Production of P(3HB-co-3HHx) with Controlled Compositions by Recombinant Cupriavidus necator Re2058/pCB113 from Renewable Resources

Abstract

Mixtures of crude palm kernel oil (CPKO) and oil palm tree trunk sap (OPTS) as carbon sources for the biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (P(3HB-co-3HHx)) by the recombinant Cupriavidus necator strain Re2058/pCB113 were investigated. Shake flask cultures using CPKO as the sole carbon source produced 5.82 g/L of cell dry weight (CDW) with 73 wt% of PHA content and 44 mol% of 3-hydroxyhexanoate (3HHx). When the total CPKO concentration in the cultures was increased from 2.5 to 7.5 g/L, a general pattern of decreasing 3HHx molar fraction (from 42 to 27 mol%) was observed in the resulting copolymer. In order to produce biopolymer with properties that resemble commodity plastics, the 3HHx molar fraction should be lowered to an optimal level of ∼12–20 mol%. The lowest 3HHx molar fraction (14 mol%) was obtained when a mixture of 4% (w/v) OPTS sugars and 1.5 g/L CPKO was used as the carbon source. The highest molar fraction of 3HHx produced (42 mol%) was observed when the cells were cultivated in 5 g/L of CPKO. 1H-NMR and 13C-NMR analyses confirmed the structure and monomer fraction of the different P(3HB-co-3HHx) polymers. This study has established a method to control the monomer molar fractions in P(3HB-co-3HHx) produced using a mixture of CPKO and sugars from OPTS.