Oral Abstracts

Improving the suitability of milk for cheese production

Svetlana Valerevna Pavlenkova^a, Galina Pavlovna Shuvaeva^a, Anna Aleksandrovna Tolkacheva^a, Lidiya Aleksandrovna Miroshnichenko^b, Tatyana Vasilevna Sviridova^a, and Olga Sergeevna Korneeva^a

^aDepartment of Biochemistry and Biotechnology, Voronezh State University of Engineering Technologies, Voronezh, Russia

^bLTD Russkaya oliva, Russia email address: [email protected]

The suitability of milk for cheese production is an important factor in the production of high quality cheese. The suitability of milk for cheese production is characterized by the following parameters: high protein content, including casein, high fat content, and the ability of milk to coagulate with rennet. The urgent problem of improving the quality of milk in the stall period is the preparation of balanced diets. Amaranth is a promising crop, because its green mass surpasses other fodder plants in amino acid balance. Our team of authors found that when high-protein amaranth silage was included in the diets of dairy cows, the protein content in milk increased by 0.2% (including the casein content increased by 0.33%); the mass fraction of fat increased by 0.38% in the prototype compared with the control, which indicates an increase in the suitability of milk for cheese production. At the same time, 14.3% less rennet and 5.5% less milk were spent on the production of 1 kg of cheese from our milk. Work supported by the state task No. 40.4149.2017/PCh.

Investigation of the physiological and biochemical changes in two carrot genotypes at different boron concentrations

Mehmet Hamurcu^a, Ayse Humayra Omay^a, Zuhal Zeynep Avsaroglu^a, Fevzi Elbasan^a, Ceyda Ozfidan Konakcı^b, Evren Yıldıztugay^c, Anamika Pandey^a, Mohd Kamran Khan^a, Erdogan Esref Hakki^a, and Sait Gezgin^a

^aFaculty of Agriculture, Department of Soil Science and Plant Nutrition, Selcuk University, Konya, Turkey

^bFaculty of Science, Department of Molecular Biology and Genetics, Necmettin Erbakan University, Konya, Turkey

^cFaculty of Science, Department of Biology, Selcuk University, Konya, Turkey email address: [email protected]

Boron is a major source of nutrients for plants that causes various physiological and biochemical changes under deficiency or excess conditions. In particular, boron plays an important role in the developmental stages of plants but it also affects the yield under toxic conditions. In this study, two different carrot genotypes, one tolerant and another sensitive to boron toxicity were used The plants were grown under controlled hydroponic conditions and three different B treatments, 0, 0.5, 1, and 2 mM boron were applied to compare with the control conditions. The growth rate, proline content, enzymatic antioxidants (SOD-POX-APX), non-enzymatic antioxidants, like ascorbate (tAsA), reduced and oxidized glutathione (GSH and GSSG), dehydroascorbate (DHA), and also, lipid peroxidation (MDA) were investigated. The results showed that growth parameters were decreased regardless of the differences in the boron doses. The non-enzymatic antioxidant activity was induced due to the increase in the boron doses of both genotypes and this was directly affected by the MDA accumulation. Though SOD enzyme activity in sensitive genotype was decreased in 1 mM boron treatment, POX activity increased. However, SOD and POX activity in the tolerant genotype of all the groups was found to be higher than the control group.

‘Influenza a/H1sw2009 virus circulation among swine populations in the republic of Kazakhstan (2019)’

Nailya Klivleyeva^a, Nuray Ongarbayeva^b, Nurbol Saktaganov^a, Tatyana Glebova^a, Galina Lukmanova^a, Mira Shamenova^a, and Assem Baimukhametova^a

^aLLP Scientific Production Center for Microbiology and Virology, Almaty, Republic of Kazakhstan

^bAl-Farabi Kazakh National University, Almaty, Republic of Kazakhstan email address: [email protected]

Influenza virus causes an acute, highly contagious disease due to its constant variability, prevalence, and availability of natural reservoirs. Swine can act as a ‘melting pot’ or an intermediate host in which reassortment takes place between avian, porcine, and human genes. Reassortment contributes to the emergence of new antigenic variants of influenza virus. Monitoring of influenza A virus circulation in the swine population is necessary for timely anti-epidemic and anti-epizootic measures to prevent epidemics, pandemics, and epizootics.

183 nasopharyngeal swabs were collected from swine during the 2018 spring season at livestock farms and private farmsteads located in the Kostanay and Aktobe regions. The primary screening of biological samples in RT-PCR using AmpliSens reagents (Moscow, Russian Federation) demonstrated the presence of influenza A virus genetic material in five nasopharyngeal swabs (2.73% of the total number of examined samples). Subtyping showed the presence of influenza A/H1N1 virus RNA in three samples (1.64%), influenza A/H3N2 virus RNA in two samples (1.09%). Therefore, the RT-PCR results obtained with the samples collected in spring 2018 from the northern and western regions of the Republic of Kazakhstan indicate the co-circulation of influenza A/H1N1 and A/H3N2 viruses in the swine population.

Acacetin inhibits NRF2 signalling pathway leading to decreased ovarian cancer proliferation

Ibrahim Hamza Kankia^a and Yusuf Deeni^b

^aDepartment of Biochemistry, Umaru Musa Yar’adua University, Katsina, Nigeria

^bDepartment of Biotechnology and Microbiology, Federal University, Dutse, Nigeria email address: [email protected]

Acacetin an organic compound of flavonoids has been shown to inhibit cancer proliferation. Nuclear factor (NRF2) regulates the normal cellular growth and is a determinant of cancer initiation and progression. Overexpression of NRF2 is reported in ovarian cancer leading to resistance of numerous chemotherapeutic agents. Human ovarian cancer cell lines, OVCAR3 and OVCAR4 were maintained in RPMI 1640 media supplemented with 10% foetal bovine serum (FBS) and 1%pen/strep in 5% CO2 and incubated at 37 °C. Before experimental treatments, cells were grown for 24 h in media and then treated with either tert-Butylhydroquinone (tBHQ; Sigma) or Acacetin (Carbosynth) to a final concentration as required with media. Following this, luciferase assay, cytotoxicity assay, ROS detection, Glutathione assay and immunoblotting were performed. The study demonstrated that Acacetin inhibited the NRF2 activity, which led to decreased cell growth, increase in ROS level and depletion of total glutathione in all the cell lines. These data suggest that Acacetin may inhibit the cell growth of ovarian cancer cells through NRF2 inhibition and could be a novel avenue Acacetin induced NRF2 inhibition, which leads to inhibition of cancer cell growth.

Gene expression alternation in multi-drug resistant E. coli cultivated with a nanomolecular iodine-containing complex inducing antibiotic resistance reversion

Sergey Shilov^a, Ilya Korotetskiy^a, Oleg Reva^b, Tatyana Kuznetsova^a, Natalya Suldina^a, and Aleksandr Ilin^a

^aScientific Center for Anti-Infectious Drugs (SCAID), Almaty, Kazakhstan

^bCentre for Bioinformatics and Computational Biology (CBCB), University of Pretoria, Pretoria, South Africa email address: [email protected]

Currently, resistant bacterial infections appear at an increasing rate, whereas the development of new antibiotics has dramatically declined over the past two decades. The purpose of this study was to gene expression profiling of multidrug-resistant strain Escherichia coli ATCC® BAA-196™ to assess transcriptomic alterations during the cultivation of the culture with a new antimicrobial nanomolecular iodine-containing complex FS-1 developed in SCAID. Total RNA was isolated using a commercial KIT according to the manufacturer’s recommendations and sequenced using IonTorrent technology. It was shown that the cultivation of E. coli for 10 passages with FS-1 leads to a general inhibition of the bacterium’s metabolism. The vast majority of the detected changes were associated with a decrease in metabolic activity. Significant suppression of the main genes involved in the tricarboxylic acid cycle and glyoxylate shunt was recorded. In view of the general suppression of the TCA, it was expected that the biosynthesis of amino acids should decrease also. Negative regulation of genes involved in the β-oxidation of fatty acids, cyoABCD oxidase complex, NADH dehydrogenase I components, and sdhCDAB operon encoding quinone oxidoreductase (SQR) also were suppressed. This general suppression of important metabolic pathways may explain the restored sensitivity to antibiotics by FS-1.

Isolation and identification of clinical isolates that cause nosocomial infections in Almaty during 2018–2019

Natalya Zubenko^a, Ilya Korotetskiy^a, Tatyana Kuznetsova^a, Sergey Shilov^a, Auyes Myrzabayeva^a, Oleg Reva^b, and Lyudmila Ivanova^a

^aScientific Center for Anti-Infectious Drugs (SCAID), Almaty, Kazakhstan

^bCentre for Bioinformatics and Computational Biology (CBCB); University of Pretoria, Pretoria, South Africa email address: [email protected]

The problem of nosocomial infections is currently in the focus of science and health care. Despite anti-epidemic measures, the frequency of outbreaks, mortality, and the cost of treatment of drug-resistant nosocomial infections continue to increase. The purpose of this study was to identify clinical isolates in order to characterize and create a collection of bacterial isolates causing nosocomial infections in hospitals in Kazakhstan to study the distribution of multidrug resistant pathogens. Primary identification of isolates was performed on selective and differential diagnostic media using generally accepted methods, followed by 16S rRNA typing. The antibiotic sensitivity was performed by the disc diffusion method using 12 the most common antibiotics. In total, 19 strains showing an extended pattern of antibiotic resistance were selected from clinical isolates of nosocomial infections from hospitals in Almaty, Kazakhstan. The selected strains include a representative of Enterobacteriaceae, Staphylococcaceae, Streptococcaceae, Pseudomonadaceae, and Enterococcaceae. The clinical isolates were characterized by a varying degree of sensitivity to antimicrobial drugs. What is worrying, several strains were resistant to the reserved group of antibiotics such as azithromycin, meropenem, and imipenem. The whole-genome sequencing of 11 isolates was performed using the Ion Torrent PGM platform. The results are deposited in the NCBI database.

Collagen nanowires loaded with ginger essential oil

Mariana Daniela Berechet^a, Maria Stanca^a, Roxana Constantinescu^a, Demetra Simion^a, Olga Niculescu^a, Dana Gurau^a, Alecsandra Miletic^b, Branca Pilic^b, and Maria Rapa^c

^aINCDTP – Division Leather and Footwear Research Institute, Bucharest, Romania

^bFaculty of Technology, Novi Sad, Serbia

^cPolitehnica University of Bucharest, Bucharest, Romania email address: [email protected]

In this research nanowires of hydrolyzed collagen loaded with ginger essential oil were obtained by electrospinning. Collagen hydrolysate was obtained from bovine leather by-products processed by alkaline-enzymatic hydrolysis in the presence of 10% CaO and 0.4% Alcalase 0.4 L, characterized by 60.40% dry matter, 82.43% protein, 14.67% total nitrogen and 1623 cP viscosity. DLS analysis of collagen hydrolysate showed particle sizes of 109.3 nm (52.3%), 9.7 nm (22.3%) and 1.9 nm (14.3%). GC-MS analysis showed a ginger essential oil rich in limonene 22.63%, camphene 22.32%, α-pinene 12.35%, cineole 11.29% and zingiberene 10.14%. Collagen mixed with 10% ginger essential oil was electrospun on waxed paper support at 9.4 kV, with a debit of 550 µL/h and the collector-needle distance was 13 cm. SEM analysis showed an average diameter of 464.2 nm in collagen nanowires and 665.5 nm in collagen nanowires loaded with ginger essential oil with a porous 3D structure and a large surface related to volume. Microbiological analysis of obtained nanowires loaded with ginger essential oil showed 96.8% antibacterial activity against Staphylococcus aureus, 73.33% against Escherichia coli and 95.51% antifungal activity against Candida albicans. Collagen nanowires loaded with ginger essential oil can be used in the medical, pharmaceutical, cosmetic or niche fields.

The present work was carried out under Short Term Scientific Mission (STSM) ECOST-STSM-Request-CA15114-44124, ‘Antimicrobial nanofiber-based membranes from biopolymers obtained from leather industry by-products’ and with support of Faculty of Technology Novi Sad, University of Novi Sad, Serbia, Grant Holder, and under 4 N/2019_ PN 19 17 01 02 and 6PFE/2018_ PERFORM-TEX-PEL projects, funded by the Ministry of Education and Research.

The sensitivity of the cells of green microalgae ankistrodesmus sp. B-11 to cadmium

Asemgul Sadvakasova, Bolatkhan Zayadan, Meruert Bauenova, Kenzhegul Bolatkhan, Huma Balouch, and Zhuldyz Mustapaeva

Department of Biotechnology, Al-Farabi Kazakh National University, Almaty, Kazakhstan

email address: [email protected]

Cadmium, a toxic heavy metal, is lethal to living organisms for its adverse effects such as inhibiting enzymes activity and damaging the cell’s membrane structures. Microalgae are widely used for biomonitoring, since the rapid reactivity of its photosynthesis machinery to the pollutants reflects its higher sensitivity. The effect of low concentrations of cadmium ions on the growth, photosynthesis and ultrastructure of microalgae Ankistrodesmus sp B-11cells was studied. A significant decrease in the number of Ankistrodesmus sp B-11 cells was observed in medium with cadmium at concentration of 0.005–0.02 mg/l, with complete ceased growth at 0.05 mg/l. Ultrastructural changes in the arrangement of stroma thylakoids, membrane divergence with formation of inter-thylakoid spaces and a significant increase in vacuolization of cells were noted. The MREA-2 fluorimeter showed that cadmium ions inhibit electronic transport in PhSII. There was a decrease in the quantum yield of electronic transport in PhSII (φEo), the performance index (PIABS), and a slowdown in the recovery of the P700 pigment, as well as an increase in energy dissipation (DI0/RC) and ΔpH-dependent non-photochemical quenching (qE). Being the most sensitive parameter, the Performance Index (PIABS) can be good indicator of the toxic action of cadmium ions on microalgae.

Production and purification of polyclonal antibody: experiences from the lab benchtop

Sehime G. Temel^a,b,c

^aFaculty of Medicine, Department of Histology & Embryology, Bursa Uludag University, Bursa, Turkey

^bFaculty of Medicine, Department of Medical Genetics, Bursa Uludag University, Bursa, Turkey

^cDepartment of Translational Medicine, Institute of Health, Bursa Uludag University, Bursa, Turkey, email address: [email protected]

Immunohistochemical localization of different peptides in the central nervous system and peripheral organs is important for functional studies. For this reason, new peptides are being discovered day by day and it is possible to synthesize these peptides in sufficient quantities to immunize different animal species. The amino acid sequence of many neuropeptides or receptor proteins shows homology between species. This feature provides ease of use of an antibody created in different animal species. However, this homology between species and small peptide size is responsible for low antigenicity. Although there are few researchers who obtain a specific antibody by generating an immune response with unconjugated peptides, the vast majority of success has been achieved by immunization by binding the peptide with a large immunogenic carrier protein. These carriers are usually from serum proteins. An agent capable of forming covalent binding between the amino acids of two different molecules is required for conjugation of carrier proteins to the peptide. In this talk, the polyclonal antibody that we produce in the rabbits against the kainate-selective glutamate receptor subunit, kainate-2, Gonodotropin Releasing Hormone and Alpha-1 B adrenergic receptor, etc. will be discussed. The obtained antibodies were then purified and their specificity was investigated.

Study of the effect of a extract Kazakhstani desert plant Cistanche deserticola on the viability and regenerative ability of human skin fibroblast cells

Mariana Ualikhanovna Sarsembayeva^a, Kanat Nurullaevich Sarsenbayev^b, Kulzada Mergenbayevna Lakhanova^c, and Gulzhaina Baikonyssovna Alpamyssova^a

^aSouth Kazakhstan Pedagogical University, Shymkent, Kazakhstan

^bL.N. Gumilyov Eurasian National University, Astana, Kazakhstan

^cYassawi International Kazakh-Turkish University, Turkestan, Kazakhstan email address: [email protected]

We studied the effect of the extract from Kazakhstan desert plant Cistanche deserticola on wound healing and increase the vitality of human skin fibroblasts. The biological activity of Cistanche deserticola is five times higher than that of ginseng. Fibroblast cultures of the human embryo were used. Alcohol extract of Cistanche deserticola was used at a concentration of 30 mg/1 ml. The study of the wound healing properties of the extract was carried out on white mice. It was shown that Cistanche deserticola increases the viability of human fibroblast cells within the concentration 5 to 500 μg/ml till to 50% viability. It was revealed that when applying liquid soap, sealing liquid soap with petroleum jelly with the addition of an extract of Cistanche deserticola, the process of wound growth is faster. The use of Cistanche deserticola extract will help improve the healing processes of wounds, enhance the regeneration processes of fibroblast.

Development of methods for humus accelerated enrichment of old plowland blacksoil

Kanat Nurullaevich Sarsenbayev^a, Mariana Ualikhanovna Sarsembayeva^b, Kulzada Mergenbayevna Lakhanova^c, Yerubay Baibekovich Baibekov^c, and Elmira Begimbayevna Zhapparbergenova^b

^aL.N. Gumilyov Eurasian National University, Astana, Kazakhstan

^bSouth Kazakhstan Pedagogical University, Shymkent, Kazakhstan

^cYassawi International Kazakh-Turkish University, Turkestan, Kazakhstan email address: [email protected]

On the vast territory of Kazakhstan, there are practically no untouched lands with normally developed humus and high productivity. This is especially characteristic of Central Kazakhstan, where after intensive exploitation, land productivity has plummeted. The soil of this region is characterized by a weak regenerative ability. Therefore, industrially used land will be in its former state for a long time.

The main function of the soil is the complete degradation or conservation of plant and animal waste that is not metabolizable by microorganisms. These include lignin, bitumen and keratogenic structures. Microorganisms use the majority of compounds for nutrition and they are responsible for the disposal of dead organisms. However, in the case of complex, complex compounds, they cannot utilize them. These compounds in the form of humic compounds are deposited in the soil. A technology is being developed to regulate the process of humus formation using simple and widely available methods of reclamation: the introduction of straw and alfalfa plant residues and the use of biologically active substances to accelerate the transformation of organic matter in the soil.

The study of ferule smelly in Turkestan region of Kazakhstan

Mariana Ualikhanovna Sarsembayeva, Gulzhaina Baikonyssovna Alpamyssova, Gulmira Satybaldievna Khalikova, Gulmira Turdyevna Zhussipova, and Asia Yrsymbetovna Mamytova

South Kazakhstan Pedagogical University, Shymkent, Kazakhstan email address: [email protected]

One of the promising species in Kazakhstan, which is in great demand as a medicinal raw material, is smelly ferula. The species grows on sandy, gravel-clay soils of deserts of Kazakhstan, Uzbekistan, Turkmenistan, Tajikistan, and Afghanistan. Despite the centuries-old experience of practical use, smelly ferula has not been fully studied. It is believed that it is an ancient Persian medicinal plant that spread from Iran to many other countries. The differences between them are so significant and population polymorphism is so high that a special study is needed on the biochemistry, morphology and medicinal properties of individual populations.

Scientific novelty lies in the fact that the implementation of research in full compliance with the priorities of scientific, technical and socio-economic development of Kazakhstan. The data obtained made it possible to determine the localization and polymorphism of ferula smelly populations; classify populations of a species, establish distribution patterns and reserves of underground organs. When analyzing the phytochemical composition of all parts of plants, data were obtained on the content of useful substances in them. The composition of gum, alkaloids and phenolic compounds was studied, the biological activity and acute toxicity of extracts and individual ferula compounds were determined.

Perspectives of antiviral defense in crops through CRISPR/CAS9 genome editing

Olga Kershanskaya, Aralbek Rsaliyev, Gulvira Esenbaeva, Gulzhamal Mukiyanova, Darya Nelidova, Zarina Sadullayeva, and Sergey Nelidov

Institute Plant Biology and Biotechnology SC, MES RK, Almaty, Kazakhstan email address: [email protected]

Plant–virus interactions based-studies have contributed to increase our understanding on plant resistance mechanisms, providing new tools for crop improvement. The new genome editing systems CRISPR/Cas9 is revolutionizing the scope of tools available to confer virus resistance in plants. The task of our research is to explore the perspectives of antiviral defense in barley through CRISPR/Cas9 genome editing – knock and modification of eIF4E – the key gene of complex of initiation of protein translation. We have created a genetic constructs for CRISPR/Cas9 editing the genomes of five local Kazakhstan barley cultivars with eIF4E barley gene, NCBI, GenBank: AJ704454.1. For this we have elaborated sgRNA target sequences using in silico program CRISPRdirect. sgRNA was synthesized and cloned into the CRISPR-plant Sigma-Aldrich vector at EcoRi and HindIII sites under the WhU6 promoter, Cas9 nuclease under the 35S promoter, and contained hptII and nptII as marker genes. Created constructs were introduced into barley cells via agrobacterium transformation technique, which we have elaborated and patented. Confirmation of edited events was carried out by PCR, DNA sequencing, testing of edited plants on virus and micropathogens resistance. Using CRISPR/Cas9 editing technology, we have developed eIF4E non‐transgenic barley mutants that exhibit resistance to multiple economically important viruses.

The study of Ferula foetida in Turkestan region of Kazakhstan

Mariana Ualikhanovna Sarsembayeva, Gulzhaina Baikonyssovna Alpamyssova, Gulmira Satybaldievna Khalikova, Gulmira Turdyevna Zhussipova, and Asia Yrsymbetovna Mamytova

South Kazakhstan pedagogical university, Shymkent, Kazakhstan email address: [email protected]

One of the promising species in Kazakhstan, which is in great demand as a medicinal raw material, is smelly ferula. The species grows on sandy, gravel-clay soils of deserts of Kazakhstan, Uzbekistan, Turkmenistan, Tajikistan, and Afghanistan. Despite the centuries-old experience of practical use, smelly ferula has not been fully studied.

The scientific and practical significance of the work is to identify bioecological, morphological, genetic and biochemical, features of various smelly ferula populations growing in the southern region of Kazakhstan. Morphometric, chemical and biochemical methods revealed differences between the populations of ferule smelly in the composition of essential oils, phenolic compounds, and organic acids. The composition of DNA amplicons shows significant differences between smelly ferula populations that grow in vivo. Smelly ferula populations most productive in biomass accumulation were identified. A number of chemical compounds that can be used to create new drugs and food additives have been identified and identified. The studies carried out suggest the influence of growing conditions on the formation of genetic differences in the studied populations.

Study of the effect of Cistanche deserticola extract on the viability and regenerative capacity of human skin fibroblast cells

Kanat Nurullaevich Sarsenbayev^a, Mariana Ualikhanovna Sarsembayeva^b, Kulzada Mergenbayevna Lakhanova^c, Gulzhaina Baikonyssovna Alpamyssova^b, and Gulnar Makulbekovna Sartbayeva^b

^aL.N. Gumilyov Eurasian National University, Astana, Kazakhstan

^bSouth Kazakhstan Pedagogical University, Shymkent, Kazakhstan

^cYassawi International Kazakh-Turkish University, Turkestan, Kazakhstan email address: [email protected]

We studied the effect of the extract from Kazakhstan desert plant Cistanche deserticola on wound healing and increase the vitality of human skin fibroblasts. The biological activity of Cistanche deserticola is five times higher than that of ginseng. Fibroblast cultures of the human embryo were used. Alcohol extract of Cistanche deserticola was used at a concentration of 30 mg/1 ml. The study of the wound healing properties of the extract was carried out on white mice. It was shown that Cistanche deserticola increases the viability of human fibroblast cells within the concentration 5 to 500 μg/ml till to 50% viability. It was revealed that when applying liquid soap, sealing liquid soap with petroleum jelly with the addition of an extract of Cistanche deserticola, the process of wound growth is faster. The use of Cistanche deserticola extract will help improve the healing processes of wounds, enhance the regeneration processes of fibroblast.

Does DL-sulforaphane induce oxidative stress in breast cancer cell lines in high glucose media?

Günnur Demircan^a, Sule Beyhan Ozdas^a, Ummugulsum Guzelsoy^b, and Demet Akin^c

^aMedical Faculty, Medical Biology and Genetics, Demiroglu Bilim University, Istanbul, Turkey

^bYildiz Technical University, Istanbul, Turkey

^cBahcesehir University, Medical Faculty, Pharmacology, Istanbul, Turkey email address: [email protected]

Impaired glucose metabolism is an important risk factor for cancer formation by increasing oxidative stress at the cellular level. Therefore, the diabetes is an important risk factor in the progression of cancer. In many studies, it has been shown that sulforaphane (SFN), which is obtained from broccoli, has anticancer properties. In this study, we aimed to demonstrate the antioxidant and anticarcinogenic effects of SFN in breast tissue healthy (MCF-10A) and cancer cell lines (MDA-MB 231 and MCF-7) incubated in a high glucose media. Cells were incubated for 24, 48 h in high glucose medium using different SFN concentrations. Oxidative stress responses in the cell lysates were evaluated with total oxidant and antioxidant measurement kits. Anti-apoptotic (Bcl2) and apoptotic protein (Apaf1,Caspase3) expressions were evaluated by Western Blot analysis. İntracellular insulin levels were shown by ELISA. According to our study, SFR induces apoptosis via a reactive oxygen species (ROS)-dependent mechanism. The production of ROS has been postulated to be a key mechanism by which SFN induces apoptosis. SFR acts strengthening the cellular defenses against oxidative damage and promoting the removal of carcinogens. There is substantial and compelling evidence for SFN mediated apoptosis in both of breast cancer cells, but this may depend on SFN dose.

Development of a feed additive made of the vegetative mass of amaranth with the properties of a phytobiotic and its use for obtaining import-substituting food products

Ivan Sapelkin, Svetlana Pavlenkova, Anna Derkanosova, Alisa Korotaeva, Aleksey Drannikov, and Vladimir Sharshov

Voronezh State University of Engineering Technologies, Voronezh, Russia email address: [email protected]

This project is aimed at studying the properties and scientific evaluation of the effectiveness of the use of a feed additive made of the vegetative mass of amaranth, as a functional component of compound feed to increase the meat productivity and immune status of Mulard duck, as well as the production of import-substituting products. Due to the increase in population, income and urbanization, the demand for poultry meat is constantly growing. When breeding Mulard duck, it is possible to obtain several types of gourmet foods: foie gras liver, magret de canard and meat. They are a source of macronutrients and trace elements, vitamins, have high biological value, have a unique amino acid composition. The inclusion of duck meat and liver in the diet will satisfy the human need for animal proteins and polyunsaturated fatty acids. It is assumed that the introduction of a protein feed additive from the amaranth vegetative mass into the feed composition will have a positive effect on the meat productivity and immune status of Mulard duck. The main advantage of feeding herbal supplements will be the possibility of its use as a taste and stimulating appetite, while the poultry will not be exposed to any risks.

Development of polyclonal antibodies specific to wheat ribosomal protein S6 kinase 1

Sanzhar Dossanovich Alybayev, Bauyrzhan Smailov, Aibanu Baktygaliyeva, and Amangeldy Bissenbaev

Scientific Research Institute of Biology and Biotechnology Problems, Al-Farabi Kazakh National University, Almaty , Kazakhstan email address: [email protected]

Mammalian ribosomal protein S6 kinase 1 (S6K1) is a key enzyme in the regulation of cell growth and proliferation. In contrast to animal S6K1, there are not many studies published on plant S6K1, despite the importance of their function. Wheat S6K1 (TaS6K1) protein has yet to be detected in wheat tissues, due to lack of the appropriate molecular tools for characterization of TaS6K1 function and protein interactions in signal transduction. This study is focused on the production of polyclonal antibodies specific to TaS6K1 protein. To produce an anti-TaS6K1 antibody, we expressed and purified a soluble 6xHis-tagged TaS6K1. Purified recombinant TaS6K1 was injected into New Zealand white rabbits to generate polyclonal antiserum. TaS6K1 antibodies were purified by ammonium sulfate precipitation, followed by affinity chromatography using protein A-agarose beads. In wheat seedling extracts polyclonal antibodies specifically recognize a protein with a molecular mass close to predicted molecular weight of endogenous TaS6K1 protein. The purified polyclonal antibodies are demonstrated to be effective in immunodot assay, western-blot hybridization, and immunoprecipitation. Thus, the purified polyclonal antibody produced from the TaS6K1protein is a specific, sensitive, and could be a useful tool for future insight into downstream components of S6K1-regulatory mechanisms in wheat plants.

Ultrastructural change in epithelial cells of the proximal kidney during tumor growth and autophagy in epithelial cells of the proximal kidney

Assel Rakhmetova^a, Natalia Bgatova^b, and Sholpan Zhumadina^c

^aPavlodar State University. S. Toraigyrov, Pavlodar, Kazakhstan

^bResearch Institute of Clinical and Experimental Lymphology – Branch of the Federal State Budgetary Scientific Institution ‘Federal Research Center Institute of Cytology and Genetics of the Siberian Branch of the Russian Academy of Sciences’, Novosibirsk, Russia

^cJoint Stock Company Kazakh Agrotechnical University. S. Seifullina, Nursultan, Kazakhstan email address: [email protected]

In an experiment on CBA mice, structural changes in the kidneys were studied during the development of an experimental hepatocarcinoma transformed into the femoral region – 29. Swelling of the proximal kidney epithelial cells was noted during tumor growth, a decrease in mitochondria, an increase in primary and secondary lysosomes were also observed. The data obtained indicate that under conditions of distant tumor growth in the kidneys, changes are observed in the ultrastructure of the proximal kidney epithelial cells in animals with tumor growth and autophagy is formed to maintain intracellular homeostasis of the proximal epithelial cells.

Expression levels of genes involved in PI3K/AKT in PCOS mouse models

Pinar Tulay^a, Tuna Onal^b, and Seda Vatansever^b

^aNear East University, Department of Medical Genetics, Nicosia, Cyprus

^bFaculty Medicine, Department of Histology and Embryology, Manisa Celal Bayar University, Manisa, Turkey email address: [email protected]

Polycystic ovary syndrome (PCOS) is the most common heterogeneous endocrine-hormonal disorder. This syndrome has major characteristics of hyperandrogenism, polycystic ovaries and ovulatory dysfunction. It has been suggested that both environmental factors as well as genetic factors are involved in the development of PCOS. PI3K/AKT pathway has been shown to be involved during gametogenesis, especially during migration of primordial germ cell. The aim of this study was to examine the expression levels of genes involved in PI3K/AKT pathway in PCOS mouse models pre- and post-treatment. The expression levels of Irs, Akt1 and Akt2, mTorch and Pdpk1were investigated in four mouse models; PCOS, PCOS treated with clomiphene citrate, PCOS treated with clomiphene citrate, metformin and pioglitazone and the control group. Ovarian samples were obtained in each group and real time PCR was performed to investigate the gene expression levels. The histological analysis showed that normal follicle differentiation was obtained in the PCOS models following treatment. Akt1 expression level was significantly different between control and the PCOS group. Furthermore, both Akt1 and mTOR expression levels were significantly different in the control group and the treated groups. Thus, it may indicate that these two genes are involved in the follicular development.

Natural antimicrobial compounds on Staphylococcus aureus strains

Elif Kalpar and Meltem Yeşilçimen Akbaş

Department of Molecular Biology and Genetics, Gebze Technical University, Gebze‐Kocaeli, Turkey email address: [email protected]

Staphylococcus aureus is a common pathogenic microorganism found on different surfaces of the food industry and also produces dangerous en-terotoxins. Enterotoxins can cause serious food poisoning. Phytochemicals found in fruit wastes are extracted by different methods and may be used in many fields such as pharmaceutical, cosmetic and food industries as anti-microbial and antioxidant compounds. In this study, seven different extracts were prepared using different solvents (ethanol, methanol, ethanol:acid, methanol:acid, acetone or water). Antibac-terial activities of the extracts at different ratios (1/5, 1/10, 1/25) were investigated on S. aureus strain. In the agar well diffusion method, the highest inhibition zone diameter was 31 mm with methanol: acid-containing extract while the lowest inhibition zone diameter was 14 mm with acetone-containing extract against S. aureus bacteria. When the dilution rate of extracts increased, zone diameter decreased. Antibacterial activity was not observed with water-containing ex-tract. As a result, it was determined that fruit waste extracts could be have important potential antimicrobial compounds against S. aureus bacteria.

A preliminary results to propose the prognostic algorithm for PET/CT SUVmax values and BRCA1 and BRCA2 mutations in breast cancers

Semra Ozdemir^a, Fatma Silan^b, Yılmaz Akgun^c, Fulya Koc Ozturk^a, Ozturk Ozdemir^b

^aDepartment of Nuclear Medicine, Faculty of Medicine, CanakkaleOnsekiz Mart University, Canakkale, Turkey

^bDepartment of Medical Genetics, Faculty of Medicine, CanakkaleOnsekiz Mart University, Canakkale, Turkey

^cDepartment of General Surgery, Faculty of Medicine, CanakkaleOnsekiz Mart University, Canakkale, Turkey email address: [email protected]

BRCA1 and BRCA2 are tumor suppressor genes that play crucial role which predispose to breast and ovarian cancers. The maximum standardized uptake value (SUVmax), which is an indicator of metabolic activity obtained from PET/CT, has also an important role in determining the prognosis of tumoral cell aggressiveness and viability. In the current preliminary results we aimed to develop a prognostic algorithm for PET/CT SUVmax values criteria and BRCA1 and BRCA2 mutations in breast cancers. METHODS: Forty-three breast cancer patients who underwent PET/CT for primary staging were evaluated retrospectively. Germ-line DNA samples were genotyped for BRCA1 and BRCA2 genes by IonTorrent S5 NGS platform and analysed with IonReporter and Parseq-VariFind™AIP assay and compared to SUVmax values for the current results. RESULTS: In a cohort of patients, 2,3% BRCA1 VUS, 2,3% BRCA1 patogen mutation, 11,6% BRCA2 VUS, 6,9% BRCA2 patogen mutation were determined. There were no statistically significant differences in the SUV max value between the patients with and without BRCA1 (p = 0.281) and BRCA2 mutation (p = 0.094) carriers. CONCLUSION: According the current preliminary results, there is a limited association between BRCA mutations and SUVmax criteria that indicating poor prognosis for breast cancer. Results need to be supported by large scale of tumoral breast biopsies.

A case of Coffin-Siris syndrome with atypical phenotype caused by a novel De novo mutation in ARID1B gene

Burcu Albuz, Nihan Ecmel Akbas, Ahmet Kablan, Fatma Silan, and Ozturk Ozdemir

Department of Medical Genetics, Faculty of Medicine, Canakkale Onsekiz Mart University, Canakkale, Turkey email address: [email protected]

Coffin-Siris syndrome (CSS) is a genetic disorder classically characterized by sparse scalp hair, distinctive facial features, aplasia/hypoplasia of the distal phalanx or nail of the digits, developmental/cognitive delay of varying degree, hypotonia, hirsutism or hypertrichosis. Here we aimed to present a CSS case with a novel frameshift mutation in ARID1B gene and an atypical symptom, congenital hypothyroidism. A 3-years-old girl referred to our outpatient clinic because of global developmental delay. Physical examination revealed facial dysmorphism, muscle weakness, hypopigmentations in lower abdomen, global developmental delay, epilepsy and congenital hypothyroidism. Her parents were non-consanguineous and both healthy. After DNA isolation from peripheral blood sample, clinical exome analyse was performed using the Solution®(CES) kit and results analyzed with Sophia DDM 5.5.1 and IGV Software. We detected heterozygous frameshift c.2545dupG(p.Val849Glyfs*80) variant in ARID1B gene. This mutation wasn’t detected in her parents. No genetic variant associated with congenital hypothyroidism was detected. CSS is an autosomal dominant genetic disorder that affects multiple systems. Here we report a novel germline mutation in ARID1B gene in a CSS case with an atypical presentation, with congenital hypothyroidism. Current report contributed literature with a novel variant in ARID1B gene and expanding the clinical spectrum of phenotype.

Chromatin remodelling dysfunction and CHD2 related epilepsy: reports of two unrelated child

Ahmet Kablan, Burcu Albuz, Nihan Ecmel Akbas, Taner Karakaya, Volkan Sonmez, Mehmet Berkay Akcan, Fatma Silan, and Ozturk Ozdemir

Department of Medical Genetics, Onsekiz Mart University, Çanakkale, Turkey email address: [email protected]

Epilepsy is a common neurologic disorder. Genetic is responsible for nearly 20–50% of patients. More than 500 genes described for seizure disorders. Here we report two chromodomain helicase DNA-binding protein2 (CHD2) gene mutated male patients with unconsanguineous parents. Patient1 was born uneventfully at term, first seizure at 14 months old; neurodevelopmental delay, microcephaly recognized after 1 year of age. Neuroimaging and EEG was non-specific. After normal karyotyping, CES revealed heterozygous missense variant c.2095C > T p.(Arg699Trp) on CHD2 gene.ClinVar:likely pathogenic(2), VUS; ACMG:VUS, Mutation Taster:disease causing and was not found on 1000 G or ExaC. Patient2 was born at term, treated for respiratory stress, first seizure at 6 months of age. Mother and grandfather also epileptic and she had difficulties at school. Patient was delayed neurodevelopmentally, showed autistic behaviors in age 3–4.Imaging were normal. Although initial EEG was normal, later showed abnormal epileptic areas on right hemisphere furtherly. Karyotype, Microarray, FMR1, SCN1A gene tests resulted normal, CES revealed heterozygous missense variant c.3938G > A p.(Arg1313Gln) on CHD2 gene. ClinVar; VUS, ACMG; VUS, Mutation Taster disease causing and not found on 1000 G or ExaC. We identified two missense mutations in CHD2 gene in two unrelated pediatric patients with unknown seizure etiology.CHD2 associated with Epileptic encephalopathy. The current report highlighted the fact that epilepsy by failure of chromatin remodeling.

Investigation of macroporous bacterial cellulose scaffold as potential human artificial dermis

Aylin Basaran Eroglu^a, Gokhan Coral^a, and Sakir Necat Yılmaz^b

^aFaculty of Science and Letters, Department of Biotechnology, Mersin University, Mersin, Turkey

^bFaculty of Medicine, Department of Histology and Embryology, Mersin University, Mersin, Turkey email address: [email protected]

Bacterial cellulose (BC) is a pure form of extracellular cellulose synthesized the most effectively by Acetobacter xylinum. Due to its noticeable properties for example, three dimensional network structure, fine mechanical properties, high water holding capacity and biocompatibility, BC founds a wide variety of applications in many different industries, especially in biomedical fields such as wound dressings and scaffolds for tissue engineering. Hence, BC has a great potential for skin tissue engineering. However, the pore sizes of BC are not enough for cellular growth. This limitation restricts its practical usage as tissue engineering scaffold. Therefore BC needs some surface modifications by using freze-drying technique. The aim of this study is to create a BC scaffold to grow and proliferate human fibroblast cell line (ATCC, Bj-5ta) in the structure. Different BC concentrations were used to determine the most appropriate pore diameters for fibroblast cell growth. For this purpose, 0.25, 0.50, 0.75 wt% BC concentrations were tested. Cell viability was detected by using MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] kit and cells were visualized with SEM (Scanning Electron Microscope). Our results showed that 0.50 wt% BC was the optimal pore size for fibroblast cell growth.

Application of Fourier transform infrared spectroscopy for developing, monitoring and control of microbial bioprocesses

Simona Dzurendova, Boris Zimmermann, Achim Kohler, Valeria Tafintseva, Gergely Kosa, Kristin Forfang, Johanna Karin Hillevi Blomqvist, Anne Marie Langseter, and Volha Shapaval

Norwegian University of Life Sciences, Faculty of Science and Technology, As, Norway email address: [email protected]

Development of the novel bioprocesses based on the microbial valorization of various rest materials and by-products requires extensive screening studies for selecting microbial cell factories and optimization of process parameters. In order to fully assess the metabolic potential of microorganisms and evaluate sustainability of the bioprocess, chemical analysis of all intra- and extracellular metabolites is performed.

Fourier transform infrared spectroscopy (FTIR) allows non-invasive and rapid determination of a wide spectrum of intra- and extracellular metabolites as well as monitoring of substrate consumption. This can be achieved in a single step measurement, due to the fact that an IR spectrum contains fingerprint-type contributions from all kinds of chemical constituents. In a high-throughput screening studies, FTIR combined with the microcultivation systems provides a multi-analyte information on a high number of samples, with minimal amount of the biomass/culture media required for the analysis. Therefore, FTIR can greatly contribute to the development, monitoring and control of bioprocesses without time-consuming separation and determination of metabolites and media components. FTIR was shown to be reliable for monitoring of microbial lipid production and lipid extraction in all stages of the bioprocess development. Thus, FTIR could be considered as a powerful analytical method for evaluation of bioprocesses.

Biogenic synthesis and structural characterization of selenium nanoparticles from halophilic bacteria

Sena Kardelen Dinç^a, Nalan Oya San Keskin^a, and Figen Esin Kayhan^b

^aDepartment of Biology, Ankara Hacı Bayram Veli University, Ankara, Turkey

^bDepartment of Biology, Marmara University, Istanbul, Turkey email address: [email protected]

Since nanoparticle synthesis is expensive and dangerous with chemical methods, green synthesis which is cheaper, more efficient and safe has begun to be used as an alternative to traditional methods. Due to important functions in cellular metabolism of selenium, especially SeNPs have come to forefront between nanoparticles.

In this study, we report synthesizing of selenium nanoparticles using bacterial supernatant which is obtained from a halophilic bacteria Citricoccus sp. and optimum parameters for production. pH, reaction temperature, stirring speed and time parameters were study done. The optimum results of the study were found 8 pH, 24 h, 37 °C, 150 rpm. Selenium nanoparticles were characterized by UV-Vis Spectroscopy, Dynamic Light Scattering, Energy-Dispersive X-Ray Spectroscopy, Transmission Electron Microscopy, Scanning Electron Microscopy and Fourier-Transform Infrared Spectroscopy analyses. UV-Vis analysis showed characteristic broad peaks between 500–700 nm. In TEM and SEM analyses, spherical shaped selenium nanoparticles were observed. DLS analysis indicated that selenium nanoparticles have an average size of 300 nm. EDX analysis confirmed the presence of selenium. In FTIR spectral bands showed medium absorption peak at 1636 cm⁻¹ attributed to amide bands and broad absorption peak at 3336 cm⁻¹ correspond to O-H stretching in cell proteins. Results demonstrate that formation is originated from proteins.

Mechanical properties of three-dimensional microenvironment activate specific mechano-sensing circuits in MCF-7 breast cancer cells

Sara Sergio^a, Addolorata Maria Luce Coluccia^a, Enrico Domenico Lemma^b, Barbara Spagnolo^b, Daniele Vergara^a, Massimo De Vittorio^b, Ferruccio Pisanello^b, and Michele Maffia^a

^aDepartment of Biological Environmental Sciences and Technologies, University of Salento, Lecce, Italy

^bItalian Institute of Technology, Center for Biomolecular Nanotechnologies, Arnesano (Le), Italy email address: [email protected]

Cells sense and adapt to exogenous mechanical stimuli by activating nuclear transcription events and related phenotypic outcomes including proliferation, adhesion and sprouting. An interesting signalling player in this framework is β-catenin which can be aberrantly accumulated in the nucleus wherein it binds to and activates transcription factors of the LEF/TCF family. However, little is known about how mechanical cues are integrated into cells harbouring impaired mechanisms of β-catenin’s nuclear uptake and/or retention. We engineered 3D scaffold, by two-photon lithography, to study the influence of mechanical cues on MCF-7 cells which are known to fail in relocating β-catenin into the nucleus. We found that 3D microenvironments are per se sufficient to prime a TCF-dependent rescuing of β-catenin nuclear activity that boost cell proliferation and invasiveness. This, let us suggest that our system could provide a mechanism-based rationale to dissect further aspects of mechanotranscription in breast cancerogenesis and progression.

Lead tolerance of Aspergillus alliaceus isolated from green turtle nesting site

Esra Deniz Candan

Department of Medical Services and Techniques, Vocational School of Health Services, Giresun University, Giresun, Turkey email address: [email protected]

Industrial outputs of lead to the environment outweigh all natural sources, and lead reaches the terrestrial and aquatic environment. In this study, Sugözü Beaches, which host different industrial activities as a study area, and also an important nesting site for the green turtle (Chelonia mydas), were determined. The green turtles qualify for Endangered (EN) status under criteria of International Union for Conservation of Nature’s (IUCN) Red List with a decreasing population trend. This species is vulnerable to heavy metals during all life-stages: from eggs to adults. In this study, Aspergillus alliaceus isolated from sand samples in this nesting site was identified by internal transcribed spacer sequencing (ITS1-ITS4). Mycelia growth of the fungal strain was subjected to a range of (0–7000) lead concentrations (ppm) incorporated into sabouraud dextrose agar (SDA) in triplicates. Fungal radial growths were recorded every days over a 10-days’ incubation period. The development of the strain is unaffected by lead up to 2000 ppm concentrations, and can also tolerate up to 7000 ppm. In this study; A. alliaceus has a high tolerance to lead. A. alliaceus was firstly evaluated to determine heavy metal tolerance, and has a good potential for bioleaching.

The effect of biologically activated cyclophosphamide on amniotic fluid derived stem cells – in vitro study

Paulina Modrakowska, Łukasz Kaźmierski, Klaudia Bonowicz, Karolina Balik, and Anna Bajek

Department of Tissue Engineering, The Ludwik Rydygier Collegium Medicum, Nicolaus Copernicus University, Toruń, Poland email address: [email protected]

Cyclophosphamide (CP) is a pro-drug metabolized, in living organism, by P450 to its active forms. Due to its strong alkylating properties, it found application in breast cancer therapy. Studies employing amniotic fluid derived stem cells (AFSCs) in vitro might deliver crucial information necessary to assess CP as a potential drug for pregnancy associated breast cancer and ensure fetal safety. CP was activated using rat S9 fraction, NADPH regenerating system and cofactors. AFSCs were treated with CP in 500, 100 and 10 µM concentrations for 24 and 48 h. Viability of treated cells was measured using MTT assay and ScienCellTM Live/Dead Cell Staining Kit. To measure proliferation of treated cells BrdU assay and xCELLigence RTCA system was used. AFSCs apoptosis was estimated using CellEventTM Caspase-3/7 fluorescent assay. 500 µM CP was determined to have the biggest impact on viability, proliferation and apoptosis of AFSCs. The results indicate that AFSCs are hardly affected by 100 and 10 µM CP concentrations and retain the ability to proliferate. Effect of CP on AFSCs needs to be further investigated in terms of possible concentrations, individual variation and effect on 3D cellular model, but these results deliver a promising tool for further research.

Effect of bacterial hemoglobin expression on bioethanol production from malt extract by repeated batches

Gamze Şeker and Meltem Yeşi̇Lçi̇Men Akbaş

Department of Molecular Biology and Genetics, Gebze Technical University, Kocaeli, Turkey email address: [email protected]

Energy sources alternative to fossil fuels must be eco-friendly, renewable, sustainable even provide waste disposal in production process as carbon source. Bioethanol is a bio-molecule produced from carbon-rich materials by ethanologenic microorganisms like yeasts or bacteria. Especially using biotechnologically modified microorganisms and some methods like immobilization in bioethanol production are become widespread in industry as well.

In present study 4.5%maltose and 1% glucose containing maltose (MM) or malt extract (MEM) growth mediums were used as carbon sources to produce ethanol. To investigate effects of bacterial hemoglobin (VHb) expression; 20 day stored of immobilized E. coli strains (FBR5 (VHb (-)) and TS4 (VHb (+)) were used to produce ethanol with malt extract from barley. Immobilized E. coli cells were incubated for seven successive batches with 72 h periods to determine reusability of alginate beads for industrial production. As a result, MEM medium is more effective than MM medium in ethanol production by FBR5, and TS4 strains during 7 batches (2.12 and 2.46% respectively). Moreover immobilized TS4 (vgb+) is more efficient (11%) than immobilized FBR5 cells in ethanol production from malt extract containing medium. Malt extract can be used for effective ethanol production with repeated use of immobilized E. coli expressing VHb.

Overexpression of Crassostrea gigas proline-rich peptide in Pichia pastoris

Mine Erdem Büyükkiraz and Zülal Kesmen^b

^aGastronomy and Culinary Arts Department, School of Applied Sciences, Cappadocia University, Nevşehir, Turkey

^bFaculty of Engineering, Food Engineering Department, Erciyes University, Kayseri, Turkey email address: [email protected]

Antimicrobial peptides have become powerful alternative agents to meet the need for novel anti-infectives. In this study, we performed the overexpression of an oyster-derived antimicrobial peptide, Crassostrea gigas proline-rich peptide (Cg-Prp), in P. pastoris (GS115) by using the recombinant technique. Hence, the nucleotide sequence encoding the Cg-Prp was obtained by the recursive PCR and then ligated into a pPICZaA vector. Multi-copy transformants of P. pastoris were produced by post-transformational vector amplification (PTVA). The multi-copy transformants were selected from yeast extract-peptone-dextrose agar containing a high concentration of antibiotic (4 mg/mL of zeocin). The production of recombinant Cg-Prp (rCg-Prp) by P. pastoris was performed with methanol induction in a buffered methanol-complex medium for 96 h. The concentrations of rCg-Prp, which purified from the culture medium, were determined using a His-Tag ELISA kit. Multi-copy integration increased the peptide accumulation approximately threefold when compared to single-copy integration. The recombinant peptide successfully inhibited the growth of Staphylococcus aureus, Methicillin-resistant S. aureus, Listeria monocytogenes and Bacillus cereus, with the minimum inhibition concentrations in a range from 18.75 to 32.50 pg/mL of rCg-Prp. This study demonstrated that the transformation of Cg-Prp genes into the P. pastoris genome with PTVA approach may an efficient strategy to increase production of recombinant Cg-Prp peptide.

Homemade soaps produced from fried oils

Dani Dordevic^ab, Bojan Antonic^a, Simona Jancikova^a, Karolina Tesikova^a, and Bohuslava Tremlova^a

^aDepartment of Plant Origin Foodstuffs Hygiene and Technology, University of Veterinary and Pharmaceutical Sciences Brno, Brno, Czechia

^bDepartment of Technology and Organization of Public Catering, South Ural State University, Chelyabinsk, Russia, email address: [email protected]

Food waste, including edible oils, from the sources such as industry, restaurants and homes represents significant portion of the whole generated wastes. The literature data are indicating that soaps biodegrade more easily and rapidly in comparison with the degradation of used oils. The aim of the study was to evaluate the production of soaps out of used/fried oils. Soaps were produced out of sunflower, rapeseed and palm oils. Oils were fried (french fries were fried) until the achievement of 24% total polar materials (TPM). Following analysis were performed on soaps: malondialdehyde content, textural parameters and foam evaluation. Malondialdehyde levels in soaps were very low in samples produced from highly fried oils (over 24% TPM). Almost all homemade soaps produced from palm fried oil had the lowest level of malondialdehyde (under 1 µg/g). The textural parameters indicated that soaps produces from fried rapeseed and palm oils had lower hardness in comparison with soaps from fresh oils. The highest disadvantage of produced soaps from fried oil is lower foam formation. Though, on the other side this disadvantage can be easily solved with the addition of foaming agent. The study confirmed reasonable possibility to produce eco-friendly homemade soaps. Supported by: 228/2020/FVHE

Isolation of rhizospheric actinomycetes and determination of their plant growth promoting activities

Hüseyin Evlat, Sultan Kübra Toker, and Ali Koçyiğit

Basic and Industrial Microbiology Section, Biology Department, Ege University, Izmir, Turkey email address: [email protected]; [email protected]

The area that directly surrounds the root system of plants is defined as rhizosphere. The bacterial group colonized in this region is called rhizobacteria, and actinomycetes are a large part of this group. Microbial activity in rhizosphere affects the presence of nutrients available for plants and changes the quality and amount of root secretions. In this study, actinomycetes were isolated from the rhizospheres of different plants in terrestrial and aquatic habitats. The strains were screened for the production of plant growth promoting (agroactive) compounds. Forty-two rhizospheric actinomycetes were isolated and screened for indole acetic acid (IAA), siderophore production (SP), phosphate solubilizing activity (PSA) and hydrogen cyanide (HCN) production. The highest IAA production was obtained from the isolate Nocardiopsis sp. CYP39 (51.04 µg/ml). Of 42 isolates, 32 were determined as siderophore producers on Chrome azurol S containing agar. The best HCN producer was determined as isolate Nocardiopsis sp. CYP15 with glycine containing ISP-2 medium. The isolates Streptomyces moderatus CYP35 and Streptomyces chartreusis CYP19 performed the highest halo-zone diameters in the screening of phosphate solubility test. The presence of antagonistic effects of agroactive compounds of actinomycetes isolated in this study is important to eliminate dependence on harmful chemicals in agriculture.

Anionic monolith column for separation of ε-Poly-L-Lysine

Yaprak Petek Koraltan and Ali Özhan Aytekin

Department of Genetics and Bioengineering, Yeditepe University, Istanbul, Turkey email address: [email protected]

Monolith columns are highly promising material to use as perfusion bioreactor or separation and purification columns for Biotechnology Industry. The ε-Poly-L-Lysine (εPL) is highly cationic polymer that is composed of L-Lysine monomers with 25–35 monomers. In this study, cryogel polymerization of acrylamide (AA) monomers was performed under different initiator, monomer ratio, and temperature. After that cryogel monolith columns were activated by carboxymethyl groups from sodium chloroacetate. Pore size, chemical structure, swollen, ionic charge, and static binding affinity to εPL of columns were characterized by Scanning Electron Microscopy (SEM), Fourier-Transform Infrared Spectroscopy (FTIR), water retention assay, and titration methods, respectively. Pore sizes of AA were in the range of 20–100 µm that was very broad and highly effected by initiator and temperature. FTIR spectra showed that stretching in – OH and NH2 groups were higher than native monoliths. Swollen ratio of monoliths was around 250% that was very high and it sustain no leakage in column header. Ionic charge of activated columns reached to 0.285 meq/ml. Static binding affinity of activated column was 42 mg εPL/mL of column. The activated column was highly promising material for using separation and purification of cationic materials.

Obtaining strains of hybrid cells producing monoclonal antibodies to PD-1 receptor for human T-lymphocytes

Zhansaya Adish^ab, Kanat Tursunov^a, Kassym Mukanov^a, Yerlan Ramankulov^a, and Kanatbek Mukantayev^a

^aNational Center for Biotechnology, Nur-Sultan, Kazakhstan

^bL.N. Gumilyov Eurasian National University, Nur-Sultan, Kazakhstan email address: [email protected]

Immunological therapy of many oncological pathologies includes the use of therapeutic vaccines, poly- and monoclonal antibodies, cytokines, immune response inhibitors and adoptive administration of T cells.

Of particular interest are monoclonal antibodies against the programmed cell death protein 1 (PD-1) receptor. The PD-1 receptor of T-cells is a regulator that can activate or repress immunological responses depending on the underlying context. As a result of the research, a strain of Escherichia coli producing recombinant extracellular domain of PD-1 receptor with a molecular weight of 21 kDa was obtained. Comparison of the results of MS-MS spectrometry with the database of NCBI and SwissProt revealed the 38 most probable proteins (Score 4950) corresponding to the PD-1 receptor. The obtained strains of hybrid cells 5B6C9H, 7F5D11G, 8E4D7E, 10E3F5D producing monoclonal antibodies to PD-1. The specific interaction of monoclonal antibodies was confirmed by Western blot and fluorescence microscopy with T-lymphocytes. The binding constant of monoclonal antibodies ranged from 7 × 10⁻⁸ M⁻¹ to 8 × 10⁻⁸ M⁻¹. This research was performed within the framework of the budget program 217 of the Ministry of Education and Science of the Republic of Kazakhstan for the period 2018–2020. Project No. AP05130036.

Specific cell lines from epidermolysis bullosa patients: the disease modelling and genome editing perspectives

N. G. Gurskaya^a, A. K. Beilin^a, N. E. Evtushenko^a, E. T. Ambarchian^b, N. N. Murashkin^b, K. V. Savostyanov^b, and E. A. Vorotelyak^c

^aCenter for Precision Genome Editing Technologies for Biomedicine, Pirogov Russian National Research Medical University, Moscow, Russia

^bNational Medical Research Center for Children’s Health, Moscow, Russia,

^cKolzov Institute of Developmental Biology of Russian Academy of Sciences, Moscow, Russia email address: [email protected]

The rare group of genodermatoses named inherited epidermolysis bullosa attracts the attention of both geneticists, cell biologists and clinicians, as well. It covers more than 20 genes with a wide spectrum of mutations with different types of inheritance and the severity of symptoms. We described the panels of cell lines from primary cell cultures of patients with recessive form of dystrophic EB-RDEB and of healthy donors as a control. The mutations in the COL7A1 gene were found to be registered in the DEB register for most of the patients. We confirmed splicing impairments of COL7A1 gene by sequencing for one of the patients. The strategy of immortalization by exogenous hTERT cDNA was applied and immortalized cell lines were obtained. The genome editing with CRISPR/Cas9 system was applied to test the opportunity to edit the mutation in COL7A1 gene specific for one of the patient cell line. Several specific sgRNAs were analyzed concerning the activity and specificity on the COL7A1template with c.425G < A mutation. The profiles of the Indels and off-target sites in the case of each sgRNAs were analyzed, that allow to choose the best strategy of gene therapy for correction of RDEB mutation.

Acknowledgements: This work was reformed according to RFBR, project no. 19-29-04044, the sequencing was performed by grant 075-15-2019-1789 from Ministry of Science and Higher Education of the Russian Federation.

Engineering the interior and exterior of Pseudomonas putida for biotechnological processing of lignocellulosic substrates

Pavel Dvorak^a and Víctor de Lorenzo^b

^aDepartment of Experimental Biology (Microbiology Section), Faculty of Science, Masaryk University, Brno, Czech Republic

^bSystems and Synthetic Biology Program, Centro Nacional de Biotecnología (CNB-CSIC), Madrid, Spain email address: [email protected]

Pseudomonas putida KT2440, the best-characterized and safe pseudomonad, has recently drawn considerable attention as a promising bacterial platform for lignocellulose biotechnology. The strain has been employed for the production of value-added chemicals from glucose and it is becoming a host of choice for the valorization of lignin-derived aromatics. However, the lack of certain metabolic traits and functions hinders the application of P. putida for utilization of a wider spectrum of (hemi)cellulosic carbohydrates. In this study, we aimed at empowering P. putida with novel catabolic pathways and a system for efficient display of cellulases on bacterial surface. Metabolic engineering and synthetic biology approaches were adopted for this purpose and applied on P. putida KT2440-derived strains with reduced genomes and altered physiological properties. Isomerase pathway from Escherichia coli and cytoplasmic β-glucosidase from Thermobifida fusca implanted in P. putida mutant enabled rapid growth of the recombinant on D-xylose (μ = 0.17 h⁻¹) and D-cellobiose (μ = 0.35 h⁻¹) and co-utilization of these biotechnologically relevant sugars. The P. putida surface was also efficiently decorated with β-glucosidase molecules using designer protein scaffolds mimicking the function of natural cellulosomes. The project provides a showcase of expanding the catalytic scope of environmental bacterium toward biotechnological applications.

Protective bioproducts for fresh soft fruits

Alina Butu, Steliana Rodino, Adelina Ghincea, Melania Bica Popi, and Marian Butu

National Institute of Research and Development for Biological Sciences, Bucharest, Romania email address: [email protected]

The food industry challenges are to provide nutritional food while implementing sustainable production processes. The importance of fruits in nutrition, health and economy is well established. They are the best carriers of vitamins, essential minerals, dietary fiber, phenolic antioxidants, glucosinolates and other bioactive substances. Soft fruits therefore play an important role in the daily diet, and it is advisable to be consumed fresh. However, soft fruits are extremely perishable goods. Currently, up to 23 percent of the most perishable are lost during their agri-food chain journey as they deteriorate. The objective of our research was to study the silver nanoparticles by the synthesis mediated with aqueous plant extracts and to evaluate their preservation effect on soft fruits. The silver nanoparticles were formed by reducing silver nitrate solution with the plant extracts. The protective effect was tested by immersion and spraying. Various concentrations of silver nanoparticles – plant extracts solutions were investigated for their effect on prolongation of fruits shelf life. The silver nanoparticles produced by green synthesis showed the good results on prolongation of tested fruits shelf life.

Advanced biotechnology for production of biocomposites based on macromycetes mycelium

Marian Butu, Steliana Rodino, Gina Fidler, Alexandra Dobranici, Bogdan Miu, and Alina Butu

National Institute of Research and Development for Biological Sciences, Bucharest, Romania email address: [email protected]

Due to the increasing demand for ecological materials and productive processes nowadays, technology based on production of biocomposite and bio-based materials was developed. According to recent available studies it was demonstrated that mycelium bio-composites coupled with other materials derived from biological processes, such as plant extracts, represent an alternative for production of different materials used in construction industry, textile industry or food industry. It was demonstrated that mushrooms are economically important biotechnological products, representing a good source of protein, vitamins, minerals, and biologically active substances. Pleurotus ostreatus, known as the oyster mushroom, is a white-rot edible fungus, which is strongly saprophytic and highly adaptable. The research carried out was aimed to capitalize of some wastes from agriculture and food in the cultivation of macromycetes. The results obtained from these researches showed that the tested waste can be used for the development of macromycetes mycelium from Pleurotus sp and Ganoderma sp. The novelty and relevance of the preliminary results are given by the possibility of using macromycetes mycelium to design new biomaterials which could be a promising alternative for production of manufacturing products.

Characterization of monoclonal antibodies against receptor 161 associated with protein G

Kanat Tursunov, Zhansaya Adish, Kanatbek Mukantayev, Yerlan Ramankulov, and Kassym Mukanov

National Center for Biotechnology, Nur-Sultan, Kazakhstan email address: [email protected]

Triple-negative breast cancer (TNBC) is the most common type of cancer and the leading cause of death among women. Monoclonal antibodies are recognized as one of the most promising agents for the treatment of human malignant tumors. The G-protein coupled receptor 161 (GPR161) has been identified as a prognostic biomarker for TNBC. The GPR161 receptor has been found to be an important regulator of cell proliferation and migration in breast cancer. As a result of studies, a recombinant GPR161 receptor was obtained, including amino acids 64–344, with a molecular weight of 49 kDa. MS/MS spectrometry showed a high correspondence of the amino acid sequence of the protein presented in international databases. The obtained hybrid cell strains H3D8, F10G7, C1G8, stably produced monoclonal antibodies against recombinant GPR161 protein. The productivity of the obtained clones in the cell culture was 25 μg/ml, in vivo – 8 mg/ml. Monoclonal antibodies specifically reacted with the recombinant GPR161 protein and had a binding constant of 8.8 × 10⁻⁸ M⁻¹. This research is carried out within the framework of the budget program 217 of the Ministry of Education and Science of the Republic of Kazakhstan for the period 2018–2020. Project No. AP05130053.

Preparation of salicylic acid imprinted cryogel membranes for the releasing of salicylic acid

Sevgi Aslıyüce^a and Neslihan Idil^b

^aDepartment of Chemistry, Hacettepe University, Ankara, Turkey

^bDepartment of Biology, Hacettepe University, Ankara, Turkey email address: [email protected]

Cryogels are one of the most widely used materials for the preparation of controlled releasing systems. Cryogels are synthesized in the presence of monomeric or polymeric precursors at temperatures below the freezing point of the solvent. Nowadays, cryogels have been used as chromatographic materials, carriers for immobilization of molecules and cells, matrices for cell separation and matrices for drug release in many fields of biotechnology. Molecularly imprinted polymers mimicking the recognition processes found in nature, and the research area on molecular imprinting to create materials with the ability to recognize similar to natural systems have become an important and attractive field in recent years. This technique has been applied for several purposes such as purification, chemical sensors, catalysis, biological antibodies, receptor systems and drug release. In the present study, salicylic acid imprinted cryogel membranes were prepared. In addition, their controlled release profiles, kinetic analysis and antibacterial properties were examined. ‘Accordingly, it has been indicated that salicylic acid imprinted cryogel membranes have antibacterial properties against Enterococcus faecalis, Staphylococcus aureus and Escherichia coli, by the way, they are active against both Gram negative and positive bacteria’. In conclusion, cryogels are promising tools as potential carriers for sustained release and antibacterial materials.

Determination of optimum reduced graphene oxide synthesis method for conductive sensing material production

Didem Aycan, Fatma Karaca, and Neslihan Alemdar

Department of Chemical Engineering, Marmara University, Istanbul, Turkey email address: [email protected]

In recent years, graphene-family materials have been commonly used in the production of conductive materials for biosensor applications. Since they offer high electrical conductivity, excellent mechanical flexibility and, biocompatibility, they are accepted as a promising material for biosensors. Reduced graphene oxide (RGO) is superior to other graphene-derivative materials due to its restored sp2-hybridized carbon system and improved conductivity. In the present study, we aim to determine the optimum RGO synthesis method for the production of the conductive material used in biosensor applications. Firstly, graphene oxide (GO) was synthesized starting from graphite by the modified Hummers method. Following that, obtained GO was reduced through two different methods as chemical and thermal reduction. Graphite, GO and RGO samples obtained via different reduction methods were characterized by FT-IR, Raman, and UV-Vis spectroscopy. The reduction efficiency of each method was evaluated according to Raman results. Finally, the most efficient method to obtain RGO, which is a good candidate for the production of conductive materials used as biosensors, was determined based on all results.

Stability of lycopene during the beer fermentation process

Enriqueta Martinez Rojas^a, Hamza Gadhoumi^b, Eridani Fuentes^c, and Sabrina Mann^d

^aDepartment of Food Technology and Food Chemistry, Lab of Brewing Science, Berlin Technical University, Berlin, Germany

^bFaculty of Mathematical, Physical, and Natural Sciences of Tunis, University Tunis El Manar, Tunis, Tunisia

^cDepartment of Pharmacology CINVESTAV, Mexico City, Mexico

^dDepartment of Agriculture and Food Sciences, Neubrandenburg University of Applied Sciences, Neubrandenburg, Germany email address: [email protected]

The presented work displays the stability of bioactive pigments added during the beer fermentation process. As part of an interdisciplinary work for the feasibility analysis of a functional beer, which is enriched with the pigment lycopene. Lycopene is very well known as bioactive compound regard to its high antioxidative activity and nutritional properties that can have a positive effect on the human body. However, during the brewing process, the lycopene could be metabolized by the yeast, therefore, the aim of the work was to investigate the stability of lycopene during the beer fermentation process. For this purpose, beers with different malt compositions were produced and analysed according to the MEBAK® analytical protocols in order to elucidate the relative effect of ethanol, and pH-value on pigment stability on beer. The hyprosochromic shift (L*a*b-colour-value) and isomerization of the pigment degradation were more pronounced at the end for fermentation time with high ethanol concentration (up to 4% alc.). Statistical analysis indicated that pigment degradation was affected more by ethanol concentrations than by pH-value (4.5 and 3.5). Taking all together, we conclude that at the end of beer fermentation with lycopene, a product with complementary bioactive ingredients is formed.

Screening of microorganisms for chitinase production by solid-state fermentation using insect meal

Ana Margarida De Matos Gonçalves^a, Patrícia Marlene Alves Ferreira^b, Maria Helena Tabuaço Rego Martins Peres^c, Isabel Maria Pires Belo^a, and José Manuel Salgado Seara^a

^aCEB-Centre of Biological Engineering, University of Minho, Braga, Portugal

^bDepartamento de Biologia, Faculdade de Ciências, Universidade do Porto, Porto, Portugal

^cCIMAR/CIIMAR-Centro Interdisciplinar Marinha e Ambiental, Universidade do Porto, Porto, Portugal email address: [email protected]

The use of insect meal as a new and sustainable feed ingredient has received intensive research effort, but its dietary inclusion levels are limited to its chitin content. Chitin is a complex cross-linked molecular structure, which cannot be digested by animals. To digest chitin, chemical methods may be used, presenting, however, low yield, high cost and environmental impact. Thus, the development of natural chitinases has important biological relevance, allowing it incorporation in animal feed. In this work, a screening of fungi producers of chitinase was performed in agar plates with insect meal (Hermetia illucens) and Aspergillus niger 01UAs183 had the best performance. In insect meal fermentation during 7 days, A. niger produced 50 units of chitinase per gram of dry meal. The mixture of insect meal with brewerýs spent grain (1:1, w/w) fermented during 14 days lead to a 10-fold increase of chitinase activity. Mixtures of insect meal with other agro-industrial wastes are promising substrates for chitinase production, opening new strategies to improve the production of chitinase and the nutritional properties of new alternative protein sources. Acknowledgement: This study was supported by InovFeed (ref. MAR-02.01.01-FEAMP-0111; Mar 2020), SPO3 (ref. POCI-01-0145-FEDER-030377; FCT), BiotecNorte (NORTE-01-0145-FEDER-000004 and strategic funding (UIBD/04469/2020).

Isolation and characterization of a new bacteriophage against Rhodococcus erythropolis

Rute Ferreira, Sílvio Santos, Inês Pacheco, Vânia Ferreira, Luís Melo, and Ana Nicolau

CEB – Centre of Biological Engineering, University of Minho, Campus de Gualtar, Braga, Portugal email address: [email protected]

Activated-sludge is the most common biotechnology to treat wastewaters. In the aerating tank of these systems, a true ecosystem develops, being bacteria the key players in the depuration process. Frequently, filamentous bacteria grow exaggeratedly, being the source of two well phenomena that cause significant throwbacks in the process: bulking and foaming. (Rhodococcus erythropolis) is a gram positive filamentous bacterium previously identified as responsible for the formation of foams in activated-sludge wastewater treatment plants (WWTP). Bacteriophages (phages) are viruses that specifically infect bacteria, using the bacterial machinery to reproduce themselves in a very efficient way. They are the most abundant entities in the biosphere, being found in every environment where their bacterial hosts are present. In this work, one lytic phage for (R. erythropolis) was isolated, using sewage water from an urban WWTP. The phage was tested against other species found in WWTP to conclude about its lytic spectra. Stability of the phage solution at different temperatures and pH was determined. Morphologic analysis by Transmission Electron Microscopy (TEM) and Genomic Sequencing were also performed.

Results clearly indicate the potential of this bacteriophage for future studies in wastewater treatment system management.

Investigation of growth properties of Paenibacillus strains in the presence of 2-HBP

Taner Sar, Murat Ozturk, and Meltem Yesilcimen Akbas

Department of Molecular Biology and Genetics, Gebze Technical University, Kocaeli, Turkey email address: [email protected]

Crude oil and fossil fuels contain sulphur compounds. Consumption of these compounds could cause acid rain and environmental pollution. Dibenzothiophene (DBT) is considered as model compound for investigation of fossil fuel desulphurization. This compound is converted to 2-HBP following bio-desulphurization (BDS) by 4S pathway. This work aimed to investigate bacterial growth properties in the presence of 2-HBP which is a final product of DBT desulphurization. For this, two Paenibacillus strains (32 O-W and 32 O-Y) were incubated in 2-HBP (to give the final concentrations at 0.05, 0.1, and 0.2 mM) containing growth media. Although the OD600 value of the 32 O-W strain was low at all 2-HBP concentrations, the 32 O-Y strain could survive, and its OD levels were 3–4 fold higher than 32 O-W strain. As a result, it can be interpreted that the final products of DBT metabolism did not too toxic, and thus bio-desulfurization can be successfully completed by Paenibacillus strains. Acknowledgments: This work was supported by the Scientific and Technological Research Council of Turkey (TUBITAK, 118Y416).

An evaluation of vitamin D and bone turnover markers levels in postmenopausal women in Albania

Esmeralda Agim Hoxhaj^a and Anila Mitre^b

^aDepartment of Mikrobiology, Intermedica Center, Tirana, Albania

^bDepartment of Biology, University of Tirana, Tirana, Albania email address: [email protected]

Vitamin D is an essential lipophilic prohormone that is synthesized in the skin in response to sunlight, although diet may be a source of much lower amounts of Vitamin D. Receptors of the active form of Vitamin D (VDR), have been identified in the cells of the intestinal epithelium, renal tubules, bone and other tissues and organs, which indicates a broad spectrum of 25(OH)D3. The consequences of vitamin D deficiency are secondary hyperparathyroidism and bone loss, leading to osteoporosis and fractures, mineralization defects, which may lead to osteomalacia in the long term, and muscle weakness, causing falls and fractures. The vitamin D status within different European countries shows a high variation. In this two-years study (2017–2019), we described the epidemiology of vitamin D status across women population in Albania and its potential associations with bone biomarkers (OC, CTX, PTH, ALP). From the results we notice a reduced 25(OH)D concentration in Albanian women (23.25 ± 10.6 ng/mL), and showed that a deficient 25(OH)D concentration is associated with significantly increased markers of bone resorption. Our study showed also a negative Pearson correlation between serum 25(OH)D and osteocalcin (OC) (p < 0.05).

Small molecule inhibitor library screening for MCL-1

Oznur Atay^a, Sehime G. Temel^b, Sehime G. Temel^c, Sehime G. Temel^d, Huveyda Basaga^a, and Ozgur Kutuk^e

^aFaculty of Engineering and Natural Sciences, Molecular Biology, Genetics and Bioengineering Program, Sabanci University, Tuzla, Istanbul, Turkey

^bFaculty of Medicine, Department of Histology & Embryology, Bursa Uludag University, Bursa, Turkey

^cFaculty of Medicine, Department of Medical Genetics, Bursa Uludag University, Bursa, Turkey

^dInstitute of Health, Department of Translational Medicine, Bursa Uludag University, Bursa, Turkey

^eAdana Dr. Turgut Noyan Medical and Research Center, School of Medicine, Baskent University, Yuregir, Adana, Turkey email address: [email protected]

Mitochondrial priming level determines the fate of cell and it facilitates the tendency of undergoing apoptosis. MCL-1 protein is a member of anti-apoptotic BCL-2 family proteins, and it plays a crucial role on chemoresistance mechanism, either in targeted therapy or chemotherapy. Our project aimed to investigate the mitochondrial cell death priming status via BH3 profiling method to find potent inhibitors for anti-apoptotic MCL-1 protein in non-small lung cancer cell lines. The small-molecule inhibitor library (1280 compounds) was screened by the help of an automated pipetting system. To report the effects of these molecules on the mitochondrial charge, non-linear regression analysis was conducted, and the depolarization percentage of each inhibitor was revealed using Graph-pad Prism 6.2 software. We categorized the responses of the molecules into five profiles, according to the revealed mitochondrial outer membrane potential loss. These classes are depolarization (P1), hyperpolarization (P2), depolarization at low dose (P3), depolarization at high dose (P4) and non-conclusive (P5). 72% of total 1280 molecules caused hyperpolarization on the mitochondria, while 24% of the molecules depolarized the mitochondria and caused mitochondrial outer membrane potential loss.