable plastics.