Automatable downstream purification of the benzohydroxamic acid D-DIBOA from a biocatalytic synthesis

ElsevierVolume 72, 25 December 2022, Pages 48-57New BiotechnologyHighlights•

Design of a 3-steps D-DIBOA purification method without organic solvent wastes.

Novel affinity chromatography for the purification of benzohydroxamic acids.

Prototype for the automation of D-DIBOA purification from whole-cell biocatalysis.

Both biological and synthetic D-DIBOA have the same herbicide activity.

Abstract

Herbicides play a vital role in agriculture, contributing to increased crop productivity by minimizing weed growth, but their low degradability presents a threat to the environment and human health. Allelochemicals, such as DIBOA (2,4-dihydroxy-(2H)-1,4-benzoxazin-3(4 H)-one), are secondary metabolites released by certain plants that affect the survival or growth of other organisms. Although these metabolites have an attractive potential for use as herbicides, their low natural production is a critical hurdle. Previously, the synthesis of the biologically active analog D-DIBOA (4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one) was achieved, using an engineered E. coli strain as a whole-cell biocatalyst, capable of transforming a precursor compound into D‐DIBOA and exporting it into the culture medium, although it cannot be directly applied to crops. Here a chromatographic method to purify D-DIBOA from this cell culture medium without producing organic solvent wastes is described. The purification of D-DIBOA from a filtered culture medium to the pure compound could also be automated. Biological tests with the purified compound on weed models showed that it has virtually the same activity than the chemically synthesized D-DIBOA.

AbbreviationsDIBOA

2,4-dihydroxy-(2H)-1,4-benzoxazin-3-(4H)one

D-DIBOA

4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one, or 2-deoxy-DIBOA

wcb-D-DIBOA

D-DIBOA obtained by whole-cell biocatalysis

cs-D-DIBOA

D-DIBOA obtained by chemical synthesis

PC

ethyl-2-(2´-nitrophenoxy)acetate (D-DIBOA precursor)

IMAC

immobilized-metal affinity chromatography

IEX

ion-exchange chromatography

Mono Q

anion exchange column

Mono S

cation exchange column

RPLC

Reversed Phase Liquid Chromatography

Keywords

D-DIBOA

Green chemistry

Whole-cell biocatalyst

Downstream processes

Chromatography

Ecological herbicides

© 2022 The Authors. Published by Elsevier B.V.

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