Biotechnologia Acta

16-22

Authors: S. R. Budnikov, O. O. Soldatkin, A. L. Kukla, I. I. Khomenko, S. V. Dzyadevych, O. P. Soldatkin

Research was aimed at the optimization of the urease biosensor for analysis of heavy metals and determining the opportunities of its reactivation. A differential pair of gold interdigitated electrodes deposited on a ceramic substrate was used as the conductometric transducer. As a bioselective membrane served urease, coimmobilized with bovine serum albumin by glutaraldehyde cross-linking on the surface of conductometric transducer. 1.0 mM urea was selected as an optimal substrate concentration for the inhibition analysis of heavy metals. The biosensor was tested for its sensitivity to different heavy metals, the calibration curves were plotted. The proposed biosensor was shown to have high reproducibility of signals prior and after inhibition, the measurement error was less than 3%. It was proved a possibility of reactivation of the bioselective membrane after urease irreversible inhibition by heavy metals, using the ethylenediamine tetraacetic acid solution. The optimum conditions of reactivation, i.e. the dependence of its level on time and concentration of heavy metals, were determined. The additional stage, post-inhibition reactivation, was shown to increase significantly the selectivity of biosensor determination of heavy metals.

32-40

Authors: M. D. Lootsik, R. O. Bilyy, M. M. Lutsyk, R. S. Stoika

The aim of research was the evaluation of blood clotting activity and hemostatic effect of chitosan specimens using developed set of laboratory tests and preparation of chitosan with high blood clotting activity. Chitosan specimens were powdered to the particle size less than 0.25 mm and characterized by in vitro tests including determination of coagulation time of heparinized blood, bulk density of material, swelling of particles in water and Tris-buffered saline, as well as in vivo tests based on estimation of efficacy in controlling hemorrhage from wounds in mice and rats. Using of the developed tests it was revealed that available chitosan specimens (reagent grade, dietary supplements, samples prepared in laboratory) are hemostatically inactive as compared with Celox preparation. We developed a method of activation of above mentioned chitosan specimens to active hemostatic preparations. Some characteristics of chitosan that are significant for blood clotting activity are described. It was demonstrated that high molecular weight chitosan specimens are more effective as hemostatic agents comparing with low molecular weight chitosan specimens.

48-55

Authors: V.V. Sokolik, S.М. Shulga

The purpose of the study was the investigation of curcumin liposome form effect on angiotensinconverting enzyme activity, cytokines and mnestic features of rats with experimental model of Alzheimer’s disease. In the animals with intrahippocampal injection of Аβ42_Human, nasal therapy with curcumin liposome form was used. Cytokine concentration and angiotensin converting enzyme activity in brain regions (cerebral cortex and hippocampus) and in blood serum as well as indicators of conditioned avoidance response were registered. It was found that as a result of curcumin therapy the rats with Alzheimer’s disease had suppressed cytokine and angiotensin converting enzyme activities and recovered mnestic indices. Nasal therapy with curcumin liposome form gave reduction of angiotensin-converting enzyme activity and anti-cytokine effect in the target regions of the brain (cerebral cortex and hippocampus), which helped the rats mnestic features and memory recovery.

63-70

Authors: A. K. Gulevsky, Yu. S. Akhatova, А. А. Sysoev, I. V. Sysoeva

To study the bioenergy indicators of leykokontsentrate cells after cryopreservation and the possibility of their recovery after incubation in a medium with a low-molecular fraction from cow cord blood were the aim of research. Leykokontsentrate was obtained from donor blood by sedimentation; cryopreservation was carried in the slow freezing regimen with 5% dimethylacetamide; amount of ATP, ADP and AMP was determined by chemiluminescence; glycogen — by cytochemical method. Evidence of energy disbalance of leukoconcentrate cells after cryopreservation was obtained. It was shown that the cord blood low-molecular fraction (0.15 mg/ml) activated glycogenolysis and increased contents of ATP, ADP and AMP in leukoconcentrate cells after cryopreservation. It was also shown that the cord blood low-molecular fraction contained energy substrates and metabolites, hormones and macro- and micronutrients. Use of the low-molecular fraction (below 5 kDa) from cord blood as a component of rehabilitating medium for frozen-thawed leukoconcentrate cells contributed to improvement of their energy status, which was manifested as augmentation in the total adenylic pool and glycogenolysis activation.

77-86

Authors: О. Yu. Маyorova, L. R. Hrytsak, N. М. Drobyk

The objective of the research was to develop the technology of introduction of the obtained by microclonal propagation Gentiana lutea L. plants into conditions in situ. Methods of cultivation of plant objects in vitro were used. There were chosen optimal conditions for rooting G. lutea shoots obtained through microclonal propagation in vitro: МS/2 medium with twice decreased concentration of NH4NO3 without vitamins and sucrose supplemented with 3 g/l of mannite and 0.05 mg/l kinetin, and agar (4 mg/l) in combination with perlite (16 g/l) used as a maintaining substrate; or the nutrient medium (MS/2 without vitamins and smaller concentration of N4NO3) with gradual decrease of carbohydrates from 10 g/l to 2 g/l, and further rooting experimental shoots in tap water. Rooted plants were adapted to conditions ex vitro through planting them into flowerpots with soil and gradual changing hothouse regime for exposed one. The share of adapted to in situ conditions plants (21% after a year of planting) proves the suggested method to be efficient and promising. There was suggested this technology is the most efficient ones for revival of disturbed G. lutea populations that includes repatriation of rooted and adapted to ex vitro conditions plants obtained through microclonal propagation in vitro.

19-26

Authors: А. V. Bazalii, A. V. Vorotnikov, P. A. Тiurin-Кuzmin, V. А. Тkachuk, S. V. Коmisarenko, L. B. Drobot

The aim of this study was to design the expression vector encoding fluorescent sensor of hydrogen peroxide HyPer fused with adaptor protein Ruk/CIN85 as well as to check its subcellular distribution and ability to sense hydrogen peroxide. It was demonstrated that in transiently transfected HEK293 and MCF-7 cells Ruk/CIN85-HyPer is concentrated in dot-like vesicular structures of different size while HyPer is diffusely distributed throughout the cell. Using live cell fluorescence microscopy we observed gradual increase in hydrogen peroxide concentration in representative vesicular structures during the time of experiment. Thus, the developed genetic construction encoding the chimeric Ruk/CIN85-HyPer fluorescent protein represents a new tool to study localized H2O2 production in living cells.

39-48

Authors: A.A.Peterson, M. Yu. Vasylenko, N. A. Matvieieva, M. V. Kuchuk

Determination of the presence of the recombinant fusion protein (ESAT6-Ag85B(ΔTMD)-6His) and its accumulation level in duckweed plants (Lemna minor L.) was the aim of the research. ESAT6 and Ag85B are secretory proteins of Mycobacterium tuberculosis and are considered as potential candidates for development of new vaccine against tuberculosis (TB). Transgenic duckweed plants were obtained previously by Agrobacterium rhizogenes-mediated transformation and possessed fusion gene sequence esxA-fbpBΔTMD. Specific polyclonal antibodies were produced in immunized mice to identify levels of accumulation of TB antigens in plants. Recombinant antigen used for mice immunization was obtained in our laboratory by expression in E. coli. Western blot analysis revealed the recombinant tuberculosis antigen ESAT6-Ag85B(ΔTMD)-6His in extracts from transgenic L. minor plants. The level of accumulation of the protein corresponds to 0.4-0.5 µg protein per 1 g of fresh weight of plant. Additionally, the accumulation of recombinant protein was investigated in lyophilized transgenic plants after 1.5 year storage. Duckweed plants accumulating a recombinant analogue of M. tuberculosis secretory proteins can be used for development of plant-based edible vaccines.

55-63

Authors: L. A. Kotyuk

The aim of the paper is to research biochemical composition of Hyssopus officinalis L. (Lamiaceae) in relation to plant age and phenological growth stage under conditions of Ukrainian Polissya, bin order to determine the optimal harvest dates of the herbal material and its application spheres. The raw material samples under analysis were cut at various growth stages: the vegetative, budding, blooming, ripening stages. To study the hyssop oil composition, areal parts of H. officinalis were used. The composition analysis was aimed at determining absolute dry matter (by drying samples at 105 °C up to the constant mass), “crude” cellulose, amounts of protein, fats, calcium, potassium, phosphorus, ascorbic acid, carotene, discernible sugars and tannins and essential oil. The present study has proved that in the plant ontogenesis the amount of essential oil, obtained from H. officinalis areal parts, does not markedly decrease: volatile oil yield in plants of the first, second and third years of life amounted to 1.007%, 0.75% and 0.71% respectively. The composition of volatile oil in the plants of the first year of life reveals 46 components, of which pinocampone (53.73%), isopinocampone (4.66%) myrtenol (9.35%) and camphor (3.86%) prevailed. In H. officinalis volatile oil of the third year 30 components were identified, the prevailing of which were isopinocampone (44.43%), pinocampone (35.49%), myrtenol (5.26%), germacrene D (3.15%), pulegone (2.93%) and bicyclogermacrene (1.35%). We could observe the change in the quantitative and qualitative composition of H. officinalis volatile oil throughout the entire vegetation period. Thus, in the phase of vegetative growth one can identify 25 compounds, the most predominant being elemol (33.25%), germacren D (21.59%) and bicyclogermacrene (15.78%). In the phase of blossoming 30 components can be identified, a high amount of isopinocampone and pinocampone (44.43% and 35.49%) and somewhat lover amount of myrtenol (5.26%), pulegone (2.93%) and bicyclogermacrene (1.35%) can be noted. During the fruit-bearing period 21 compounds with the prevalence of elemol (44.46%), bicyclogermarcren (10.30%), germacren D (5.86%), spatulenol (4.36%), β-eudesmol (4.34%), α-eudesmol (4.04%) and γ-eudesmol (3.92%) can be identified. H. officinalis raw material from one-year-old plants is to be used in food industry, whereas plants collected at the blooming stage are preferable for cosmetics and perfumery.

71-77

Authors: A. R. Banya, O. Y. Karpenko, V. I. Lubenets, V. I. Baranov, V. P. Novikov, O. V. Karpenko

The aim of the work was to study the influence of rhamnolipid biocomplex and ethylthiosulfanilate on field pea and sorghum plants when growing in petroleum contaminated soil. Plant seeds were treated with solutions of rhamnolipid biocomplex or ethylthiosulfanilate (0.01 g/l) before planting and grown in containers with soil artificially contaminated with petroleum (5, 8 and 10%). Effect of rhamnolipid biocomplex and ethylthiosulfanilate was determined by the determination of growth (weight, length plants) and biochemical parameters (content of photosynthetic pigments, hydrogen peroxide, malondialdehyde). The stimulating effect of rhamnolipid biocomplex and ethylthiosulfanilate on growth parameters of field pea was shown: the shoot mass has significantly increased on 39%, root mass – on 26% if compared with the control. For sorghum somewhat smaller increase of growth parameters was observed. Under the influence of rhamnolipid biocomplex and ethylthiosulfanilate the content of photosynthetic pigments in field pea and sorghum has also increased. It was determined that the action rhamnolipid biocomplex and ethylthiosulfanilate promoted the decrease of indicators of plant oxidative reactions if compared with control: the content of hydrogen peroxide – in average on 15% and 16%, malondialdehyde – on 13.5% and 16% respectively. The results of the study testify to the effectiveness of rhamnolipid biocomplex and ethylthiosulfanilate as growth stimulators for field pea and sorghum, as well as improvement of adaptive capability of plants to unfavorable conditions. It creates the prospects of their application as effective and ecologically safe substances for the intensification of contaminated soil phytoremediation.

86-92

Authors: I. A. Blayda, T. V. Vasilyeva, L. I. Slyusarenko, V. F. Khytrych

The study's aims were to isolate and establish pure cultures of prevailing microorganisms from the aboriginal consortium in coal fly ash (FAAC), describe their physiology, biochemistry and practically-useful properties, and compare the efficiency of bioleaching metals from fly ash using pure cultures and the consortium. Through enrichment cultures on standard media we isolated pure cultures of the microorganisms which were then preliminarily identified using standard techniques. This allowed us to isolate from FAAC pure cultures of three prevailing strains of mesophilic and moderately thermophilic acidophilic chemolithotrophic bacteriae, belonging to Acidithiobacillus, such as Acidithiobacillus ferrooxidans, and Sulfobacillus. The strains exhibited high oxidative activity in leaching the rare metals Gallium and Germanium, as well as some heavy metals, from fly ash substrate. A comparison of oxidative activity of the isolated strains and the aboriginal consortium under mesophilic conditions led to the conclusion about advantage of consortium, because it had arisen from syntrophy of microbes in the community. This should be taken into account at the developing of bacterial preparations that are optimal for the technogenic substrate.

103-11

Authors: Govorukha V.M., Tashyrev O.B.

The aim of the study was regulation of metabolism of Citrobacter freundii Ml-31.1/1 on the basis of thermodynamic calculations. The essence of metabolism regulation was in ability to perform two opposite processes, namely mobilization and immobilization of iron compounds, depending on the culture conditions by single strain C. freundii Ml-31.1/1. The following methods were used: potentiometric measuring of pH and Eh values, colorimetric determination of microbial biomass increase and changes of iron compounds concentration, gas chromatography. The possibility of theoretically grounded regulation of iron transformation pathways was experimentally confirmed. Single culture (C. freundii Ml-31.1/1) provided both immobilization and mobilization of iron compounds due to specific conditions for microbial growth in nutrient medium.. Obtained regularities are the basis for assessing the role of microorganisms in iron biogeochemical cycles. This data could be used as a basis of effective biotechnologies of iron extraction from depleted deposits and water purification from iron compounds.

21-39

Authors: T. P. Pirog, A. D. Konon, I. V. Savenko

It was shown literature and own experimental data concerning the use of microbial surface active glycolipids (rhamno-, sophoro- and trehalose lipids) and lipopeptides for water and soil purification from oil and other hydrocarbons, removing toxic heavy metals (Cu2+, Cd2+, Ni2+, Pb2+), degradation of complex pollution (oil and other hydrocarbons with heavy metals), and the role of microbial surfactants in phytoremediation processes. The factors that limit the use of microbial surfactants in environmental technologies are discussed. Thus, at certain concentrations biosurfactant can exhibit antimicrobial properties and inhibit microorganisms destructing xenobiotics. Microbial biodegradability of surfactants may also reduce the effectiveness of bioremediation. Development of effective technologies using microbial surfactants should include the following steps: monitoring of contaminated sites to determine the nature of pollution and analysis of the autochthonous microbiota; determining the mode of surfactant introduction (exogenous addition of stimulation of surfactant synthesis by autochthonous microbiota); establishing an optimal concentration of surfactant to prevent exhibition of antimicrobial properties and rapid biodegradation; research both in laboratory and field conditions.

53-62

Authors: Komisarenko S. V., Volochnyuk D. M., Shymanskyy I. O., Ivonin S. P., Veliky M. M.

The aim of the study was to investigate the effectiveness of nitrogen-containing bisphosphonates synthesized as promising substances for correction of mineral metabolism in osteoporosis. The study was carried out on a model of alimentary osteoporosis that was characterized by hypocalcaemia, hypophosphatemia, decreased 25-Hydroxyvitamin D3 content in blood serum and severe bone tissue demineralization (reduced ash content and mineral components). It was found that synthesized novel nitrogen bisphosphonates (pyrazole-containing analogues), like reference drugs — metylene bisphosphonate (disodium salt of metylene bisphosphonic acid) and alendronate (4-amino-1-hidroxybutyliden bisphosphonate), inhibit with the different efficiency demineralization of the bone tissue and increase the mineral metabolism in rats with alimentary (nutritional) osteoporosis that was assessed by the marker parameters of bone formation. In particular, drug administration (bisphosphonates І-12, І-40, І-42) resulted in elevation of calcium and phosphate levels and decreased the total activity of alkaline phosphatase and its isoenzymes in blood serum. The ash content and the levels of calcium and phosphorus in the ash of tibia and femur bones were shown to be markedly elevated. Bisphosphonate І-12 has shown more profound antiresorbtive activity and ability to correct mineral metabolism in alimentary osteoporosis, including such of reference drugs. It was found a significant decrease of 25-Hydroxyvitamin D3 content in the serum that is considered as a profound vitamin D3 deficiency associated with nutritional osteoporosis. As it was not compensated by bisphosphonates, we suggest that further investigations should be directed to the combined use of both: bisphosphonates as inhibitors of osteoclast activity that diminish bone resorption and vitamin D3 as a key regulator of bone remodeling process and osteosynthesis activator.

71-81

Authors: Burlaka O. M., Pirko Ya. V., Kolomys O. F.2, Smertenko P. S., Glazunova V. O., Konstantinova T. E., Yemets A. I., Blume Ya. B.

To select the most effective methods for functionalizing carbon nanotubes and the to compare the ability of a number of biological molecules (plasmid DNA, ATP, deoxyribonucleoside triphosphates, bovine serum albumin, compounds of vitreous humor extract and sodium humate) to interact noncovalently with carbon nanotubes and mediate their dispersion in an aqueous medium was conducted was the aim of research. Properties of carbon nanotubes-biomolecules conjugates were characterized using ultraviolet, visible and near infrared spectroscopy, Raman spectroscopy, transmission electron and atomic force microscopy. Formation of stable aqueous polydisperse colloidal systems of single-walled and multi-walled non-covalently functionalized carbon nanotubes was shown. The appearance of extended functionalization covering consisting of biomolecules on the surface of carbon nanotubes was demonstrated. Changes in morphology and structure of carbon nanotubes, namely shortening and the appearance of defects in sp2-hybridized surface caused by functionalization were revealed. As a result, the range of molecules of biological origin suitable for noncovalent functionalization of carbon nanotubes was chosen, with correspondence to the specific use in plant biotechnology and the properties of formed complexes were characterized.

92-97

Authors: Tigunova O. O., Shulga S. M.

The aim of this work was to enlarge accumulation of butanol from switchgrass Panicum virgatum L. biomass using strains-producers obtained from grounds and silts of Kyiv lakes. The objects of the study were strains of C. acetobutylicum ІМВ B-7407 (IFBG C6H), Clostridium acetobutylicum IFBG C6H 5М and Clostridium tyrobutyricum IFBG C4B from the "Collections of microbial strains and lines of plants for food and agricultural biotechnology" of the Public Institution "Institute of Food Biotechnology and Genomics" of the National Academy of Sciences of Ukraine. Gas chromatography was used to determine the alcohol concentration at the stage of solvent synthesis. To determine the effect of butanol precursors during cultivation, butyric, lactic and acetic acids were used. Optimization of processing parameters, which was based on the needs of cultures, allowed us to increase the yield by 20 and 50% for the initial and mutant strain respectively. Using synthetic precursors (such as lactic, butyric and acetic acid) during cultivation increased total concentration of butanol by 1.7 times. To optimize the process, a study was carried out using acetone- butyl grains. Using of acetone-butyl grains in concentrations up to 60% does not affect the synthesis of butanol by C. acetobutylicum IFBG C6H 5M. Increasing the concentration of grains led to decrease in accumulation of butanol. Almost double increase in accumulation of the target product (butanol) was achieved using two-stage fermentation and/or precursors of synthesis. It was shown the possibility of using acetone-butyl grains in fermentation. As a result the mass fraction of the waste was reduced.

108-112

Authors: Kuznietsova H. M., Ogloblya O. V., Cherepanov V. V., Prylutskyy Yu. I., Rybalchenko V. K.

The toxicity of С60 fullerene, traditional cytostatic cisplatin and С60 fullerene-cisplatin complex on honeybee Apis mellifera L. toxicity estimation test system was assessed. Water-soluble pristine C60 fullerenes were nontoxic for honeybee when consumed with the food in doses equivalent nontoxic and effective ones for mammalian. Cisplatin toxicity for honeybee in the doses exceed the same for mammalian in 2 times was observed as fallows: honeybee 56% death occurred after consumption of 60 mg/kg of bee weight. С60 fullerene-cisplatin complex proved to be more toxic for honeybee in comparison with free cisplatin and caused honeybee 50% lethality after consumption of 40 mg/kg bee weight.

122-127

Authors: Isayenkov S. V., Miam A., Maathuis F. J. M.

The results of screen and analysis of two OsTPKb rice mutant lines were described. The phenotypes and growth rate level of homozygous mutant plants of both rice lines were estimated. The electron microscopy of aleurone layer from forming seeds was performed. The OsTPKb mutant plants demonstrate lower growth rate in comparison with wild type plants. The loss of function OsTPKb mutations invariably led to (semi)sterile rice plants. The functional disruption of OsTPKb channel has negative impact on plant growth and development. It might completely change the cell morphology of aleurone layer.

135-140

Authors: Suslova O. S., Rokitko P. V., Bondar K. M., Golubenko O. O., Tashyrev A. B.

The aim of this study was to determine homeostasis quantitative parameters (maximum permissible concentrations of xenobiotic and types of cell response) of microbial communities from cave clays influenced by p-nitrochlorobenzene. It was determined the ability of cave bacteria to transform xenobiotic. General bacteriological methods were used as well as gas chromatography-mass-spectric method. Chemoorganotrophic karst caves microbial communities isolated from Mushkarova Yama (Podolia, Ukraine) and Kuybushevskaya (Western Caucasus, Abkhazia) were highly resistant to p-nitrochlorobenzene. For Mushkarova Yama representative strain Rhodococcus erythropolis P3 the influence of p-nitrochlorobenzene (in concentration range 50–300 mg/l) on physiological parameters of bacterial cells was shown. The efficiency of p-nitrochlorobenzene degradation was proportional to the decreasing of redox potential. Thus, cave microorganisms could potentially be used to create new environmentally friendly biotechnologies, for example enterprises for wastewater treatment from nitrochloraromatic compounds.

28-44

Authors: Skivka L. M.

Literature data and own author experiments concerning the influence of microbiota on the immune system are summarized. The mechanisms of the diversification of immune response to pathogenic and commensal microorganisms are described. Effect of microorganisms of normal flora on innate and adaptive immunity is characterized. Human inflammatory diseases associated with microbiota disorders are reviewed. Biological properties of probiotic preparations are discussed in context of its modulatory effect on inflammatory response. Prospects of use of immunomodulatory potential of probiotic microorganisms are being analyzed.

56-66

Authors: Kvesitadze E.

The work is devoted to the isolation, purification, determination of taxonomical characteristics and application in soil improvement and other biotechnological processes halophilic microscopic fungi strains isolated from saline soils of Eastern Georgia (middle part of South Caucasus), where their existence is maximally supposed. In all soilclimatic zones the dominate forms of spread fungi are genera Aspergillus, Penicillium and Fusarium, followed by Trichoderma and Mucor. Other genera are met less intensively. The genera Aspergillus is widely spread in chestnut soils and in chernozem, in green forest soils the genera Penicillium is prevailing. The salinity of soil, lake or any other objects from which the isolation of microscopic fungi is performed greatly determines halophilisity of isolated strains. Finally, the collection of halophilic microscopic fungi has been created accounting 96 isolates of extreme halophiles, halophiles and week halophiles.

78-88

Authors: Sokolik V. V., Shulga S. M.

The purpose of this study was to investigate the effect of curcumin on cytokine response and angiotensin-converting activity in terms of intrahippocampus administration of ß-amyloid peptide in rats. Animals with experimental model of Alzheimer’s disease received nasal therapy aqueous solution of curcumin. Recorded concentration of cytokines (interleukin-1ß, interleukin-6, interleukin-10, tumor necrosis factor-α) and angiotensin-converting activity in the parts of the brain (cerebral cortex and hippocampus) and blood serum, as well as indicators of conditional reflex escapers action. Install a reduction in cytokine response intrahippocampus administering Aß42_Human and angiotensin-converting activity inhibition in the brain but not in the blood serum of animals under the action of curcumin. Recorded improvement in cognitive performance in rats with a model of Alzheimer’s disease as a result of curcumin therapy. Nasal treatment with an aqueous solution of curcumin has anti-inflammatory effect in the targeted parts of the brain (сerebral cortex and hippocampus), but inhibits angiotensin-converting activity.

95-103

Authors: Babenko L. P., Lazarenko L. M., Demchenko Z. K., Konarbaeva O. A., Veccio G. Lo, Spivak M. Ja

The aim of work was to determine antibacterial activity of Lactobacillus casei ІМV В-7280 probiotic strain on the experimental urogenital tract infection of mice. The influence of intravaginal and/or per os administration of this strain once per day during 7 days on the microflora of vagina, kidneys and intestinal contents of Staphylococcus aureus 8325-4 infected mice was studied. It was established, that in cases of experimental staphylococcal infection of urogenital tract L. Casei IMV B-7280 had effective antagonistic activity against S. aureus 8325-4 and opportunistic bacteria. After L. casei IMV B-7280 introduction into infected mice reduction or complete elimination of S. aureus 8325-4 in vagina, kidneys and intestinal contents in different periods of observation was established. Under the influence of L. casei ІМВ В-7280 the number of coliform bacteria, streptococci and staphylococci in the vagina was normalized, and fungal flora — decreased even in comparison with intact mice. Normalization of kidneys microflora was also observed. In the intestinal contents of infected mice trea ted with L. casei IMV B-7280 the number of streptococci did not change, staphylococci number decreased, but the number of fungal and coliform flora remained relatively low during the observation period. L. casei IMV B-7280 probiotic strain is promising to create immunobiotics with antibacterial action, which can be used for the prevention and treatment of urogenital infections caused by opportunistic microorganisms.

110-115

Authors: Bal-Prylypko L. V., Leonova B. I.

The purpose of this work was to study reduction of sodium nitrite in cooked sausages by adding of the optimized amount of denitrifying microorganisms to the bacterial preparation maintaining quality characteristics of the product. To develop biotechnology of boiled sausages «Naturel» we selected bacterial preparation based on nitrite-reducing strains of Staphylococcus carnosus and S. carnosus ssp.utilis. It was used generally accepted and special methods. The content of total pigments and nitrozopigments was determined by a method based on the extraction of meat pigments by aqueous acetone; color stability of final products was evaluated as the difference in optical density of nitroso pigment extracts before and after exposure (40 min) of the sample under the light source; analytical processing of the experimental data was carried out using modern software; quantitative evaluation of color characteristics was performed in the RGB using a multifunctional device Epson Stylus TX400. Mathematical modeling was carried out on the basis of full factorial experiment such as 22, the optimization was performed by Box–Wilson. According to the study, using of the bacterial preparation based on nitrite-reducing strains of Staphylococcus carnosus and S. carnosus ssp. utilis in biotechnology of boiled sausages «Naturel» has a positive effect on the formation of the complex of required color characteristics of final products (for prototypes of sausages the index redness was 1. 61 times higher compared to the control). Degradation of sodium nitrite and formation of nitroso pigments were intensified that improved the stability of color during the storage (the index of color fastness of experimental cooked sausages was higher by 19%). The results of performed investigations illustrate the possibility of production of cooked sausages with a minimized content of synthetic food additives and ingredients.

123-128

Authors: Govorukha V. M., Havrylyuk O. A., Tashyrev O. B.

The aim of the work was quantitative determination of Fe(III)-reducing bacteria in natural ecosystems of the Antarctic, the Arctic, the Dead and the Black Sea, middle latitude (Ukraine, Abkhazia) and the equatorial zone (Ecuador). It was used the method of quantitative determination of microorganisms by McCready and the colorimetric method for determination of Fe(II) compounds . Results. The systemic study of the number of Fe(III)-reducing bacteria of both hemispheres in the ecosystems of six geographic regions was carried out for the first time. High number of Fe(III)-reducing bacteria in natural ecosystems was experimentally shown. The number of Fe(III)-reducing bacteria ranged from 1.1•102 to 2.8•107 cells/g of absolutely dry sample. Conclusions. The presented data showed that Fe(III)-reducing bacteria are an integral part of natural ecosystems and can significantly affect the biogeochemical cycles of iron and carbon compounds transformation.

26-35

Authors: N. A. Matvieieva

The review focused on the data concerning current state in the field of Compositae “hairy” roots and transgenic plants construction using A.tumefaciens- and A. rhizogenes-mediated transformation to obtain biologically active compounds, including recombinant proteins. The article presents data on the results of genetic transformation of Cichorium intybus, Lactuca sativa, Artemisia annua, Artemisia vulgaris, Calendula officinalis, Withania somnifera and other Compositae plants as well as studies on the artemisinin, flavonoids, polyphenols, fructans and other compounds accumulation in transgenic plants and roots. The data show that the use of biotechnological approaches for construction of "hairy" roots and transgenic plants with new features are of great interest. The possibility of increase in the accumulation of naturally synthesized bioactive compounds and recombinant proteins production via A. tumefaciens and A. rhizogenes-mediated transformation have been shown. In vitro cultivation of transgenic plants characterized by high level of bioactive compounds accumulation and synthesis of recombinant proteins makes it possible to obtain guaranteed pure raw material. Using of biotechnological approaches preserved natural populations of plants is particularly important for rare and endangered plant species.

52-57

Authors: A. M. Taranenko, L. O. Sakhno, B. V. Morgun, M. V. Kuchuk

The purpose of research was to analyze 10 developed at the Institute of Cell Biology and Genetic Engineering lines of rapeseed to confirm the presence and functionality of the transferred transgene CP4 epsps, as well as the differences among those lines from registered transformation events GT73 and GT200 (Monsanto). During the study extraction of total rapeseed DNA, PCR analysis, electrophoretic separation and visualization of amplicons in agarose gel were conducted, as well as testing of green plants for resistance to glyphosate in greenhouse. The structural difference among 7 transgenic lines from registered transformation events GT73 and GT200 was revealed. Plants showing the presence of synthetic CP4 epsps sequence were resistant to the herbicide in a closed soil. The uniqueness of the obtained transformation events was confirmed, as well as the prospect of using them in breeding.

69-77

Authors: S. V. Pykalo, M. О. Zinchenko, S. I. Voloshchuk, O. V. Dubrovna

The direct and step-type in vitro selection with application of selective system based on low molecular mannitol and the selection of callus lines of winter triticale being resistant to simulated water deficit were carried out. As a result, from line 38/1296 and variety Obriy respectively, 5 and 4 resistant callus lines were identified that had a high survival rate on the selective medium with 0.6 M mannitol and maintained morphogenetic potential. From the resistant lines plant regenerants were induced and their rearing, rooting and transfer to in vivo conditions were optimized and followed by testing to water deficit tolerance. During the testing under simulated 3-weeks drought it was revealed that plant regenerants derived from resistant lines were characterized with reliably higher relative water content and free proline content as compared with non-resistant plants. A step-type in vitro selection was more effective, because resulted from the selection more resistant callus forms were identified. Evaluation of plant regenerants of triticale obtained from resistant calli showed increased level of their tolerance to water deficit.

84-90

Authors: E. V. Маtseliukh, L. А. Safronova, L. D. Varbanets

Proteases from probiotic strains of the genus Bacillus, just like the antibiotics, bacteriocins and other hydrolytic enzymes, are one of the main factors that determine their biological activity. The aim of this work was to study the synthesis and biochemical properties of proteases from two strains Bacillus amyloliquefaciens subsp. plantarum UCM B-5139 and UCM B-5140 that included in the probiotic Endosporin. The cultivation of strains was carried out in flasks under rotating for two days. The influence of physico-chemical parameters of the reaction medium on proteolytic activity was studied on partially purified protease preparations. Lytic activity was determined by turbidimetric method. On the second day of cultivation B. amyloliquefaciens subsp. plantarum UCM В-5139 and UCM В-5140 synthesized the metal-dependent peptidase and serine protease, respectively. The optimum conditions of their action were the following: temperature 37–40 °C and pH 6.5–7.0. Isolated proteases are able to lyse the living cells of Staphylococcus aureus and Candida albicans. Thus we demonstrated that B. amyloliquefaciens subsp. plantarum UCM B-5140 and UCM B-5139, included in the probiotic veterinary preparation Endosporin, produced proteolytic enzymes that hydrolyze the native insoluble proteins (elastin, fibrin and collagen). These enzymes belong to the group of neutral metal-dependent and serine proteases. They are active under physiological conditions against gram-positive bacteria and yeasts. The application of these proteases in biotechnology is considered.

96-106

Authors: V. V. Turov, I. M. Todor, N. Yu. Lukyanova, T. V. Krupskaya, A. P. Ugnivenk, V. F. Chehun

The aim was to study the changes in the structure of intracellular water in the intact rat liver and liver with Guerin carcinoma. The state of water in lyophilized liver tissue of rats – intact and affected by Guerin carcinoma at different stages of tumor development was studied by low-temperature 1H NMR spectroscopy. It is shown that in progress of tumor the contribution from the water clusters whose size is 3–12 nm is grows. It was established that in intact tissue present 6 types of water clusters capable of dissolving different amounts of trifluoroacetic acid. In the liver with Guerin carcinoma clusters of strongly associated water more uniform, and their dissolving power with respect to trifluoroacetic acid and the electron donor molecules significantly decreased. These results are important for elucidating the mechanism of action of water on the development of neoplastic processes.

113-119

Authors: A. Yu. Galkin, 2, A. G. Komar, A. A. Grigorenko

In article we analyzed national and international regulations concerning the quality and safety of medical devices for in vitro diagnostics. We discussed the possibility of a partial application of the recommendations of the State Pharmacopoeia of Ukraine to this type of product. The main guiding regulatory documents establishing requirements for quality and safety tools for the serological diagnosis products are The technical regulation on medical devices for the diagnosis in vitro, DSTU ISO 13485 “Medical devices. Quality management system. Regulatory requirements”, and DSTU ISO/IEC 17025 “General requirements for the competence of testing and calibration laboratories”. Similar requirements of the State Pharmacopoeia of Ukraine which are used for drug standardization can not be directly applied to the medical devises for in vitro diagnostics due to a number of features, namely, the serological diagnosis products pre-designed to determine the unknown concentration of a particular analyte in a biological material, the diagnostic kits has to include the control samples (internal standard systems) that need to be calibrated. It was determined following parameters of bioanalytical standardization and validation characterization for of qualitative (semi quantitative) test-kits for serological diagnosis: precision (convergence, intralaboratory precision and reproducibility), diagnostic and analytical specificity, diagnostic sensitivity. It’s necessary to inspect additional parameters for quantitative test-kits such as accuracy (precision), linearity, analytical sensitivity and range.