Vol 54, No 5 (2018)

The influence of the conditions of the formation of genipin cross-linked chitosan cryostructurates on the porous morphology and physicochemical properties of these cryostructurates and on the possibility of their use as biopolymer 3D scaffolds for tissue engineering was studied. The chitosan cryostructurates were obtained by freeze-drying a chitosan acetate solution, treating the resulting sponge with an alcohol solution of ammonia to transform the polyaminosaccharide from a salt into a chitosan-base, and then cross-linking the polymer with genipin (the molar ratios of genipin to the number of chitosan amino groups were 0.05, 0.033, and 0.02, respectively). The pore sizes, water-holding capacity, and in vitro biodegradation rate of the cryostructurates were shown to depend on the aforementioned ratio. The properties of the prepared chitosan cryostructurates, the hydrogels formed by chitosan cross-linking with genipin at positive temperatures, and the films cast from genipin-containing chitosan solutions after solvent evaporation were studied and compared. The biocompatibility of the obtained macroporous sponge materials was demonstrated using L929 mouse fibroblasts. Confocal laser microscopy showed that the cells in all of the 3D scaffolds obtained were evenly distributed; they grew and proliferated when cultured in vitro for seven days.

The conditions to obtain materials with elastic-viscous properties based on chitosan succinamide have been studied. A decreased polymer content and a transition from visco-elastic liquids to elastic-viscous systems were shown upon the addition of glycerol to an aqueous solution of chitosan succinamide. The systemic response, biological compatibility, and dynamics of bioresorbability of the obtained materials were studied during implantation in laboratory animals.

Conjugates of chitosan (molecular weight 28 and 830 kDa) with gallic acid were synthesized by a reaction initiated by a free radical. The conjugates contained 106 and 119 mg of polyphenol per g of polymer. Thin films were obtained from solutions of synthesized chitosan derivatives, and their physico-chemical characteristics (thickness of 0.058–0.076 mm and moisture content of 7.92–9.44%), as well as the antioxidant (inhibiting activity with respect to 2,2-diphenyl-1-picrylhydrazyl) and antimicrobial properties, were studied in relation to Staphylococcus epidermidis and Escherichia coli.

Silver nanoparticles have been obtained in a matrix of 6-O-carboxymethyl chitin in the presence of D-glucose as a reducing agent by microwave hydrothermal synthesis. The TEM results show that the silver nanoparticles have a spherical shape; the particle size range is 3–20 nm. The resulting colloidal solution of silver nanoparticles had a strong bacterial effect on gram-positive bacteria Staphylococcus aureus ATCC 21027 (=209 P), Bacillus subtilis ATCC 6633, and, to a lesser extent, on gram-negative bacteria Escherichia coli АТСС 25922. The synthesized silver nanoparticles showed pronounced fungistatic activity against A. niger INA 00760.

The paper reports on the isolation of an extracellular chitinase produced by the alkaliphilic Bacillus mannanilyticus IB-OR17 B1 strain grown in media containing crab shell and bee chitin at a pH of 8–11. The enzyme was 860-fold purified by ultrafiltration and chitin sorption. The molecular weight of the purified chitinase was shown by denaturing electrophoresis to be 56 kDa. The enzyme showed maximum activity at a pH of 7.5–8.0 and 65°C and was stable within a pH range of 3.5–10.5 and temperature range of 75–85°C. With colloidal chitin as substrate, the kinetic characteristics of the chitinase were determined as follows: K_M ~ 1.32 mg/mL and V_max ~ 5.05 μM min^–1. N-acetyl-D-glucosamine and its dimer were the main products of enzymatic chitin cleavage, while the trisaccharide was detected just in minor quantities. The chitinase actively hydrolyzed p-nitrophenyl-GlcNAc₂ according to the exo-mechanism of substrate hydrolysis characteristic of chitobiosidases.

Probability-based surveys of the application of chitosan succinate as an adjuvant added to the vaccine against animal necrobacteriosis have been performed. The addition of 3.7% chitosan succinate (MM 330 kDa) to the standard-dose vaccine formulation could contribute to improvement of the vaccine immunogenicity, as measured by the hemagglutination test (HA): 1 : 512 as compared to 1 : 128 in the control range. The use of 0.5-% chitosan succinate solution as a protective medium for drying in the production of a dried inactivated vaccine against infectious bovine rhinotracheitis could ensure the preservation of the biopreparation’s high immunogenic activity. Chitosan succinate was shown to have quite good solubility in water, allowing the possible use of saline solution to reconstitute the vaccine into the suspension for injection and to abandon expensive solvents.

The effect of chitooligosaccharides (ChOSs) with an acetylation degree (AD) of 65% and 30% on the formation of defense reactions of potato plants (Solanum tuberosum L.) against Phytophthora infestans (Mont.) De Bary is studied. The content of hydrogen peroxide, the activities of peroxidases, allene oxide synthase, and hydrolase inhibitors, and the transcriptional rate of pathogenesis-related (PR) proteins are studied. ChOSs with an AD of 65% reduced the degree of late blight symptoms on the leaves of potato plants via the stimulation of H₂O₂ accumulation and the transcriptional activity of genes encoding peroxidase (M21334), chitinase (U49969.1), and inhibitors of amylase (XM006351484) and proteinase (JX683427). It is found that ChOSs with an AD of 30% induced the activity of anionic isoperoxidases with a pI of ~3.5–4.5 in infected plants. The mechanisms of potato resistance to Phytophthora infestans formed under the influence of different ChOSs are discussed.

The treatment of wheat with polyfunctional biopreparations based on the Bacillus subtilis strains, chitosans, and their complexes, was shown to be highly efficient. These preparations activate the adaptive potential, productive capacity, and resistance of wheat to the causative agents of root rot, powdery mildew, and leaf rust. Plants treated with a Vitaplan complex preparation containing the culture liquid and chitosan II had the best adaptive potential to agroecological environmental factors. In this case, the growth rate and the weight of the vegetative part and ear had the highest values. At the same time, the development of root rot and the intensity of damage to the flag and preflag leaf with wheat leaf blotch were minimal in comparison with the control. The stress index, as determined according to the results of spectrometric analysis, can be an integral estimation of the effect of polyfunctional biopreparations.