A model system for increasing the intensity of whole-cell biocatalysis: investigation of the rate of oxidation of D-sorbitol to L-sorbose by thin bi-layer latex coatings of non-growing Gluconobacter oxydans

We developed a novel <50-microm thick nano-porous bi-layer latex coating for preserving Gluconobacter oxydans, a strict aerobe, as a whole cell biocatalyst. G. oxydans was entrapped in an acrylate/vinyl acetate co-polymer matrix (T (g) approximately 10 degrees C) and cast into 12.7-mm diameter patch coatings (cellcoat) containing approximately 10(9) CFU covered by a nano-porous topcoat. The oxidation of D-sorbitol to L-sorbose was used to investigate the coating catalytic properties. Intrinsic kinetics was studied in microbioreactors using a pH 6.0 D-sorbitol, phosphate, pyruvate (SPP) non-growth medium at 30 degrees C, and the Michaelis-Menten constants determined. By using a diffusion cell, cellcoat and topcoat diffusivities, optimized by arresting polymer particle coalescence by glycerol and/or sucrose addition, were determined. Cryo-FESEM images revealed a two-layer structure with G. oxydans surrounded by <40-nm pores. Viable cell density, cell leakage, and oxidation kinetics in SPP medium for >150 h were investigated. Even though the coatings were optimized for permeability, approximately 50% of G. oxydans viability was lost during cellcoat drying and further reduction was observed as the topcoat was added. High reaction rates per unit volume of coating (80-100 g/L x h) were observed which agreed with predictions of a diffusion-reaction model using parameters estimated by independent experiments. Cellcoat effectiveness factors of 0.22-0.49 were observed which are 20-fold greater than any previously reported for this G. oxydans oxidation. These nano-structured coatings and the possibility of improving their ability to preserve G. oxydans viability may be useful for engineering highly reactive adhesive coatings for multi-phase micro-channel and membrane bioreactors to dramatically increase the intensity of whole-cell oxidations.