Production and Characterization of Poly-β-hydroxyalkanoate Copolymers from Burkholderia cepacia Utilizing Xylose and Levulinic Acid

Abstract

Poly(beta-hydroxybutyrate-co-beta-hydroxyvalerate) (P(3HB-co-3HV)) copolymers were prepared via shake-flask fermentations of Burkholderia cepacia (formerly Pseudomonas cepacia) containing 2.2% (w/v) xylose and concentrations of levulinic acid ranging from 0.07% to 0.67% (w/v). Periodic harvest of shake-flask cultures from 48 to 92 h post-inoculation yielded 4.4-5.3 g/L of dry cell biomass, containing 42-56% (w/w) P(3HB-co-3HV), with optimal product yield occurring between 66 and 74 h. Growth and PHA accumulation enhancement were observed with concentrations of levulinic acid from 0.07 to 0.52% (w/v), producing dry cell biomass and P(3HB-co-3HV) yields of 9.5 and 4.2 g/L, respectively, at the 0.52% (w/v) concentration of levulinic acid. Representative samples were subjected to compositional analysis by 300 MHz 1H and 150 MHz 13C NMR, indicating that these random copolymers contained between 0.8 and 61 mol % 3-hydroxyvalerate (3HV). Solvent-cast film samples were characterized by differential scanning calorimetry, which demonstrated melting temperatures (Tm) to decrease in a pseudoeutectic fashion from 174.3 degrees C (0.8 mol % 3HV) to a minimum of 154.2 degrees C (25 mol % 3HV) and the glass transition temperatures (Tg) to decrease linearly from 2.1 to -11.9 degrees C as a function of increasing mol % 3HV. Thermogravimetric analysis of the copolymer series showed the temperature for onset of thermal decomposition (T(decomp)) to vary as a function of mol % 3HV from 273.4 to 225.5 degrees C. Intrinsic viscosities (eta) varied from 3.2 to 5.4 dL/g, as determined by dilute solution viscometry. Viscosity average molecular weights (Mv) of the copolymers were determined to range from 469 to 919 kDa, indicating that these P(3HB-co-3HV) copolymers are of sufficient molecular mass for commercial application.