Screening of Fungi Isolates for C-4 Hydroxylation of R-2-Phenoxypropionic Acid Based on a Novel 96-Well Microplate Assay Method

R -2-(4-Hydroxyphenoxy)propionic acid ( R -HPPA) is a pivotal intermediate for the synthesis of aryloxyphenoxypropionate (APP) herbicide. To rapidly screen microbial isolates with the capacity of hydroxylating R -2-phenoxypropionic acid to R -HPPA from various environmental samples, a convenient and...

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Bibliographic Details
Published in:Applied biochemistry and biotechnology 2020-09, Vol.192 (1), p.42-56
Main Authors: Zhou, Hai-Yan, Jiang, Rui, Li, Yi-Zuo, Xu, Wei, Wang, Yuan-Shan, Xue, Ya-Ping, Zheng, Yu-Guo
Format: Article
Language:English
Subjects:
Acids
Agar - chemistry
Animals
Aspergillus - genetics
Aspergillus - metabolism
Assaying
Biochemistry
Bioconversion
Biomass
Biotechnology
Bombyx - microbiology
Catalysis
Chemistry
Chemistry and Materials Science
China
Color
Cultivation
DNA, Ribosomal - genetics
Fermentation
Fungi
Herbicides
Herbicides - chemistry
Hydrogen-Ion Concentration
Hydroxylation
Methods
Microorganisms
Penicillium - genetics
Penicillium - metabolism
Phenyl Ethers - chemistry
Phylogeny
Potassium
Propionates - chemistry
Propionic acid
Reaction time
Reagents
Screening
Sewage - microbiology
Sodium nitrite
Sodium Nitrite - chemistry
Soil Microbiology
Strains (organisms)
Temperature
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Summary:R -2-(4-Hydroxyphenoxy)propionic acid ( R -HPPA) is a pivotal intermediate for the synthesis of aryloxyphenoxypropionate (APP) herbicide. To rapidly screen microbial isolates with the capacity of hydroxylating R -2-phenoxypropionic acid to R -HPPA from various environmental samples, a convenient and safe 96-well microplate assay method with sodium nitrite (NaNO 2 ) as chromogenic reagent was proposed and optimized. The optimum assay conditions were as follows: the detection wavelength was 420 nm, the concentration of NaNO 2 solution was 6.0 g/L, color reaction temperature was 60 °C, the pH of the NaNO 2 solution was 2.4, and the reaction time was 40 min. With the aid of this method, screening for microorganisms with C-4-specific hydroxylation activity of R -PPA was conducted. As a result, 23 strains among 3744 single colonies isolated from various samples exhibited the hydroxylation activity. Among these strains, the highest bioconversion rate was achieved by Penicillium oxalicum A5 and Aspergillus versicolor A12, respectively. After 72-h cultivation in shake flask, their conversion rates of R -HPPA from 10 g/L R -PPA reached 21.18% and 40.24%, respectively. The established method was effective in rapid screening of microbes capable of biosynthesizing R -HPPA through hydroxylation of R -PPA, and the obtained two fungi species could be potentially used for R -HPPA production.
ISSN:0273-2289
1559-0291
DOI:10.1007/s12010-020-03303-z