Production of (R)-3-hydroxybutyric acid by Burkholderia cepacia from wood extract hydrolysates

Abstract

(R)-hydroxyalkanoic acids (R-HAs) are valuable building blocks for the synthesis of fine chemicals and biopolymers because of the chiral center and the two active functional groups. Hydroxyalkanoic acids fermentation can revolutionize the polyhydroxyalkanoic acids (PHA) production by increasing efficiency and enhancing product utility. Modifying the fermentation conditions that promotes the in vivo depolymerization and secretion to fermentation broth in wild type bacteria is a novel and promising approach to produce R-HAs. Wood extract hydrolysate (WEH) was found to be a suitable substrate for R-3-hydroxybutyric acid (R-3-HB) production by Burkholderia cepacia. Using Paulownia elongate WEH as a feedstock, the R-3-HB concentration in fermentation broth reached as high as 14.2 g/L after 3 days of batch fermentation and the highest concentration of 16.8 g/L was obtained at day 9. Further investigation indicated that the composition of culture medium contributed to the enhanced R-3-HB production.

Keywords: (R)-3-hydroxybutyric acid; Burkholderia cepacia; Wood extract hydrolysate.

Figure 1

2D HSQC spectrum for wood exact hydrolysate (WEH) (A: Paulownia elongate WEH before B.cepacia fermentation; B: Paulownia elongate WEH after 11 days B.cepacia fermentation; C: Acer saccharum WEH before B.cepacia fermentation; D: Acer saccharum WEH after 10 days B.cepacia fermentation).

Figure 2

Proton NMR spectrum of reagent 3-HB.

Figure 3

Proton spectrum of B. cepacia fermentation broth after 8 days of fermentation using Paulownia elongate WEH as substrate.

Figure 4

Standard curve of normalized peak area of 3-HB in proton spectrum corresponding to its concentration.

Figure 5

3-HB concentrations in the fermentation broth determined by NMR readings (■) and standard curve (blue circle symbol).

Figure 6

R-3-HB production during batch fermentation of B. cepacia using 50% Acer saccharum WEH or 50% Paulownia elongate WEH as substrate (R-3-HB in Acer saccharum WEH: •; R-3-HB in Paulownia elongate WEH: ■; Acetic acid in Acer saccharum WEH: blue up pointing triangle symbol; Acetic acid in Paulownia elongate WEH:blue down pointing triangle symbol).

Figure 7

R-3-HB production during batch fermentation using 50 g/L glucose or 50 g/L xylose as substrate (xylose: green diamond symbol; glucose: ■; xylose with extra nitrate and chloride ions red circle symbol; xylose with extra nitrate ion blue up pointing triangle symbol).

Figure 8

Biomass yield form batch fermentations.

Figure 9

PHA content in dried biomass from batch fermentations.

Figure 10

Metabolic pathways of the biosynthesis and intracellular degradation of PHA. ① PhaA, b-ketothiolase; ② PhaB, NADPH-dependent acetoacetyl-CoA reductase; ③ PhaC, PHA synthase; ④ PhaZ, PHA depolymerase; ⑤ (R)-3-Hdyroxybutyrate dehydrogenase; ⑥ Acetoacetyl-CoA synthetase.