High-Yield Production of Polyhydroxybutyrate and Poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) from Crude Glycerol by a Newly Isolated Burkholderia Species Oh_219

Abstract

Crude glycerol (CG), a major biodiesel production by-product, is the focus of ongoing research to convert it into polyhydroxyalkanoate (PHA). However, few bacterial strains are capable of efficiently achieving this conversion. Here, 10 PHA-producing strains were isolated from various media. Among them, Burkholderia sp. Oh_219 exhibited the highest polyhydroxybutyrate (PHB) production from glycerol and was therefore characterized further. Burkholderia sp. Oh_219 demonstrated significant tolerance to major growth inhibitors in CG and metabolized the fatty acids present as impurities in CG. Furthermore, the Oh_219 strain was genetically engineered using phaCBP-M-CPF4 and phaJPa to enable the fatty acid-based production of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx), a component of CG. The resulting strain produced PHBHHx containing 1.0–1.3 mol% of 3HHx from CG. Further supplementation with capric and lauric acids increased the 3HHx molar fraction to 9.7% and 18%, respectively. In a 5 L fermenter, the Oh_219 strain produced 15.3 g/L PHB from 29.6 g/L biomass using a two-stage fermentation system. This is the highest yield reported for PHA production from glycerol by Burkholderia spp. Additionally, PHB produced from CG had a lower melting point than that from pure glycerol and fructose. Taken together, Burkholderia sp. Oh_219 is a promising new candidate strain for producing PHA from CG.