The products of chitosan hydrolysis are chitooligosaccharides and are used mainly for medical applications due to their specific biological activities. The objective of this study was to detect and identify the products of enzymatic hydrolysis of chitosan (dimers to hexamers) using a crude extract of chitosanolytic enzymes produced by the fungus Metarhizium anisopliae. These fungus was able to produce, during 48 h cultivation in a medium containing chitosan, chitooligosaccharides ranging from dimers, trimers, tetramers and pentamers at concentrations 0.2, 0.19, 0.06, 0.04 mg/mL, respectively, and the enzymatic activity was 2.5 U/L. Using the crude enzyme extract for chitosan hydrolysis, we detected the presence of dimers to hexamers at hydrolysis times of 10, 20, 30, 40, 50 and 60 min of enzymatic reaction, but the yields were higher at 10 min (54%). The hexamers was obtained only with 30 min of reaction with concentration of 0.004 mg/mL.

In this study, the kinetic behavior of Sf9 and Sf21 cells used in the production of a baculovirus biopesticide to control the pest of corn Spodoptera frugiperda was analyzed. Kinetic variables such as maximum specific growth rate, cell productivity, mean rate of infection, as well as the mean rate of occlusion body production were determined during the infection of these cell-lines with the extracellular virus of the S. frugiperda nucleopolyhedrovirus (SfMNPV). The Sf9 cell-line resulted in better viral production results (5.0 x 108 OB/mL) than the Sf21 cell-line (2.5 x 10 8 OB/mL).

Internalization of hydrophilic molecules into yeast cytosol is required for different applications such as cell transformation or preservation of water soluble components by bioencapsulation. However, these molecules are not able to cross the plasma membrane and strategies have to be developed. Recent works revealed that osmotic perturbations could induce non-lethal transient permeabilization of the plasma membrane. In this work, we endeavored to clarify the phenomenon of permeabilization during rehydration after a mild hyperosmotic perturbation in order to evaluate the possibility of hydrophilic molecule internalization in yeast by this treatment. Rehydration step is particularly interesting because the large entry of water into the cells could help the internalization of molecules. The internalization of a fluorescent molecule [fluorescein isothiocyanate Dextran (FITC-Dextran), 20 kDa], added during the rehydration after a sublethal hyperosmotic treatment, was studied in Saccharomyces cerevisiae yeast cells. The internalization kinetic and the localization of the fluorescent molecules were studied by flow cytometry and fluorescence confocal microscopy. Our results show that the rehydration leads to the rapid internalization of FITC-Dextran due to a transient plasma membrane permeabilization. Thus, osmoporation, i.e. plasma membrane poration by modifications of osmotic pressure of the extracellular medium, could be a new and simple way to deliver molecules of particular interest into yeasts.