A high-throughput screening method for improved R-2-(4-hydroxyphenoxy)propionic acid biosynthesis
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R-2-(4-hydroxyphenoxy)propionic acid (R-HPPA) is a key intermediate of the enantiomerically pure phenoxypropionic acid herbicides. R-HPPA could be biosynthesized through selective introduction of a hydroxyl group (–OH) into the substrate R-2-phenoxypropionic acid (R-PPA) at C-4 position, facilitated by microorganisms with hydroxylases. In this study, an efficient high-throughput screening method for improved R-HPPA biosynthesis through microbial hydroxylation was developed. As a hydroxylated aromatic product, R-HPPA could be oxidized by oxidant potassium dichromate to form brown-colored quinone-type compound. The concentration of R-HPPA can be quantified according to the absorbance of the colored compound at a suitable wavelength of 570 nm; and the R-HPPA biosynthetic capability of microorganism strains could also be rapidly evaluated. After optimization of the assay conditions, the high-throughput screening method was successfully used in identification of Beauveria bassiana mutants with enhanced R-HPPA biosynthesis capacity. A positive mutant C-7 with high tolerance to 20 g/L R-PPA was rapidly selected from 1920 mutants. The biomass and R-HPPA titer were 12.5- and 38.19-fold higher compared with the original strain at 20 g/L R-PPA. This high-throughput screening method developed in this work could also be a potential tool for screening strains producing other important phenolic compounds.
KeywordsHigh-throughput screening R-2-(4-hydroxyphenoxy)propionic acid R-2-phenoxypropionic acid Hydroxylation Potassium dichromate
The authors gratefully acknowledge the financial support of the National Natural Science Foundation of China (No. 31500031) and the Postgraduate Teaching Reform Project of Zhejiang University of Technology (No. 2018114).
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Conflict of interest
The authors declare that they have no conflict of interest.
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