Permeabilization of Microbacterium oxylans shifts the conversion of puerarin from puerarin-7-O-glucoside to puerarin-7-O-fructoside

The main product of the conversion of puerarin by unpermeabilized cells of bacterium Microbacterium oxydans CGMCC 1788 was puerarin-7-O-glucoside (241 +/- 31.9 A mu M). Permeabilization with 40% ethanol could not increase conversion yield, whereas it resulted in change of main product; a previous trace product became a main product (213 A +/- 48.0 A mu M) which was identified as a novel puerarin-7-O-fructoside by electrospray ionization time-of-flight MS, C-13 NMR, H-1 NMR, and GC-MS analysis of sugar composition, and puerarin-7-O-glucoside became a trace product (14.8 A +/- 5.4 A mu M). However, the extract from cells of M. oxydans CGMCC 1788 permeabilized with ethanol converted puerarin to form 113.9 A +/- 27.7 A mu M puerarin-7-O-glucoside and 187.8 A +/- 29.5 A mu M puerarin-7-O-fructoside under the same conditions. When unpermeabilized intact cells were recovered and used repeatedly for the conversion of puerarin, with increase of reuse times, the yield of puerarin-7-O-glucoside gradually decreased, whereas the yield of puerarin-7-O-fructoside increased gradually in the conversion mixture. The main product of the conversion of puerarin by the tenth recycled unpremerbilized cells was puerarin-7-O-fructoside (288.4 A +/- 24.0 A mu M). Therefore, the change of permeability of cell membrane of bacterium M. oxydans CGMCC 1788 contributed to the change of conversion of the product's composition.