Process for producing 4-vinylguaiacol by biodecaroxylation of ferulic acid

Abstract

4-vinylguaiacol is produced using recombinant E. coli containing a decarboxylase gene from Bacillus pumilis in an aqueous fermentation broth and in an immobilized whole cell system. The 4-vinylguaiacol is extracted and recovered from an organic hydrocarbon solvent, preferably n-octane, whereby the product can readily be separated.

Description

BRIEF DESCRIPTION OF DRAWINGS

The process of the invention is described in greater detail with reference to the following examples, and the accompanying drawings, wherein:

FIG. 1 is a graph of VG production in various media;

FIG. 2 is a graph of VG production at various temperatures in an aqueous/organic system;

FIG. 3 is a graph of cell growth rate for B. pumilus;

FIG. 4 is a graph of VG production using FA induced B. pumilus;

FIG. 5 is a graph of recombinant E. coli cell growth rate;

FIG. 6 is a bar graph of FA biotransformations involving the multi-utilization of immobilized recombinant E. coli; and

FIGS. 7 and 8 are graphs illustrating the specific activity of E. coli cells in alginate beads.

Claims

1. A process for producing 4-vinylguaiacol comprising the steps of cultivating recombinant E. coli containing a decarboxylase gene from Bacillus pumilus in an aqueous fermentation broth; adding an organic solvent and a ferulic acid substrate to the fermentation broth whereby 4-vinylguaiacol is formed and accumulates in the organic solvent; and separating the 4-vinylguaiacol from the organic solvent.

2. The process of claim 1, wherein the recombinant E. coli is added to the aqueous fermentation broth immobilized in aliginate.

3. The process of claim 1, wherein the E. coli is E. coli JM 109.

4. The process of claim 3, wherein the B. pumilus is strain AM670.

5. The process of claim 1, including the steps of preparing a recombinant E. coli cell paste; and combining the paste with alginate to produce beads of immobilized E. coli biocatalyst for use in the cultivating step of the process.

6. The process of claim 1, wherein the organic solvent is selected from the group consisting of cyclohexane, n-hexane, n-octane, n-dodecane and n-hexadecane.

7. The process of claim 1, wherein the organic solvent is n-octane.

8. The process of claim 1, wherein the recombinant E. coli is cultivated for 4 to 48 hours, the organic solvent is added to the fermentation broth at a ratio to the broth of 1:1 to 1:20, and the ferulic acid substrate is added to the broth in an amount of 5 to 25 g/L of broth.

9. The process of claim 8, wherein the fermentation is performed at a pH of 5 to 7 and a temperature of 7 to 37° C.

10. The process of claim 9, wherein the fermentation is performed at a temperature of 30° C.

11. The process of claim 5, wherein recombinant E. coli cells are grown in a fermentation broth, the broth is centrifuged to yield the cell paste, the cell paste is suspended in an alginate solution, and the mixture thus produced is added dropwise to calcium chloride solution to produce the beads.