Radek Stloukal

Butanol production by immobilised Clostridium acetobutylicum in repeated batch, fed-batch, and continuous modes of fermentation

Clostridium acetobutylicum immobilised in polyvinylalcohol, lens-shaped hydrogel capsules (LentiKats(®)) was studied for production of butanol and other products of acetone-butanol-ethanol fermentation. After optimising the immobilisation protocol for anaerobic bacteria, continuous, repeated batch, and fed-batch fermentations in repeated batch mode were performed. Using glucose as a substrate, butanol productivity of 0.41 g/L/h and solvent productivity of 0.63 g/L/h were observed at a dilution rate of 0.05 h(-1) during continuous fermentation with a concentrated substrate (60 g/L). Through the process of repeated batch fermentation, the duration of fermentation was reduced from 27.8h (free-cell fermentation) to 3.3h (immobilised cells) with a solvent productivity of 0.77 g/L/h (butanol 0.57 g/L/h). The highest butanol and solvent productivities of 1.21 and 1.91 g/L/h were observed during fed-batch fermentation operated in repeated batch mode with yields of butanol (0.15 g/g) and solvents (0.24 g/g), respectively, produced per gram of glucose.

Immobilization of cells and enzymes to LentiKats

Biocatalyst immobilization is one of the techniques, which can improve whole cells or enzyme applications. This method, based on the fixation of the biocatalyst into or onto various materials, may increase robustness of the biocatalyst, allows its reuse, or improves the product yield. In recent decades, a number of immobilization techniques have been developed. They can be divided according to the used natural or synthetic material and principle of biocatalyst fixation in the particle. One option, based on the entrapment of cells or enzymes into a synthetic polyvinyl alcohol lens with original shape, is LentiKats® immobilization. This review describes the preparation principle of these particles and summarizes existing successful LentiKats® immobilizations. In addition, examples are compared with other immobilization techniques or free biocatalysts, pointing to the advantages and disadvantages of LentiKats®.

Recombinant α-L-rhamnosidase of Aspergillus terreus immobilization in polyvinylalcohol hydrogel and its application in rutin derhamnosylation

Abstract Recombinant α-L-rhamnosidase from Aspergillus terreus expressed in Pichia pastoris was immobilized in LentiKats® lens-shaped polyvinylalcohol (PVA) capsules with an activity of 7 U g− 1, which was 21% of its original activity. Immobilization did not significantly affect the pH and temperature profile of α-L-rhamnosidase, KM increased by a factor of 3.4 whereas Vmax decreased more than 10-fold. No decrease in activity was observed after 27 repeated batch runs of rutin derhamnosylation. The enzyme proved to have an excellent storage stability (136 days) in 60 g L− 1 ethanol with no change in its activity.

Determination of low concentration of Paracoccus denitrificans encapsulated in polyvinyl alcohol LentiKat’s pellets

The aim of this work was to compare three methods to determinate low concentrations of Paracoccus denitrificans encapsulated in polyvinyl alcohol pellets, which is important for evaluation and optimization of pellet production as well as for monitoring of biomass growth. Pellets with different and well-defined biomass concentrations were used for experiments. The following fast and simple methods were tested: (1) dissolution of polyvinyl alcohol in hot water followed by dry weight estimation, (2) dissolution of polyvinyl alcohol in hot water followed by optical density measurement, (3) and extraction and quantification of proteins. Dry weight estimation proved to be problematic as it was difficult to separate biomass from polymeric carrier. Optical density measurement showed good linearity of dependence of optical density on biomass content, but determined limits of detection and limits of quantification were not within the range necessary for intended application. The only tested method meeting the requirements for sensitivity was determination of protein concentration after protein extraction.