Production of cis-1,2-dihydrocatechols of high synthetic value by whole-cell fermentation using Escherichia coli JM109 (pDTG601): A detailed study

• Growing conditions for high cell density cultures of E. coli JM109 (pDTG601).
• Biotransformation kinetics and effect of bacterial concentration on transformation productivities.
• Biotransformation using bi-phasic procedures to decrease the toxic impact of the substrate.
• Storage of active biocatalyst as a chemical reagent for dihydroxylation of organic substrates.

Chiral cis-1,2-dihydrocatechols have been extensively used as starting materials for the synthesis of complex organic molecules. The preparation of these high added value compounds is carried out mainly by enzymatic dihydroxylation of arenes since no other efficient chemical method is known. In this paper we describe a detailed study of the biotechnological production at a preparative scale of cis-3-bromo-1,2-dihydrocatechol (BDC) from bromobenzene using Escherichia coli JM109 (pDTG601), a recombinant strain that harbors the toluene dioxygenase genes from Pseudomonas putida F1. High cell-density cultures of the microorganism (65 g/L cdw) were achieved in a 5 L bioreactor using fed-batch cultures in aerobic conditions. The influence of the biomass concentration in the volumetric and specific productivity of the system, as well as the kinetics of the biotransformation, was thoroughly studied. The use of liquid paraffin as a second phase to relieve bromobenzene's toxicity resulted in an increased specific productivity for the bi-phasic protocol, which resulted in higher final BDC concentrations. An improvement of 3.2-fold was obtained for BDC production compared to previous reports using this organism. Furthermore, storage strategies of the biocatalytic system as a reagent for organic synthesis are presented.

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