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Dextransucrase (DS) is a glucosyltransferase (E. C. 2.4.1.5) that catalyzes the transfer of glucosyl residues from sucrose to dextran polymer and liberates fructose. This enzyme isassociated with a wide application range of dextran and oligosaccharides. DS production by Leuconostoc mesenteroidesT3 was optimized using a Central Composite Design under the Response Surface Methodology. Three variables were chosen for optimization: distillery stillage, sucrose and manganese concentration. The results showed that sucrose and manganese concentrations had a positive linear effect on DS production while all variable interactions (stillage-manganese, stillage-sucrose, and sucrose-manganese) had significant influences on the DS production. The maximal DS yield of 3.391±0.131 U cm-3, was obtained in the medium with 64.33 % distillery stillage concentration, 5.30% sucrose concentration and 0.022 % manganese concentration. Our study revealed the potential of distillery stillage combined with sugar beet molasses, supplemented with sucrose and manganese to be employed as a valuable medium growth for lactic acid bacteria and production of DS. Also, taking into consideration the origin of the substrates, utilization of industrial by-products in this way has a great environmental relevance and is in accordance with circular economy., Dekstransaharaza (DS) je glukoziltransferaza (E. C. 2.4.1.5.) koja katalizuje prenos ostataka glukoze iz saharoze u polimer dekstrana, pri čemu se oslobađa fruktoza. Ovaj enzim je povezan sa širokim spektrom primene dekstrana i oligosaharida. Proizvodnja dekstransaharaze pomoću bakterije Leuconostoc mesenteroides T3 optimizovana je metodom odzivnih površina korišćenjem centralnog kompozitnog dizajna. Za optimizaciju su izabrane tri promenljive: koncentracija džibre, koncentracija saharoze i koncentracija jona mangana. Rezultati su pokazali da koncentracije saharoze i jona mangana imaju pozitivan linearni efekat na proizvodnju DS dok su sve interakcije (džibra-Mn2+, džibra-saharoza i saharoza-Mn2+) imale značajan uticaj na proizvodnju DS. Na osnovu eksperimentalnih podataka i numeričke optimizacije, dobijen je maksimalni prinos DS od 3.391 ± 0.131 U cm-3 u podlozi sa 64.33 % džibre, 5.30 % saharoze i 0.022 % jona mangana. Naše istraživanje otkrilo je da se džibra u kombinaciji sa melasom šećerne repe kao i saharozom i dodatkom jona mangana može koristiti kao dragocena hranjiva komponenta za rast bakterija mlečne kiseline i proizvodnju DS. Takođe, uzimajući u obzir poreklo supstrata, upotreba industrijskih nusproizvoda na ovaj način ima veliku ekološku važnost.

Predmet istraživanja u okviru ove doktorske disertacije je optimizacija proizvodnje enzima dekstransaharaze, dobijene iz novog soja Leuconostoc mesenteroides T3, na komercijalnoj podlozi i na otpadnom supstratu, kao i imobilizacija ovog enzima na nanočestice TiO2 i unutar agar-agar filmova u cilju proizvodnje dekstrana i glukooligosaharida (GLOS). Novi soj Lc. mesenteroides T3, izolovan iz zrna vodenog kefira korišćen je kao mikroorganizam za proizvodnju i izolaciju dekstransaharaze. Optimalna podloga za maksimalnu proizvodnju enzima dobijena je variranjem uslova gajenja (kao što su temperatura i aeracija) i komponenata podloge (koncentracija saharoze i izvora azota). Dekstransaharaza dobijena u optimalnoj podlozi je zatim izolovana i prečišćena taloženjem uz pomoć polietilen glikola 400 (PEG). Karakterizacijom delimično prečišćene dekstransaharaze utvrđeno je da pokazuje najveću aktivnost na 30 0C pri pH vrednosti 5,4 i da dodatak jona Mn2+ ima najveći pozitivan uticaj na povećanje aktivnosti od čak 73 %. Zimogramom je pokazano da dekstransahraza ima molekulsku masu od oko 180 kDa. Karakterizacija proizvoda reakcije katalizovane dekstransaharazom iz radnog mikroorganizma, kada u reakcionoj smeši nisu bili prisutni akceptorski molekuli, izvršena je FTIR spektroskopijom. Potvrđeno je da je molekul glukana koji je sintetisan dekstransaharazom iz Lc. mesenteroides T3 dekstran u kome su molekuli glukoze dominantno povezani α-(1→6) glikozidnim vezama. Za proizvodnju dekstransaharaze na otpadnom supstratu, izluženi repin rezanac (SBP, od engl. sugar beet pulp), veličina zrna 500 μm‒800 μm, korišćen je kao nosač za imobilizaciju ćelija Lc. mesenteroides T3 dok je melasa bila izvor ugljenika, vitamina i mineralnih materija. Ispitivanjem uticaja ne-tretiranog (SBP-N) i izluženog repinog rezanca nakon tretmana sa NaOH (SBP-NaOH) na proizvodnju dekstransaharaze, bolji se pokazao repin rezanac nakon alkalnog pretretmana. Optimalan sastav podloge za maksimalnu proizvodnju dekstr, The scope of this doctoral dissertation is the optimization of the production of dextransaccharase (DS) enzyme, obtained from a new strain Leuconostoc mesenteroides T3, using a commercial medium and a waste substrate, as well as the immobilization of this enzyme on TiO2 nanoparticles and within agar-agar and application of these imobilised systems for the production of dextran and glucooligosaccharides (GLOS). A new strain Lc. mesenteroides T3, isolated from water kefir grains, was used as a microorganism for production and isolation of enzyme dextransucrase. The optimal medium for maximum enzyme production was defined by varying the growth conditions (such as temperature and aeration) and the broth/medium components (sucrose concentration and nitrogen source). The enzyme, purified by polyethylene glycol 400 (PEG) fractionation, displayed the maximum activity at 30 0C and pH 5.4. The addition of Mn2+ cation caused a signficant change of DS activity, with 73% increase. Zymogram analysis showed the presence of DS of approximately 180 kDa. Characterization of the dextransucrase catalyzed reaction product, when acceptor molecules were absesent from the reaction mixture, was performed by FTIR spectroscopy. The results indicated that a glucan molecule synthesized by dextransucrase from Lc. mesenteroides T3 was dextran in which glucose molecules were dominantly linked by α- (1 → 6) glycosidic bonds. For the dextransaccharase production on waste substrated, sugar beet pulp (SBP) of grain size 500 μm ‒ 800 μm, was used as a carrier in order to immobilize Lc. mesenteroides T3 cells, while molasses was a source of carbon, vitamins, and minerals. The influence of SBP in native form and after treatment with NaOH (SBP-NaOH) on dextransucrase production was investigated. The optimal substrate composition for maximum dextransucrase production was determined by varying the concentration of molasses, SBP-NaOH and sucrose. The maximum enzyme activity of 2,02 IU/ml was obtained on the

The aim of this study was the development of a non-toxic, biosynthetic antimicrobial agent which selectively acts on only one type of microorganism, and preserves the microbiota. Antimicrobial performance of biosynthesized silver nanoparticles (Ag NPs) by horsetail (Equisetum arvense L.) extract was examined against Gram-negative bacteria Escherichia coli and Gram-positive bacteria Staphylococcus aureus, as well as yeasts Candida albicans and Saccharomyces boulardii. Also, the cytotoxicity of Ag NPs was examined toward pre-osteoblast cells. The synthetic conditions—concentration of extract, temperature, and pH—were optimized to prepare silver colloids with different particle size distributions and long-term stability. The obtained samples were characterized using transmission electron microscopy, X-ray diffraction analysis, and absorption spectroscopy. The smaller-sized Ag NPs (~ 10–20 nm), prepared at a lower temperature (20 °C), showed better antimicrobial performance against E. coli compared to larger ones (~ 40–60 nm), prepared at high temperature (100 °C). On the other hand, both samples did not display any toxic action against bacteria S. aureus, or yeasts C. albicans and S. boulardii. Non-cytotoxic behavior of Ag NPs toward pre-osteoblast cells was observed for the concentrations of silver ≤ 2.25 and ≤ 4.5 mg L−1 for 10–20 and 40–60 nm-sized Ag NPs, respectively. Biosynthesized Ag NPs by horsetail extract display selective toxic action against E. coli at the ecologically acceptable concentration level. © 2019, Springer Science+Business Media, LLC, part of Springer Nature.

The antibacterial ability of in situ prepared nanometer-sized silver particles, immobilized in agar-agar films, was studied as a function of the concentration of co-dopant, magnesium ions. Content of inorganic components in hybrid films was determined using inductively coupled plasma optic emission spectroscopy, and found to be low (<2 wt.-%). Morphology of prepared hybrid films, studied by transmission electron microscopy, revealed the presence of non-agglomerated and randomly distributed 10–20 nm silver nanoparticles (Ag NPs) within the agar-agar matrices. Fourier-transform infrared spectroscopy indicated the distinct chemical interaction between Ag NPs and polymer chains. Thermogravimetric analysis, as well as the determination of tensile strength, Young's modulus, and elongation at break showed improvement of thermal stability and mechanical properties of agar-agar matrices upon the incorporation of Ag NPs due to high compatibility between the hydrophilic organic component and inorganic components. The complete microbial reduction of Gram-positive bacteria Staphylococcus aureuswas observed for all agar-silver films, while satisfactory results were observed for Gram-negative bacteria Pseudomonas aeruginosa (≥99.6%). The release of Ag+ ions is suppressed by the increase of the concentration of Mg2+ ions and it was found to be significantly smaller (≤0.24 ppm) than the harmful ecological level (1 ppm). © 2019 Elsevier Ltd

The TiO2 based hybrid supports with different functional groups (amino, glutaraldehyde or epoxy) were prepared and their influence on immobilization of dextransucrase (DS) was studied. Novel synthetic route for surface modification of TiO2 with amino and glutaraldehyde groups was developed taking advantage of charge transfer complex (CTC) formation between surface Ti atoms and salicylate-type of ligand (5 aminosalicylic acid (5-ASA)). The proposed coordination of 5-ASA to the surface of TiO2 powder and optical properties of CTC was presented. The pristine TiO2 and amino functionalized TiO2 have higher sorption capacity for DS (12.6 and 12.0 mg g(-1), respectively) compared to glutaraldehyde and epoxy activated supports (9.6 and 9.8 mg g(-1) respectively). However, immobilized enzyme to either glutaraldehyde or epoxy functionalized TiO2 have almost two times higher expressed activities compared to pristine TiO2 support (258, 235 and 142 IU g(-1), respectively). Thermal stability of enzyme immobilized on glutaraldehyde and epoxy functionalized supports was studied at 40 degrees C, as well as operational stability under long-run working conditions in repeated cycles. After five cycles, DS imobilized on glutaraldehyde activated support retained almost 70% of its initial expressed activity, while, after five cycles, performance of DS immobilized on epoxy activated support was significantly lower (15%).

The aim of this study was to develop dextran-based edible films plasticized by sorbitol. In order to optimise the film-forming formulation, response surface methodology was used. The influence of dextran and sorbitol concentration on the mechanical and water vapour barrier properties of obtained films was investigated. The results showed that both parameters exhibited significant effect on the water vapour permeability of a film. Both dextran and sorbitol concentration had significant influence on tensile strength and elongation at break, whereas only sorbitol concentration had significant effect on Youngs modulus. After optimisation by desirability approach, it was found that a film made of 3.40 wt% of dextran and 20.43 wt% of sorbitol showed the lowest water vapour permeability and the highest tensile strength and elasticity.

In this work, antimicrobial bioinspired films made from chitosan incorporated with triclosan were investigated. The tensile strenght of these films were in the range of 33 and 39 MPa, which presented satisfied mechanical stability comparable to the synthetic-based packages commonly used in industry. The addition of triclosan enhanced thermal stability and antimicrobial activity of chitosan films against Escherichia coli and Staphylococus aureus. Results obtained in this work demonstrated that chitosan/triclosan films could be potentially used as an eco-sustainable package in healthcare sector to prevent infections/contaminations.