How are Students Really selected for a PhD? The Impact of the Thesis and Solving the Country's Problems 2 2 No Abstract 1 1 Shahin Akhondzadeh Editorial 60596.pdf Akhondzadeh S. The Need for Serious Support for Basic Medical Science in Iran. Avicenna J Med Biotechnol 2024 Jul-Sep;16(3):136. ##Dehghani M, Akhondzadeh S, Mesgarpour B, Ferdousi R. A Tool to Reduce the Problems of Iranian Health Researchers. Iran J Public Health 2020 Dec;49(12):2441-2. ##

Therapeutic Effects of Nanochelating-Based Copper Nanoparticles on Burn Wound Healing in Mouse Model 2 2 Background: The aim of the present study was to investigate the potential of Nanochelating-based copper to accelerate the wound healing process and prevent infection in burn wounds. Methods: Six to eight-week- old female BALB/c mice were burned with a 1 cm2 heated copper plate on the left flank and then divided into four treatment groups, treated with C8 (nanochelating-based CuNPs), cold cream (supplementary materials) as a control drug, Silver Sulfadiazine and no treatment, respectively. Skin tissue samples were taken from the mice on days 0, 3, 8, 15 and 24. One piece was fixed in 10% neutral buffered formalin for pathological examination and the others were stored at -80°C until used for pro-inflammatory and growth factor gene expression. Results: The healing process in the group treated with 10 mg/ml C8 was significantly faster, and the survival rate of the mice in this group was significantly higher than in the other groups. The pro-inflammatory genes were expressed and down-regulated earlier in the C8 treated mice. Histopathology confirmed the higher cure rate in the group treated with 10 mg/ml C8 compared to other control groups. Conclusion: C8 has beneficial effects on the healing of burn wounds and the effective dose of this compound should be further investigated. The present study demonstrates the anti-inflammatory properties of nano-chelate-based copper particles' on mouse skin burns. This research opens up new possibilities in dermatology and burn therapy and highlights the potential of copper-based formulations in the treatment of burn injuries. 2 13 Hossein Rezvan Seyed Masoud Zolhavarieh Department of Clinical Sciences, Faculty of Veterinary Science, Bu-Ali Sina University Department of Clinical Sciences, Faculty of Veterinary Science, Bu-Ali Sina University Iran Alireza Nourian Department of Pathobiology, Faculty of Veterinary Science, Bu-Ali Sina University Department of Pathobiology, Faculty of Veterinary Science, Bu-Ali Sina University Iran Elham Bayat Department of Pathobiology, Faculty of Veterinary Science, Bu-Ali Sina University Department of Pathobiology, Faculty of Veterinary Science, Bu-Ali Sina University Iran Somayeh Kalanaky Department of Research and Development, Sodour Ahrar Shargh Company Iran Saideh Fakharzadeh Department of Research and Development, Sodour Ahrar Shargh Company Iran Pegah Karimi Department of Research and Development, Sodour Ahrar Shargh Company Department of Research and Development, Sodour Ahrar Shargh Company Iran Maryam Hafizi Department of Research and Development, Sodour Ahrar Shargh Company Department of Research and Development, Sodour Ahrar Shargh Company Iran Sahar Hamoon Navard Department of Pathobiology, Faculty of Veterinary Science, Bu-Ali Sina University Department of Pathobiology, Faculty of Veterinary Science, Bu-Ali Sina University Iran Mohammad Hassan Nazaran Anti-inflammatory agentsBurnsCopperDermatologyWound healing 60597.pdf Hettiaratchy S, Dziewulski P. Pathophysiology and types of burns. BMJ 2004;328(7453):1427-9.##Alarifi S, Ali D, Verma A, Alakhtani S, Ali BA. Cytotoxicity and genotoxicity of copper oxide nanoparticles in human skin keratinocytes cells. Int J Toxicol 2013;32(4):296-307.##Periayah MH, Halim AS, Mat Saad AZ. Mechanism Action of Platelets and Crucial Blood Coagulation Pathways in Hemostasis. Int J Hematol Oncol Stem Cell Res 2017;11(4):319-27.##Reinke JM, Sorg H. Wound repair and regeneration. Eur Surg Res 2012;49(1):35-43.##Sorg H, Tilkorn DJ, Hager S, Hauser J, Mirastschijski U. Skin Wound Healing: An Update on the Current Knowledge and Concepts. Eur Surg Res 2017;58(1-2):81-94.##Werner S, Grose R. Regulation of wound healing by growth factors and cytokines. Physiol Rev 2003;83(3):835-70.##Cañedo-Dorantes L, Cañedo-Ayala M. Skin Acute Wound Healing: A Comprehensive Review. Int J Inflam 2019;2019:3706315.##Kumar A, Pandit V, Nagaich U. Preparation and evaluation of copper nanoparticles loaded hydrogel for burns. Int J App Pharm 2021 Mar 7;13(2):108-89.##Cangul IT, Gul NY, Topal A, Yilmaz R. Evaluation of the effects of topical tripeptide-copper complex and zinc oxide on open-wound healing in rabbits. Vet Dermatol 2006;17(6):417-23.##Borkow G, Gabbay J, Dardik R, Eidelman AI, Lavie Y, Grunfeld Y, Ikher S, Huszar M, Zatcoff RC, Marikovsky M. Molecular mechanisms of enhanced wound healing by copper oxide‐impregnated dressings. Wound repair and regeneration 2010 Mar;18(2):266-75.##Salvo J, Sandoval C. Role of copper nanoparticles in wound healing for chronic wounds: literature review. Burns Trauma 2022;10:tkab047.##Sandoval C, Ríos G, Sepúlveda N, Salvo J, Souza-Mello V, Farías J. Effectiveness of Copper Nanoparticles in Wound Healing Process Using In Vivo and In Vitro Studies: A Systematic Review. Pharmaceutics. 2022 Aug 31;14(9):1838.##Ermini ML, Summa M, Zamborlin A, Frusca V, Mapanao AK, Mugnaioli E, et al. Copper nano-architecture topical cream for the accelerated recovery of burnt skin. Nanoscale Adv 2023;5(4):1212-9.##Kranz I, Gonzalez JB, Dörfel I, Gemeinert M, Griepentrog M, Klaffke D, et al. Biological response to micron- and nanometer-sized particles known as potential wear products from artificial hip joints: Part II: Reaction of murine macrophages to corundum particles of different size distributions. J Biomed Mater Res A 2009;89(2):390-401.##Ghaffari S, Alihosseini F, Rezayat Sorkhabadi SM, Arbabi Bidgoli S, Mousavi SE, Haghighat S, et al. Nanotechnology in Wound Healing; Semisolid Dosage Forms Containing Curcumin-Ampicillin Solid Lipid Nanoparticles, in-Vitro, Ex-Vivo and in-Vivo Characteristics. Adv Pharm Bull 2018;8(3):395-400.##Mofazzal Jahromi MA, Sahandi Zangabad P, Moosavi Basri SM, Sahandi Zangabad K, Ghamarypour A, Aref AR, et al. Nanomedicine and advanced technologies for burns: Preventing infection and facilitating wound healing. Adv Drug Deliv Rev 2018;123:33-64.##Alizadeh S, Seyedalipour B, Shafieyan S, Kheime A, Mohammadi P, Aghdami N. Copper nanoparticles promote rapid wound healing in acute full thickness defect via acceleration of skin cell migration, proliferation, and neovascularization. Biochem Biophys Res Commun 2019;517(4):684-90.##Tiwari M, Narayanan K, Thakar MB, Jagani HV, Venkata Rao J. Biosynthesis and wound healing activity of copper nanoparticles. IET Nanobiotechnol 2014;8(4):230-7.##Karimi-Sales R, Ashiri M, Hafizi M, Kalanaky S, Maghsoudi AH, Fakharzadeh S, et al. Neuroprotective Effect of New Nanochelating-Based Nano Complex, ALZc3, Against Aβ ((1-42))-Induced Toxicity in Rat: a Comparison with Memantine. Pharm Res 2020;37(3):48.##Hafizi M, Kalanaky S, Moaiery H, Khayamzadeh M, Noorian S, Kaveh V, et al. A randomized, double-blind, placebo-controlled investigation of BCc1 nanomedicine effect on survival and quality of life in metastatic and non-metastatic gastric cancer patients. J Nanobiotechnology 2019;17(1):52.##Fakharzadeh S, Kalanaky S, Hafizi M, Nazaran MH, Zardooz H. DIBc, a nanochelating-based nano metal-organic framework, shows anti-diabetic effects in high-fat diet and streptozotocin-induced diabetic rats. Int J Nanomedicine 2019;14:2145-56.##Hafizi M, Hajarizadeh A, Atashi A, Kalanaky S, Fakharzadeh S, Masoumi Z, et al. Nanochelating based nanocomplex, GFc7, improves quality and quantity of human mesenchymal stem cells during in vitro expansion. Stem Cell Res Ther 2015;6:226.##Hoseini‐Alfatemi SM, Karimi A, Armin S, Fakharzadeh S, Fallah F, Kalanaky S. Antibacterial and antibiofilm activity of nanochelating based silver nanoparticles against several nosocomial pathogens. Applied Organometallic Chemistry 2018;32(5):e4327.##de Lacerda Coriolano D, de Souza JB, Bueno EV, Medeiros SMdFRdS, Cavalcanti IDL, Cavalcanti IMF. Antibacterial and antibiofilm potential of silver nanoparticles against antibiotic-sensitive and multidrug-resistant Pseudomonas aeruginosa strains. Braz J Microbiol 2021;52(1):267-78.##Ag S. Effectiveness of nano-copper solution to reduce wound bioburden in chronic wounds: a case series.##Vemuri S, Abraham S, Azamthulla M, Furtado S, Bharath S. Development of in situ gels of nano calcium oxide for healing of burns. Wounds Medicine 2020;28:100177.##Alizadeh S, Seyedalipour B, Shafieyan S, Kheime A, Mohammadi P, Aghdami N. Copper nanoparticles promote rapid wound healing in acute full thickness defect via acceleration of skin cell migration, proliferation, and neovascularization. Biochem Biophys Res Commun 2019;517(4):684-90.##Akhzari S, Rezvan H, Zolhavarieh SM. Expression of pro-inflammatory genes in lesions, spleens and blood neutrophils after burn injuries in mice treated with silver sulfodiazine. Iran J Basic Med Sci 2017;20(7):769-75.##Hernandez A, Patil NK, Bohannon JK. A Murine Model of Full-Thickness Scald Burn Injury with Subsequent Wound and Systemic Bacterial Infection. Methods Mol Biol. 2021;2321:111-120.##Vinaik R, Aijaz A, Jeschke MG. Small animal models of thermal injury. Methods Cell Biol 2022;168:161-89.##Krahwinkel DJ, Boothe HW, Jr. Topical and systemic medications for wounds. Vet Clin North Am Small Anim Pract 2006;36(4):739-57.##McCulloch JM, Kloth LC, Feedar JA. Wound healing: alternatives in management. Wound healing: alternatives in management1995. p. xxvi, 442-xxvi.##Souto EB, Ribeiro AF, Ferreira MI, Teixeira MC, Shimojo AA, Soriano JL, et al. New nanotechnologies for the treatment and repair of skin burns infections. Int J Mol Sci 2020;21(2):393.##Zarrintaj P, Moghaddam AS, Manouchehri S, Atoufi Z, Amiri A, Amirkhani MA, et al. Can regenerative medicine and nanotechnology combine to heal wounds? The search for the ideal wound dressing. Nanomedicine 2017;12(19):2403-22.##Barroso A, Mestre H, Ascenso A, Simões S, Reis C. Nanomaterials in wound healing: From material sciences to wound healing applications. Nano Select 2020;1(5):443-60.##Borkow G, Gabbay J, Zatcoff RC. Could chronic wounds not heal due to too low local copper levels? Med Hypotheses 2008;70(3):610-3.##Kornblatt AP, Nicoletti VG, Travaglia A. The neglected role of copper ions in wound healing. J Inorg Biochem 2016;161:1-8.##Paterson TE, Bari A, Bullock AJ, Turner R, Montalbano G, Fiorilli S, et al. Multifunctional copper-containing mesoporous glass nanoparticles as antibacterial and proangiogenic agents for chronic wounds. Front Bioeng Biotechnol 2020;8:246.##Ahire JJ, Hattingh M, Neveling DP, Dicks LM. Copper-containing anti-biofilm nanofiber scaffolds as a wound dressing material. PloS One 2016;11(3):e0152755.##Nethi SK, Das S, Patra CR, Mukherjee S. Recent advances in inorganic nanomaterials for wound-healing applications. Biomater Sci 2019;7(7):2652-74.##Cucci LM, Satriano C, Marzo T, La Mendola D. Angiogenin and copper crossing in wound healing. Int J Mol Sci 2021;22(19):10704.##Baino F, Hamzehlou S, Kargozar S. Bioactive glasses: where are we and where are we going? J Funct Biomater 2018;9(1):25.##Goonoo N, Bhaw-Luximon A. Mimicking growth factors: Role of small molecule scaffold additives in promoting tissue regeneration and repair. RSC Adv 2019;9(32):18124-46.##Xie M, Fan D, Zhao Z, Li Z, Li G, Chen Y, et al. Nano-curcumin prepared via supercritical: Improved anti-bacterial, anti-oxidant and anti-cancer efficacy. Int J Pharm 2015;496(2):732-40.##Xu Y, Pindolia K, Janakiraman N, Noth C, Chapman R, Gautam S. Curcumin, a compound with anti-inflammatory and anti-oxidant properties, down-regulates chemokine expression in bone marrow stromal cells. Exp Hematol 1997;25(5):413-22.##Basniwal RK, Buttar HS, Jain V, Jain N. Curcumin nanoparticles: preparation, characterization, and antimicrobial study. J Agric Food Chem 2011;59(5):2056-61.##Mofazzal Jahromi MA, Sahandi Zangabad P, Moosavi Basri SM, Sahandi Zangabad K, Ghamarypour A, Aref AR, et al. Nanomedicine and advanced technologies for burns: Preventing infection and facilitating wound healing. Adv Drug Deliv Rev 2018;123:33-64.##Sandoval C, Ríos G, Sepulveda N, Salvo J, Souza-Mello V, Farias J. Effectiveness of copper nanoparticles in wound healing process using in vivo and in vitro studies: a systematic review. Pharmaceutics 2022;14(9):1838.##Zeng Z, Lan T, Wei Y, Wei X. CCL5/CCR5 axis in human diseases and related treatments. Genes Dis 2022;9(1):12-27.##Marques RE, Guabiraba R, Russo RC, Teixeira MM. Targeting CCL5 in inflammation. Expert Opin Ther Targets 2013;17(12):1439-60.##Hannestad J, DellaGioia N, Bloch M. The effect of antidepressant medication treatment on serum levels of inflammatory cytokines: a meta-analysis. Neuropsychopharmacology 2011;36(12):2452-9.##Zhou W, Zi L, Cen Y, You C, Tian M. Copper sulfide nanoparticles-incorporated hyaluronic acid injectable hydrogel with enhanced angiogenesis to promote wound healing. Front Bioeng Biotechnol 2020;8:417.##

Moringa oleifera Docking to Estrogen Receptor α Ameliorates Placental and Brain Damage in Stressed Rats 2 2 Background: Stress during pregnancy significantly impacts offspring early physiological programming. Herbal remedies are frequently used by pregnant women to enhance their wellbeing. Moringa oleifera Leaf Extract (MoLE) is believed to have both anti-stress and antioxidant properties which can act as a Selective Estrogen Receptor Modulator (SERM) that regulate activities of estrogen, and can have different effects on different tissues. Goal of this study is to compile information on molecular docking analysis of phytochemicals found in MoLE targeting Estrogen Receptor-alpha (ER-α) and assess effects of MoLE administration on dam's and fetal brain tissues and placenta, during gestational stress. Methods: Phytochemical study of MoLE was determined using Gas Chromatography-Mass Spectrometry. Molecular docking technique was employed to predict aspects of interaction and binding affinities energy of bioactive phytocompounds in protein site of ER-α using autodock tools. 30 apparently healthy pregnant Albino-Wistar rats were randomly placed into 6 groups of 5 rats per group and exposed to Chronic Unpredictable Stress (CUS) protocol for two weeks, as follows: Group I (water and normal rat chow ad libitum), Group II (CUS protocol only), Group III (5 mg/kg body weight/day of MoLE), Group IV (10 mg/kg body weight/day of MoLE), Group V (CUS protocol +5 mg/kg body weight/day of MoLE), Group VI (CUS protocol +10 mg/kg body weight/ day of MoLE). Results: This study found that 1-Propanol, 3,3'-oxy bis- and 1, 2, 3-Trimethyldiazir-idine are most potent ligands for ER-α among all 41 compounds. Photomicrograph examination of tissues from stressed rats showed mild to severe alterations in histology. Consumption of MoLE during chronic stress showed mild to moderate protective effects. Conclusion: These findings suggest that 1-Propanol, 3,3'-oxy bis- and 1, 2, 3-Trimethyldiaziridine can be further investigated for development of novel therapeutics. 14 23 Odochi O. Chukwu Department of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, David Umahi Federal University of Health SciencesDepartment of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, Alex Ekwueme Federal University, Ndufu-Alike. Abakaliki Department of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, David Umahi Federal University of Health SciencesDepartment of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, Alex Ekwueme Federal University, Ndufu-Alike. Abakaliki NigeriaNigeria Cordilia Iyare Anthony C.U. Ezimah Department of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, Alex Ekwueme Federal University, Ndufu-Alike. Abakaliki Department of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, Alex Ekwueme Federal University, Ndufu-Alike. Abakaliki Nigeria Albert E. Okorocha Department of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, Alex Ekwueme Federal University, Ndufu-Alike. Abakaliki Department of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, Alex Ekwueme Federal University, Ndufu-Alike. Abakaliki Nigeria Nwaeze G. Konyefom Department of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, Alex Ekwueme Federal University, Ndufu-Alike. Abakaliki Department of Physiology, Faculty of Basic Medical Sciences, College of Medical Sciences, Alex Ekwueme Federal University, Ndufu-Alike. Abakaliki Nigeria Nnaemeka T. Asogwa Central Research Laboratory, University Road Central Research Laboratory, University Road Nigeria Nancy P. Igwe Department of Anatomy, Faculty of Basic Medical Sciences, College of Medicine, Alex Ekwueme Federal University, Ndufu-Alike Abakaliki Department of Anatomy, Faculty of Basic Medical Sciences, College of Medicine, Alex Ekwueme Federal University, Ndufu-Alike Abakaliki Nigeria BrainEstrogensMoringaPhytochemicalsReceptors 60598.pdf Howland MA, Sandman CA, Davis EP, Stern HS, Phelan M, Baram TZ, et al. Prenatal maternal mood entropy is associated with child neurodevelopment. Emotion 2021;21(2):489-99.##McEwen BS. Neurobiological and systemic effects of chronic stress. Chronic Stress (Thousand Oaks) 2017;(1):2470547017692328.##Nicoloro-SantaBarbara J, Busso C, Moyer A, Lobel M. Just relax and you'll get pregnant? Meta-analysis examining women's emotional distress and the outcome of assisted reproductive technology. Soc Sci Med 2018;213:54–62.##Frawley J, Adams J, Steel A, Broom A, Gallois C, Sibbritt D. Women’s use and self-prescription of herbal medicine during pregnancy: an examination of 1,835 Pregnant Women. Women’s Health Issues 2015;25(4):396-402.##Tiran D. The use of herbs by pregnant and childbearing women: a risk benefit assessment. Complement Ther Nurs Midwifery 2003;9(4):176-81.##Moses BE. Health Benefits of Moringa Oleifera-The Miracle Tree. African Science Literacy Network. February 14, 2021.##Berg AH, Rice CD, Rahman MS, Dong J, Thomas P. Identification and characterization of membrane androgen receptors in the ZIP9 zinc transporter subfamily: I. Discovery in female atlantic croaker and evidence ZIP9 mediates testosterone-induced apoptosis of ovarian follicle cells. Endocrinology 2014 Nov;155(11):4237-49.##Thomas P, Converse A, Berg HA. ZIP9, a novel membrane androgen receptor and zinc transporter protein. Gen Comp Endocrinol 2018;257:130–6.##Ye R, Pi M, Cox JV, Nishimoto SK, Quarles LD. CRISPR/Cas9 targeting of GPRC6A suppresses prostate cancer tumorigenesis in a human xenograft model. J Exp Clin Cancer Res 2017 Jun 28;36(1):90.##Clemmensen C, Smajilovic S, Wellendorph P, Bräuner-Osborne H. The GPCR, class C, group 6, subtype A (GPRC6A) receptor: from cloning to physiological function. Br J Pharmacol 2014 Mar;171(5):1129-41.##Essa MM, Subash S, Parvathy S, Meera A, Guillemin GJ, Memon MA, Manivasagam T. Brain health benefits of Moringa oleifera. Food and Brain Health 2014 Jan 1;2:113-8.##Ali MA, Yusof YA, Chin NL, Ibrahim MN, Muneer S. Development and Standardization of Moringa oleifera Leaves as a Natural Dietary Supplement. J Diet Suppl 2019;16(1):66-85.##Chukwu OO, Iyare CO, Emelike CU, Ezimah AC, Asogwa NT, Konyefom NG. GC–MS analysis of Moringa oleifera leaf extract and effects of administration on histology of reproductive organs and liver of female rats exposed to chronic unpredictable stress. Food Chemistry Advances 2024 Jun 1;4:100661.##Bhargave A, Pandey I, Nama KS, Pandey M. Moringa oleifera Lam.—Sanjana (Horseradish Tree)—A miracle food plant with multipurpose uses in Rajasthan-India—an overview. Int J Pure App Biosci 2015;3(6):237-48.##Marcoccia M, De Carlo A. Iuliano L. The potential role of phytoestrogens in the prevention and treatment of gestational diabetes mellitus. Nutrients 2017;9(1):107.##Trease GE, Evans EC. A textbook of Pharmacognosy. 13th Ed. Bailliere Tindall, Britain: English Language Book Society. 2002;1985:386–480.##O'Boyle NM, Banck M, James CA, Morley C, Vandermeersch T, Hutchison GR. Open Babel: An open chemical toolbox. J Cheminform 2011 Oct 7;3:33.##Trott O, Olson AJ. AutoDock Vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem 2010 Jan 30;31(2):455-61.##Ajayi AF, Akhigbe RE. Staging of the estrous cycle and induction of estrus in experimental rodents: an update. Fertil Res Pract 2020 Mar 14;6:5.##Chukwu OO, Emelike, CU, Konyefom NG, Ibekailo SN, Ekakitie OO, Ghasi S, et al. Histological Studies of the Heart and Biochemical Changes Due to the Perinatal Consumption of Hibiscus sabdariffa (Flavonoid-rich Extract) to Feed-restricted Rats on Offspring. Iranian Journal of Veterinary Medicine 2022;17(1):37–46.##Brenes M, Fornaguera A. Effects of social isolation on the behavior and physiology of rodents: A review. Brain Research Reviews 2009;60(1):83-107.##Charil A, Fuchs E, Flugge G. Prenatal stress and the brain: Effects on morphology and function. Neuroscience and Biobehavioral Reviews 2010;34(8): 1357-71.##Keenan KJ, Miller MW, Gunnar MR. Prenatal stress and the developing brain: implications for child development. Perspectives on Psychological Science 2013;8(4):351-68.##Muller M, Sigurdsson S, Kjartansson O, Jonsson PV, Garcia M, von Bonsdorff MB, et al. Birth size and brain function 75 years later. Pediatrics 2014 Oct;134(4):761-70.##Levin ER. Minireview: Extranuclear steroid receptors: roles in modulation of cell functions. Mol Endocrinol 2011 Mar;25(3):377-84.##

Comparison of Anticancer Effects between Platinum Levetiracetam and Platinum Azidothymidine through the Expression of Biomarker Genes on Cancer Cell Lines 2 2 Background: The utilization of biomarkers is a way to assess the efficacy of recently created anticancer drugs. MiRNAs, telomerase, and Bcl-2 are extensively utilized as biomarkers for this purpose. This study aims to evaluate the comparison of the newly synthesized platinum compounds such as Platinum Azidothymidine (Pt-AZT) with Platinum Levetiracetam (Pt-Lev) on HepG2 cancer cell lines via the biomarkers. Methods: In this study, cells were divided into four groups: Group A (HDF cells) were the normal negative control group, group B were HepG2 untreated cancer cells, and groups C and D were treated cancer cells with Pt-AZT and Pt-Lev, respectively. After evaluating the LC50 for the drugs by MTT test, the relative gene expression of the biomarkers was determined by qPCR. Results: The results showed a significant decrease for antiapoptotic genes including miRNA-21 (5.1±0.014), telomerase (0.56±0.48), Bcl-2 (0.41±0.276) in group D, whereas in group C was more than group D for miRNA-21 (6.0±0.141), telomerase (3.49±0.231), Bcl-2 (4.93±0.276) also there was a significant increase in miRNA-122 (33.97±0.04) in group D, whereas in group C was (28.36±0.007) and so lower than group D. Most of the investigated groups showed a significant difference (p<0.05). In addition, there were widespread apoptotic regions in Pt-Lev treatment compared to Pt-AZT. Conclusion: The advantages of using Pt-Lev were more powerful anticancer effects on biomarkers through inhibition of antiapoptotic and stimulation proapoptotic factors and also lower side effects and lower drug resistance than Pt-AZT; therefore, it can be considered a more effective anti-cancer therapy. 24 30 Abdolreza Sabokrouh Nasim Ghaffari Department of Biochemistry and Biophysics, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University Department of Biochemistry and Biophysics, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University Iran Zahra Karimi Tafreshi Department of Biochemistry and Biophysics, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University Department of Biochemistry and Biophysics, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University Iran Freshteh Atabi Department of Biochemistry and Biophysics, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University Department of Biochemistry and Biophysics, Faculty of Advanced Sciences and Technology, Tehran Medical Sciences, Islamic Azad University Iran Zohreh Sharifi Blood Transfusion Research Center, High institute for Research and Education in Transfusion Medicine Iran Seyyed Khalil Shokouhi Mostafavi Department of Microbiology, Faculty of Medicine, Tehran Medical Sciences, Islamic Azad University Department of Microbiology, Faculty of Medicine, Tehran Medical Sciences, Islamic Azad University Iran Bcl-2LevetiracetammiRNAsTelomeraseZidovudine 60599.pdf Koeffler HP, McCormick F, Denny C. Molecular mechanisms of cancer. West J Med 1991 Nov; 155(5):505-14.##Debela DT, Muzazu SG, Heraro KD, Ndalama MT, Mesele BW, Haile DC, Kitui SK, Manyazewal T. New approaches and procedures for cancer treatment: Current perspectives. SAGE Open Med 2021 Aug 12; 9:20503121211034366.##Hu C, Dignam JJ. Biomarker-Driven Oncology Clinical Trials: Key Design Elements, Types, Features, and Practical Considerations. JCO Precis Oncol 2019 Oct 24; 3:PO.19.00086.##Sarhadi VK, Armengol G. Molecular Biomarkers in Cancer. Biomolecules 2022 Jul 23;12(8):1021.##Ohmura Y, Aoe M, Andou A, Shimizu N. Telomerase activity and Bcl-2 expression in non-small cell lung cancer. Clin Cancer Res 2000 Aug;6(8):2980-7.##Ratti M, Lampis A, Ghidini M, Salati M, Mirchev MB, Valeri N, et al. MicroRNAs (miRNAs) and Long Non-Coding RNAs (lncRNAs) as New Tools for Cancer Therapy: First Steps from Bench to Bedside. Target Oncol 2020 Jun;15(3):261-78.##Si W, Shen J, Zheng H, Fan W. The role and mechanisms of action of microRNAs in cancer drug resistance. Clin Epigenetics 2019 Feb 11;11(1):25.##Ratnasari N, Lestari P, Renovaldi D, Raditya Ningsih J, Qoriansas N, Wardana T, et al. Potential plasma biomarkers: miRNA-29c, miRNA-21, and miRNA-155 in clinical progression of Hepatocellular Carcinoma patients. PLoS One 2022 Feb 14;17(2):e0263298.##Faramin Lashkarian M, Hashemipour N, Niaraki N, Soghala S, Moradi A, Sarhangi S, et al. MicroRNA-122 in human cancers: from mechanistic to clinical perspectives. Cancer Cell Int 2023 Feb 20;23(1):29.##Feng YH, Tsao CJ. Emerging role of microRNA-21 in cancer. Biomed Rep 2016 Oct;5(4):395-402.##Nakao K, Miyaaki H, Ichikawa T. Antitumor function of microRNA-122 against hepatocellular carcinoma. J Gastroenterol 2014 Apr;49(4):589-93.##Coulouarn C, Factor VM, Andersen JB, Durkin ME, Thorgeirsson SS. Loss of miR-122 expression in liver cancer correlates with suppression of the hepatic phenotype and gain of metastatic properties. Oncogene 2009 Oct 8; 28(40):3526-36.##Bai S, Nasser MW, Wang B, Hsu SH, Datta J, Kutay H, et al. MicroRNA-122 inhibits tumorigenic properties of hepatocellular carcinoma cells and sensitizes these cells to sorafenib. J Biol Chem 2009 Nov 13;284(46):32015-27.##Sengupta D, Cassel T, Teng KY, Aljuhani M, Chowdhary VK, Hu P, et al. Regulation of hepatic glutamine metabolism by miR-122. Mol Metab 2020 Apr;34:174-86.##Dasari S, Tchounwou PB. Cisplatin in cancer therapy: molecular mechanisms of action. Eur J Pharmacol 2014 Oct 5;740:364-78.##Sharma R, Singh VJ, Chawla PA. Advancements in the Use of Platinum Complexes as Anticancer Agents. Anticancer Agents Med Chem 2022;22(5):821-35.##Sabokrouh A, Sadeghi Motlagh B, Atabi F. Study of anticancer effects of platinum levetiracetam and levetiracetam via cancer biomarkers genes expression on HepG2 cell line. Mol Biol Rep 2023 Nov;50(11):9431-9439.##Sabokrouh A, Atabi F, Jassem RM, Mohammadi R. The Anticancer Efficacy of Platinum Azidothymidin on Hepatocellular Carcinoma Via Affecting the Telomerase and the BcL-2 Genes Expression. J Gastrointest Cancer 2020 Sep;51(3):813-7.##Oliveira Rocha AM, Severo Sabedra Sousa F, Mascarenhas Borba V, S Munchen T, Guerin Leal J, Dorneles Rodrigues OE, et al. Evaluation of the effect of synthetic compounds derived from azidothymidine on MDA-MB-231 type breast cancer cells. Bioorg Med Chem Lett 2020 Sep 1;30(17):127365.##Chiu DT, Duesberg PH. The toxicity of azidothymidine (AZT) on human and animal cells in culture at concentrations used for antiviral therapy. Genetica 1995;95(1-3):103-9.##Shahabadi N, Pour Moghadam NH. Study on the interaction of the antiviral drug, zidovudine with DNA using neutral red (NR) and methylene blue (MB) dyes. Journal of luminescence 2013 Feb 1;134:629-34.##Sabokrouh A, Vaisi-Raygani A, Goodarzi MT, Khatami S, Taghizadeh-Jahed M, Shahabadi N, et al. Comparison between Platinum-Azidothymidine and Azidothymidine Effects on Bcl-2 and Telomerase Gene Expression in Rats with Hepatocellular Carcinoma. Avicenna J Med Biotechnol 2015 Apr-Jun;7(2):50-6.##Sabokrouh A, Sadeghi Motlagh B, Atabi F. Study of anticancer effects of platinum levetiracetam and levetiracetam via cancer biomarkers genes expression on HepG2 cell line. Mol Biol Rep 2023 Nov;50(11):9431-9.##Shu XL, Fan CB, Long B, Zhou X, Wang Y. The anti-cancer effects of cisplatin on hepatic cancer are associated with modulation of miRNA-21 and miRNA-122 expression. Eur Rev Med Pharmacol Sci 2016;20(21):4459-65.##Lin L, Tu HB, Wu L, Liu M, Jiang GN. MicroRNA-21 Regulates Non-Small Cell Lung Cancer Cell Invasion and Chemo-Sensitivity through SMAD7. Cell Physiol Biochem 2016;38(6):2152-62.##Demerdash HM, Hussien HM, Hassouna E, Arida EA. Detection of MicroRNA in Hepatic Cirrhosis and Hepatocellular Carcinoma in Hepatitis C Genotype-4 in Egyptian Patients. Biomed Res Int 2017:1806069.##Xu J, Wu C, Che X, Wang L, Yu D, Zhang T, et al.Circulating microRNAs, miR-21, miR-122, and miR-223, in patients with hepatocellular carcinoma or chronic hepatitis. Mol Carcinog 2011;50(2):136-42.##Wang W, Wei C. Advances in the early diagnosis of hepatocellular carcinoma. Genes Dis 2020;7(3):308-319.##Cao F, Yin LX. miR-122 enhances sensitivity of hepatocellular carcinoma to oxaliplatin via inhibiting MDR1 by targeting Wnt/β-catenin pathway. Exp Mol Pathol 2019 Feb;106:34-43.##Wang D, Sun X, Wei Y, Liang H, Yuan M, Jin F, et al. Nuclear miR-122 directly regulates the biogenesis of cell survival oncomiR miR-21 at the posttranscriptional level. Nucleic Acids Res 2018 Feb 28;46(4):2012-29.##

The Role of Disulfide Bridges in the Interaction of E. coli -Derived Recombinant SCARB2 and EV-A71's Capsid 2 2 Background: Hand, Foot, and Mouth disease is an acute infectious disease caused by a group of enteroviruses, including Coxsackievirus A16 and Enterovirus 71. EV-A71-causing disease can give rise to severe complications in children, leading to meningitis, encephalitis, and even death. A potential approach to prevent the virus spread is inhibiting the invasion of EV-A71 into target cells, thereby helping to prevent not only the spread of EV-A71 in the community but also lessen the risk of outbreaks. EV-A71 cell’s receptor, human scavenger receptor class B member 2, SCARB2, was used as a trap to gather the virus and limit its spreading. Methods: In this study, the recombinant receptor was expressed using Escherichia coli (E. coli) system. SCARB2 proteins expressed from E. coli BL21(DE3), and E. coli SHuffle® T7 Express were in inclusion bodies and subsequently refolded into soluble forms with recovery efficiencies of 57.57, and 82.2%, respectively. The presence of intramolecular disulfide bridges in the refolded SCARB2 was examined by SDS-PAGE in combination with Dithiothreitol (DTT). The two proteins were then utilized to evaluate the interaction with EV-A71 capsid by Indirect Enzyme-Linked Immunosorbent Assay (ELISA) at different pH levels to compare the adhesion efficiency. Results: The results showed that SCARB2 protein expressed from E. coli SHuffle® T7 Express with disulfide bond modifications had better adhesion to the viral capsid. Notably, when the medium pH was lowered to acidic levels, the binding efficiency of recombinant receptors to the viral capsid increased. Conclusion: To our knowledge, this study reported for the first time the activity of SCARB2 under extreme pH conditions and also revealed the crucial role of disulfide bridges in the interaction with EV-A71’s capsid. This finding contributed to the strategy using recombinant SCARB2 as a viral trap. 31 38 Hai-Vy Vo-Nguyen Department Molecular and Environmental Biotechnology, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Biosensors, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Molecular Biotechnology, University of ScienceVietnam National University Department Molecular and Environmental Biotechnology, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Biosensors, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Molecular Biotechnology, University of ScienceVietnam National University VietnamVietnamVietnamVietnam Tran Linh Thuoc Department Molecular and Environmental Biotechnology, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Biosensors, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Molecular Biotechnology, University of ScienceVietnam National University Department Molecular and Environmental Biotechnology, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Biosensors, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Molecular Biotechnology, University of ScienceVietnam National University VietnamVietnamVietnamVietnam Hieu Tran-Van Laboratory of Biosensors, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Molecular Biotechnology, University of ScienceVietnam National University Laboratory of Biosensors, Faculty of Biology and Biotechnology, University of ScienceLaboratory of Molecular Biotechnology, University of ScienceVietnam National University VietnamVietnamVietnam Capsid proteinsEnterovirus infectionsEscherichia coli (E. coli)Foot and mouth disease 60600.pdf World Health Organization. A guide to clinical management and public health response for hand, foot and mouth disease (HFMD). 2011. WHO Regional Office for the Western Pacific.##Yamayoshi S, Iizuka S, Yamashita T, Minagawa H, Mizuta K, Okamoto M, et al. Human SCARB2-dependent infection by coxsackievirus A7, A14, and A16 and enterovirus 71. J Virol 2012 May;86(10):5686-96.##Shah J, Sijun L, Hui Z, Zeb F, Haq IU, Ullah A. Neurological Complications Of Hand, Foot And Mouth Disease In Children: A Review. J Ayub Med Coll Abbottabad 2020 Oct-Dec;32(4):562-9.##Lee KY. Enterovirus 71 infection and neurological complications. Korean J Pediatr 2016 Oct;59(10):395-401.##Bessaud M, Razafindratsimandresy R, Nougairède A, Joffret ML, Deshpande JM, Dubot-Pérès A, et al. Molecular comparison and evolutionary analyses of VP1 nucleotide sequences of new African human enterovirus 71 isolates reveal a wide genetic diversity. PLoS One 2014 Mar 5;9(3):e90624.##Tong Y, Zhang X, Chen J, Chen W, Wang Z, Li Q, et al. Immunogenicity and safety of an enterovirus 71 vaccine in children aged 36-71 months: A double-blind, randomised, similar vaccine-controlled, non-inferiority phase III trial. EClinicalMedicine 2022 Jul 29;52:101596.##Yi EJ, Shin YJ, Kim JH, Kim TG, Chang SY. Enterovirus 71 infection and vaccines. Clin Exp Vaccine Res 2017 Jan;6(1):4-14.##Zhou D, Zhao Y, Kotecha A, Fry EE, Kelly JT, Wang X, et al. Unexpected mode of engagement between enterovirus 71 and its receptor SCARB2. Nat Microbiol 2019 Mar;4(3):414-419.##Yamayoshi S, Koike S. Identification of a human SCARB2 region that is important for enterovirus 71 binding and infection. J Virol 2011 May;85(10):4937-46.##Kobayashi K, Koike S. Cellular receptors for enterovirus A71. J Biomed Sci 2020 Jan 10;27(1):23.##Dang M, Wang X, Wang Q, Wang Y, Lin J, Sun Y, Li X, et al. Molecular mechanism of SCARB2-mediated attachment and uncoating of EV71. Protein Cell 2014 Sep;5(9):692-703.##Bechtel TJ, Weerapana E. From structure to redox: The diverse functional roles of disulfides and implications in disease. Proteomics 2017 Mar;17(6):10.##Manteca A, Alonso-Caballero Á, Fertin M, Poly S, De Sancho D, Perez-Jimenez R. The influence of disulfide bonds on the mechanical stability of proteins is context dependent. J Biol Chem 2017 Aug 11;292(32):13374-13380.##o-Nguyen HV, Nguyen TT, Vu HT, Nguyen TT, Hoang QC, Tran TL, Tran-Van H. Recombinant Human SCARB2 Expressed in Escherichia coli and its Potential in Enterovirus 71 Blockage. Iran J Sci Technol Trans A Sci 2021;45(2):455-461.##Leemasawatdigul K, Gappa-Fahlenkamp H. Effect of storage conditions on the stability of recombinant human MCP-1/CCL2. Biologicals 2011 Jan;39(1):29-32.##Umbarkar CP, Thakre PS, Kashikar SA, Ghanwate NA, Agarkar VB. Cloning, expression, and refolding of Nsuccinyl-ll diaminopimelic acid desuccinylase (DapE) of Mycobacterium tuberculosis. IJLSR 2019;7:110-5.##Quynh-Anh Nguyen-Ngoc T-TTN, Minh-Anh Dang-Trinh, Khoa Van Nguyen, Bich-Phuong Thi Tran, Hanh-Lan Thi Nguyen, Khoi-Nguyen Le-Phan, et al. Enterovirus 71 (EV71) neutralization effect of IgY antibodies obtained from egg yolk. Vietnam Journal of Preventive Medicine 2015;5:144.##Thao NT, Ngoc NT, Tú PV, Thúy TT, Cardosa MJ, McMinn PC, et al. Development of a multiplex polymerase chain reaction assay for simultaneous identification of human enterovirus 71 and coxsackievirus A16. J Virol Methods 2010 Dec;170(1-2):134-9.##Zou S, Guo J, Gao R, Dong L, Zhou J, Zhang Y, et al. Inactivation of the novel avian influenza A (H7N9) virus under physical conditions or chemical agents treatment. Virol J 2013 Sep 15;10:289.##Lobstein J, Emrich CA, Jeans C, Faulkner M, Riggs P, Berkmen M. SHuffle, a novel Escherichia coli protein expression strain capable of correctly folding disulfide bonded proteins in its cytoplasm. Microb Cell Fact 2012 May 8;11:56.##Rinas U, Garcia-Fruitós E, Corchero JL, Vázquez E, Seras-Franzoso J, Villaverde A. Bacterial Inclusion Bodies: Discovering Their Better Half. Trends Biochem Sci 2017 Sep;42(9):726-37.##Rosano GL, Ceccarelli EA (2014) Recombinant protein expression in Escherichia coli: advances and challenges. Frontiers in Microbiology 5.##Kumar D. Comprehensive Biotechnology vol Volume 2: Engineering Fundamentals of Biotechnology. 2nd edn. 2011. Elsevier.##Mattei B, Lira RB, Perez KR, Riske KA. Membrane permeabilization induced by Triton X-100: The role of membrane phase state and edge tension. Chem Phys Lipids 2017 Jan;202:28-37.##Ma Y, Tonelli M, Unsworth LD (2021) Effect of carbamylation on protein structure and adsorption to self-assembled monolayer surfaces. Colloids and Surfaces B: Biointerfaces 203:111719##Organic Chemistry: Structure, Mechanism, and Synthesis. Robert J. Ouellette and J. David Rawn. 2014. Elsevier. 1203. p. 27.##Boonyuen U, Promnares K, Junkree S, Day NP, Imwong M. Efficient in vitro refolding and functional characterization of recombinant human liver carboxylesterase (CES1) expressed in E. coli. Protein Expr Purif 2015 Mar;107:68-75.##Mollania N, Khajeh K, Ranjbar B, Rashno F, Akbari N, Fathi-Roudsari M. An efficient in vitro refolding of recombinant bacterial laccase in Escherichia coli. Enzyme Microb Technol 2013 May 10;52(6-7):325-30.##SSchmid M, Prinz TK, Stäbler A, Sängerlaub S. Effect of Sodium Sulfite, Sodium Dodecyl Sulfate, and Urea on the Molecular Interactions and Properties of Whey Protein Isolate-Based Films. Front Chem 2017 Jan 18;4:49.##Zhao Y, Ren J, Padilla-Parra S, Fry EE, Stuart DI. Lysosome sorting of β-glucocerebrosidase by LIMP-2 is targeted by the mannose 6-phosphate receptor. Nat Commun 2014 Jul 14;5:4321.##

The Induction of L-lysine-α-Oxidase from Trichoderma Harzianum Rifai by Metabolic Products of Brevibacterium sp. and the Improvement of Its Isolation and Purification Techniques 2 2 Background: The enzyme L-Lysine-α-oxidase from Trichoderma harzianum Rifai is a promising anticancer, antifungal and antibacterial agent. Intensive exploring of its physico-chemical properties and possible ways of application requires sufficient amounts of the protein which in turn depends on good techniques of cultivation of the micro-organism producer, enzyme soft isolation and purification "and storage". Methods: An improved method has been suggested for isolation and purification of the enzyme. A specific combination of column sorbents was adapted and gradient elution with sodium chloride was applied to elevate the yield of the enzyme. The inductive influence of Metabolic Products (MP) of the Brevibacterium species, along with fungal metabolites of Ulocladium sp. and Trichoderma sp. was tested. The enzyme activity assay was based on the detection of oxidized dimethylbenzidine in a peroxidase reaction coupled with an L-lysine-α-oxidase reaction. Some enzyme properties were additionally explored. Results: The upgraded technique of isolation and purification resulted in a yield of enzyme of about 79%. All strains of Brevibacterium sp. proved to be potent enhancers of L-lysine-α-oxidase activity and concomitant activities. The induced enzyme appeared to be less specific but more thermostable. Possible application scopes for the enzyme with modified properties are discussed. Phosphate buffer solution (pH=5.6) appeared to be the best one for long-term storage of the enzyme. Conclusion: A significant inducing effect of MP of Brevibacterium sp. on L-lysine-α-oxidase has been detected, and its isolation and purification techniques have been improved. 39 46 Irina Smirnova T.T. Berezov Department of Biochemistry, Institute of Medicine, Peoples’ Friendship University of Russia (RUDN University) T.T. Berezov Department of Biochemistry, Institute of Medicine, Peoples’ Friendship University of Russia (RUDN University) Russian Federation Ekaterina Neborak Yuri Shneyder Federal State Budgetary Institution All-Russian Plant Quarantine Center Federal State Budgetary Institution All-Russian Plant Quarantine Center Russian Federation Ida Bashkirova Federal State Budgetary Institution All-Russian Plant Quarantine Center Federal State Budgetary Institution All-Russian Plant Quarantine Center Russian Federation Elena Karimova Federal State Budgetary Institution All-Russian Plant Quarantine Center Federal State Budgetary Institution All-Russian Plant Quarantine Center Russian Federation Daria Baranova Free University of Bozen-Bolzano, 39100 Free University of Bozen-Bolzano, 39100 Italy Maria Ploskonos Department of Fundamental Chemistry, Astrakhan State Medical University Department of Fundamental Chemistry, Astrakhan State Medical University Russia Lyudmila Gavrilyuk T.T. Berezov Department of Biochemistry, Institute of Medicine, Peoples’ Friendship University of Russia (RUDN University) T.T. Berezov Department of Biochemistry, Institute of Medicine, Peoples’ Friendship University of Russia (RUDN University) Russian Federation Valeriy Shkinev Vernadsky Institute of Geochemistry and Analytical Chemistry Vernadsky Institute of Geochemistry and Analytical Chemistry Russian Federation Victor Larichev The Gamaleya National Research Center of Epidemiology and Microbiology The Gamaleya National Research Center of Epidemiology and Microbiology Russian Federation Antibacterial agentAntifungalAntineoplastic agentsBrevibacteriumL-lysine oxidaseTrichodermaTrichoderma harzianum 60601.pdf Smirnova IP, Alekseev SB, Shevchenko AA. [Biosynthesis of L-lysine-alpha-oxidase, an antitumor enzyme, by Trichoderma spp]. Antibiot Khimioter 2009;54(5–6):8–12. Russian.##Itada N, Ichihara Akira, Makita Tsutomu, Hayaishi Osamu, Suda Masami, Sasaki N. L-Lysine Oxidase, a New Oxygenase. J Biochem 1961 Jun;50(2):118–21.##Kusakabe H, Kodama K, Kuninaka A, Yoshino H, Misono H, Soda K. A new antitumor enzyme, L-lysine alpha-oxidase from Trichoderma viride. Purification and enzymological properties. J Biol Chem 1980 Jun;255(3):976–81.##Smirnova IP, Shneider YuA, Karimova E V. Trichoderma L-Lysine-α-Oxidase Producer Strain Culture Fluid Inhibits Impatiens Necrotic Spot Virus. Bull Exp Biol Med 2016 Jun;160(3):357–9.##Smirnova IP, Karimova E V, Shneider YuA, Volina EG. L-Lysine-α-Oxidase: Acidovorax citrulli Bacterium Inhibitor. Bull Exp Biol Med 2018 Jun;164(4):459–61.##Costa MN, Silva RN. Cytotoxic activity of l-lysine alpha-oxidase against leukemia cells. Semin Cancer Biol 2022 Jun;86 (Pt 3):590–9.##Lukasheva E V, Babayeva G, Karshieva SS, Zhdanov DD, Pokrovsky VS. L-Lysine α-Oxidase: Enzyme with Anticancer Properties. Pharmaceuticals (Basel) 2021 Jun;14(11):1070.##Smirnova IP, Karimova E V, Shneider YuA. Antibacterial Activity of L-Lysine-α-Oxidase from the Trichoderma. Bull Exp Biol Med 2017 Jun;163(6):777–9.##Smirnova IP, Kuznetsov VI, Podoprigora I V, Mansur TI, Senyagin AN, Bashkirova IG. Characterization of L-Lysine-A-Oxidase, A New Antitumor and Antiviral Drug Substance Synthesized By Trichoderma. Int J Life Sci Pharma Res 2022 Jun;L15–20.##Smirnova IP, Diorditsa S V, Alekseev SB, Zaĭtsev IZ. [Effect of L-lysine-a-oxidase on reproduction of herpes simplex type I virus in vitro]. Vopr Med Khim 1998;44(4):384–7. Russian.##Smirnova IP, Alekseev SB, Podboronov VM. [Effect of L-lysine-alpha-oxidase gel on development of ophthalmic herpes and herpetic skin lesions in rabbit]. Antibiot Khimioter 2003;48(11):7–9. Russian.##Smirnova IP, Khaduev SK. [L-lysine-alpha-oxidase activity of some Trichoderma species]. Mikrobiologiia 1984;53(1):163–5. Russian.##Iankevich NB, Laugalene NF, Vesa VS, Khaduev SK, Smirnova IP. [Improved method of purification of L-lysine-alpha-oxidase from the fungus Trichoderma sp]. Vopr Med Khim 1989;35(2):84–6. Russian.##Potanova OL, Smirnova IP, Turchinskiĭ V V, Berezov TT. [Use of o-phenylenediamine in determining the activity of L-lysine- alpha-oxidase and concentration of L-lysine]. Vopr Med Khim 1992;38(5):28–30. Russian.##Potanova OL, Smirnova IP, Turchinskiĭ V V, Berezov TT. [Use of o-phenylenediamine in determining the activity of L-lysine- alpha-oxidase and concentration of L-lysine]. Vopr Med Khim 1992;38(5):28–30. Russian##Khaduev SK, Lukasheva E V, Smirnova IP, Iankevich NB, Laugalene NF. [Various physico-chemical properties of L-lysine-alpha-oxidase from Trichoderma sp]. Vopr Med Khim 1988;34(4):97–100. Russian.##Potapova OL, Smirnova IP, Vesa VS, Bykova OM, Laugalene NF. [Catalytic properties of induced L-lysine-alpha-oxidase]. Vopr Med Khim 1992;38(1):9–13. Russian.##Khaduev SK, Lukasheva E V, Smirnova IP, Berezov TT. [Isolation and purification of L-lysine-alpha-oxidase from Trichoderma sp]. Vopr Med Khim 1985;31(5):130–4. Russian.##Smirnova IP, Siatkin SP, Berezov TT. [Spectrophotometric micromethod to determine L-lysine-alpha-oxidase activity]. Vopr Med Khim 1984;30(1):133–6. Russian.##Smirnova IP, Berezov TT. [Ortho-dianizidine micromethod of determining L-phenylalanine-alpha-oxidase activity]. Vopr Med Khim 1988;34(2):129–31. Russian.##Pokrovsky VS, Treshalina HM, Lukasheva E V, Sedakova LA, Medentzev AG, Arinbasarova AY, et al. Enzymatic properties and anticancer activity of L-lysine α-oxidase from Trichoderma cf. aureoviride Rifai BKMF-4268D. Anticancer Drugs 2013 Sep;24(8):846–51.##Pokrovsky VS, Lukashev AN, Babayeva G, Karshieva SS, Arinbasarova AY, Medentzev AG, et al. Plasma pharmacokinetics and tissue distribution of L-lysine α-oxidase from Trichoderma cf. aureoviride RIFAI VKM F- 4268D in mice. Amino Acids 2021 Jun;53(1):111–8.##Arinbasarova AYu, Botin AS, Medentsev AG, Makrushin K V, Vetcher AA, Stanishevskiy YM. Synthesis of Extracellular L-lysine-α-oxidase along with Degrading Enzymes by Trichoderma cf. aureoviride Rifai VKM F-4268D: Role in Biocontrol and Systemic Plant Resistance. J Fungi (Basel) 2024 Jun;10(5):323.##Yuan H, Yuan M, Shi B, Wang Z, Huang T, Zhu J, et al. Biocontrol activity of Bacillus halotolerans strain Pl7 against Botryosphaeria dothidea causing apple postharvest decay and potential mechanisms. Front Microbiol 2023 Jun;13.##Chacón-López A, Guardado-Valdivia L, Bañuelos-González M, López-García U, Montalvo-González E, Arvizu-Gómez J, et al. Effect of Metabolites Produced by Bacillus atrophaeus and Brevibacterium frigoritolerans Strains on Postharvest Biocontrol of Alternaria alternata in Tomato (Solanum lycopersicum L.). Biocontrol Sci 2021;26(2):67–74.##Duncker KE, Holmes ZA, You L. Engineered microbial consortia: strategies and applications. Microb Cell Fact 2021 Jun;20(1):211.##Senyagin AN, Larichev AF, Smirnova IP, Podoprigora I V. A Novel Express Method to Determine Activity of Antitumor Enzyme L-Lysine-α-Oxidase of Trichoderma harzianum Rifai F-180. Bull Exp Biol Med 2020 Jun;169(1):119–21.##Sushko V.I., Shkolnik I I, Shchetkin V.V., Ter-Sargsyan E.M. Method for obtaining l-lysine. Latvia patent RU2125608C1. 1999 Jan 27.##Gusyatiner MM, Luncz MG, Ivanovskaya LV, Rostova Yu G, Bachina TA, Axverdyan VZ, et al. Shtamm bakterij Escherichia coli producent l-lejcina (varianty`) [L-Leucine producer bacteria strain Escherichia coli (variants)]. Russian Federation patent Patent RF RU2140450С1. 1999 Oct 27.##Keleshyan SK, Karapetyan Zh V, Chakhalyan AKh, Hovhannisyan NA, Avetisova GYe, Chitchyan MB, et al. Influence of glutamate dehydrogenase activity on L-proline synthesis. Appl Biochem Microbiol 2017 Jun;53(5):518–23. ##Lurie Yu. Handbook of Analytical Chemistry. 6th ed. Moscow: Chimie Publisher; 1989. 231–239 p.##Kondo H, Kitagawa M, Matsumoto Y, Saito M, Amano M, Sugiyama S, et al. Structural basis of strict substrate recognition of l-lysine α-oxidase from Trichoderma viride. Protein Sci 2020 Jun;29(11):2213–25.##Amano M, Mizuguchi H, Sano T, Kondo H, Shinyashiki K, Inagaki J, Tamura T, Kawaguchi T, Kusakabe H, Imada K, Inagaki K. Recombinant expression, molecular characterization and crystal structure of antitumor enzyme, L-lysine α-oxidase from Trichoderma viride. J Biochem 2015 Jun;157(6):549-59.##Tani Y, Miyake R, Yukami R, Dekishima Y, China H, Saito S, Kawabata H, Mihara H. Functional expression of L-lysine α-oxidase from Scomber japonicus in Escherichia coli for one-pot synthesis of L-pipecolic acid from DL-lysine. Appl Microbiol Biotechnol 2015 Jun;99(12):5045-54.##Creasy A, Barker G, Carta G. Systematic interpolation method predicts protein chromatographic elution with salt gradients, pH gradients and combined salt/pH gradients. Biotechnol J 2017 Mar;12(3).##Lo C-F, Yu C-Y, Kuan I-C, Lee S-L. Optimization of lipase production by Burkholderia sp. using response surface methodology. Int J Mol Sci 2012 Nov 13;13(11):14889–97.##Gupta N, Mehra G, Gupta R. A glycerol-inducible thermostable lipase from Bacillus sp.: medium optimization by a Plackett-Burman design and by response surface methodology. Can J Microbiol 2004 May;50(5):361–8.##Farzaneh F, Mirzaie S, Dehnavi E, Aghaeepoor M, Farzaneh S, Pourzardosht N, et al. Response Surface Methodology to Optimize the Expression Efficiency of Recombinant Reteplase. Iran J Biotechnol 2023 Apr;21(2):e3288.##Pokrovsky VS, Chepikova OE, Davydov DZh, Jr AAZ, Lukashev AN, Lukasheva E V. Amino Acid Degrading Enzymes and their Application in Cancer Therapy. Curr Med Chem 2019 Jun;26(3):446–64.##

Antidiabetic Activity of Momordica charantia Extracts Through Incretin Pathway in Streptozotocin-Nicotinamide Induced Diabetic Rat Depends on Dose Differences 2 2 Background: This study addresses the increasing prevalence of type 2 Diabetes Mellitus (T2DM) and the need for alternative, cost-effective medications. The research problem focuses on the need to further study the dose effectiveness of Momordica charantia Extracts (MCE) on T2DM parameters, including Glucagon-Like Peptide 1 (GLP-1), Dipeptidyl Peptidase 4 (DPP-4), alpha glucosidase, glucosetransporter 5 (GLUT5), and pancreatic tissue histopathology. Methods: The methodology employed an experimental research design with 30 Wistar rats divided into six groups, each receiving different inductions and doses. Parameters were measured using Elisa, and histological analysis of pancreatic tissue was conducted using HE staining. Results: The Kruskal-Wallis test revealed significant differences in each group for the GLP-1 (p=0.003). However, the DPP4 test suggested a lack of significant difference in each group (p=0.192), and the GLUT5 test showed insignificant changes between each group (p=0.119). The ANOVA analysis on alpha-glucosidase revealed no statistically significant differences among the groups (p=0.202). Additionally, a qualitative examination of the histological analysis of pancreatic tissue indicated an improvement in the condition of the pancreatic tissue. Conclusion: MCE can increase GLP-1 levels, lower DPP-4, lower alpha-glucosidase, and raise GLUT5. However, there are no significant differences between other groups and the morphology of pancreatic tissue in rat model T2DM at a dose of 300 mg/kg. 47 55 Muhammad Fadhol Romdhoni Muchsin Doewes Faculty of Medicine, Universitas Sebelas MaretDepartment of Pharmacology, Faculty of Medicine, Universitas Sebelas Maret Faculty of Medicine, Universitas Sebelas MaretDepartment of Pharmacology, Faculty of Medicine, Universitas Sebelas Maret IndonesiaIndonesia Soetrisno Soetrisno Faculty of Medicine, Universitas Sebelas MaretDepartment of Obstetrics and Gynaecology, Faculty of Medicine, Universitas Sebelas Maret Faculty of Medicine, Universitas Sebelas MaretDepartment of Obstetrics and Gynaecology, Faculty of Medicine, Universitas Sebelas Maret IndonesiaIndonesia Ratih Puspita Febrinasari Faculty of Medicine, Universitas Sebelas MaretDepartment of Pharmacology, Faculty of Medicine, Universitas Sebelas Maret Faculty of Medicine, Universitas Sebelas MaretDepartment of Pharmacology, Faculty of Medicine, Universitas Sebelas Maret IndonesiaIndonesia Dipeptidyl peptidase 4Glucagon-like peptide 1Momordica charantia extractsRatsType 2 diabetes mellitus 60602.pdf Indonesia PE. Pengelolaan dan pencegahan diabetes melitus tipe 2 di Indonesia. Pb. Perkeni. 2015;6.##Ling C. Epigenetic regulation of insulin action and secretion – role in the pathogenesis of type 2 diabetes. J Intern Med 2020;288(2):158-67.##International Diabetes Federation. IDF Diabetes Atlas. Vol 102.; 2021.##Kementrian Kesehatan Republik Indonesia. Infodatin 2020 Diabetes Melitus. Published online 2019.##Goyal R, Jialal I. Diabetes Mellitus, Type 2.; 2020.##Diabetes Canada Clinical Practice Guidelines Expert Committee; Punthakee Z, Goldenberg R, Katz P. Definition, Classification and Diagnosis of Diabetes, Prediabetes and Metabolic Syndrome. Can J Diabetes 2018;42 Suppl 1:S10-S15.##Kelly SD. Ominous Octet and Other Scary Diabetes Stories: The Overview of Pathophysiology of Type 2 Diabetes Mellitus. Physician Assist Clin 2020;5(2):121-33.##Bhat GA, Hussain SA, Bhat NA, Habib S, Bhat MI. C Bhat GA, Hussain SA, Bhat NA, Habib S, Bhat MI. Comparative effect of aqueous extract of trigonella foenum-graecum and momordica charantia on diabetic wistar rats.##Singh P, Mishra A, Singh P, Goswami S, Singh A, Tiwari KD. Diabetes mellitus and use of medicinal plants for its treatment. Indian Journal of Research in Pharmacy and Biotechnology 2015 Sep 1;3(5):351.##St Onge EL, Miller S, Clements E. Sitagliptin/Metformin (janumet) as combination therapy in the treatment of type-2 diabetes mellitus. P T 2012 Dec;37(12):699-708.##Documentary Educational Resources (Firm), John Marshall. Bitter Melons.; 1966. Accessed July 13, 2024.##Peter EL, Nagendrappa PB, Hilonga S, Tuyiringire N, Ashuro E, Kaligirwa A, et al. Pharmacological reflection of plants traditionally used to manage diabetes mellitus in Tanzania. J Ethnopharmacol 2021 Apr 6;269:113715.##Mahwish, Saeed F, Sultan MT, Riaz A, Ahmed S, Bigiu N, et al. Bitter Melon (Momordica charantia L.) Fruit Bioactives Charantin and Vicine Potential for Diabetes Prophylaxis and Treatment. Plants (Basel) 2021 Apr 8;10(4):730.##Parawansah P, Sudayasa IP, Syarifin AN, Eso A, Nuralifah N, Fathanah WO, et al. Momordica charantia L. fruit fractions inhibit malondialdehyde level and regenerate hepatic damage of hyperglycemic rats. The Indonesian Biomedical Journal 2020 Mar 19;12(1):57-61.##Hendro OS, Yudhanto E, Himawan AB, Adespin DA. Effect of Momordica charantia fruit Ethanolic Extract on Malondialdehyde (MDA) level in blood of Sprague-dawley rats induced by 7, 12-dimethylbenz [a] anthracene. Jurnal Kedokteran Diponegoro (Diponegoro Medical Journal) 2022 Jan 17;11(1):32-6.##Poovitha S, Parani M. Protein extracts from Momordica charantia var. charantia and M. charantia var. muricata show anti-lipidemic and antioxidant properties in experimental type 2 diabetic rats. J Food Biochem 2020;44(9).##Ayoub SM, Rao S, Byregowda SM, Satyanarayana ML, Bhat N, Shridhar NB, Shridhar PB. Evaluation of hypoglycemic effect of Momordica charantia extract in distilled water in streptozotocin-diabetic rats. Braz J Vet Pathol 2013 Aug 22;6(2):56-64.##Raza H, John A. Streptozotocin-induced cytotoxicity, oxidative stress and mitochondrial dysfunction in human hepatoma HepG2 cells. Int J Mol Sci 2012;13(5):5751-67.##Zhu Y, Bai J, Zhang Y, Xiao X, Dong Y. Effects of bitter melon (Momordica charantia L.) on the gut microbiota in high fat diet and low dose streptozocin-induced rats. Int J Food Sci Nutr 2016;67(6):686-95.##Bhat GA, Khan HA, Alhomida AS, Sharma P, Singh R, Paray BA. GLP-I secretion in healthy and diabetic Wistar rats in response to aqueous extract of Momordica charantia. BMC Complement Altern Med 2018;18(1):162.##Kusniawati MA. Sistematik Review Aktivitas Buah Pare (Momordica charantia L.) Terhadap Target Molekuler Antidiabetes. Jurnal Farmasi Indonesia 2021 Nov 20;18(2):141-51.##Oyelere SF, Ajayi OH, Ayoade TE, Pereira GBS, Owoyemi BCD, Ilesanmi AO, et al. Corrigendum to "A detailed review on the phytochemical profiles and antidiabetic mechamisms of momordica charantia" [Heliyon 8 4 April 2022 Article e09253]. Heliyon 2023 Nov 2;9(11):e22019.##Chang CI, Cheng SY, Nurlatifah AO, Sung WW, Tu JH, Lee LL, Cheng HL. Bitter Melon Extract Yields Multiple Effects on Intestinal Epithelial Cells and Likely Contributes to Anti-diabetic Functions. Int J Med Sci 2021 Feb 24;18(8):1848-56.##Perumal V, Khatib A, Ahmed QU, Uzir BF, Abas F, Murugesu S, Saiman MZ, Primaharinastiti R, El-Seedi H. Correlation of the GC-MS-based metabolite profile of Momordica charantia fruit and its antioxidant activity. International Food Research Journal 2022;29(1):58-66.##Arif R, Ahmad S, Mustafa G, Mahrosh HS, Ali M, Tahir Ul Qamar M, et al. Molecular Docking and Simulation Studies of Antidiabetic Agents Devised from Hypoglycemic Polypeptide-P of Momordica charantia. Biomed Res Int 2021 Sep 17;2021:5561129.##Deng Y, Zhang Y, Liu G, Zhou P, Li P, Zhao Z, et al. Saponins from Momordica charantia exert hypoglycemic effect in diabetic mice by multiple pathways. Food Sci Nutr 2023 Sep 18;11(12):7626-7637.##Dahlquist A, Jandali D, Nauman MC, Johnson JJ. Clinical application of Momordica char-antia (Bitter Melon) for reducing blood sugar in type 2 diabetes mellitus. International Journal of Nutrition 2023 Oct 19:8-26.##Krawczyk M, Burzynska-Pedziwiatr I, Wozniak LA, Bukowiecka-Matusiak M. Evidence from a Systematic Review and Meta-Analysis Pointing to the Antidiabetic Effect of Polyphenol-Rich Plant Extracts from Gymnema montanum, Momordica charantia and Moringa oleifera. Curr Issues Mol Biol 2022;44(2):699-717.##Uebanso T, Arai H, Taketani Y, Fukaya M, Yamamoto H, Mizuno A, et al. Extracts of Momordica charantia suppress postprandial hyperglycemia in rats. J Nutr Sci Vitaminol (Tokyo) 2007 Dec;53(6):482-8.##Keller AC. A characterized saponin-rich fraction of Momordica charantia shows antidiabetic activity in C57BLK/6 mice fed a high fat diet. Phytomedicine Plus 2021;1(4):100134.##Xiao G, Xiao X. Antidiabetic effect of hydro-methanol extract of prunus cerasus L fruits and identification of its bioactive compounds. Tropical Journal of Pharmaceutical Research 2019;18(3):597-602.##

Repellency Effects, Cytotoxicity Assessment and Chemical Compositions of Citrus limon Seeds Extracts and Fractions Against Anopheles gambiae sl Mosquitoes 2 2 Background: Citrus limon seeds have limited exploration, compared to other parts. The increasing prevalence of mosquito menace prompted the investigation into the repellent effects of the seed n-hexane extract and fractions of Citrus limon (Rutaceae) against adult Anopheles gambiae sl mosquitoes. The cytotoxicity and chemical constituents of the 100% n-Hexane Fraction (HF) were also evaluated. Methods: Successive extraction was done with cold-maceration using, n-hexane, Ethyl Acetate (EA) and methanol. The three solvent extracts were screened against Anopheles gambiae sl mosquitos, using human bait method, at doses, 1.5, 2.5 and 5.0 mg/ml and with Deet and acetone used as positive and negative controls respectively. The n-hexane extract was further fractionated with column chromatography. Ten fractions (100% n-hexane, 10, 20, 30, 40, 60, 80% EA in n-hexane, 100% EA, 50% methanol in EA, and 100% methanol) were collected, and all tested at a single optimized dose of 5 mg/ml for 2 hr. The cytotoxicity and the chemical constituents of HF were determined using brine shrimp lethality test and gas chromatography-mass spectrometry respectively. Results: N-hexane extract and HF, both at 5 mg/ml, showed the most protective effect of 97.52 and 89.40% respectively, compared with standard (Odomos Deet) 100% activity, with no significant difference (p>0.05) between the three. HF was toxic with LC50 values of 29.2354 ppm. Thirty compounds representing 79.25% of HF were identified. Major compounds present in the fraction were heptadecanamine (8.31%), N-(3-methylbutyl) acetamide (7.89%), 2-ethyl- 1-hexanamine (6.65%). Conclusion: This promising repellent activity further justified the traditional usage of Citrus limon as mosquito repellent. 56 63 Mukaram Akintunde Adeniyi-Akee Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ibadan, Nigeria Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ibadan, Nigeria Nigeria Kingsley Adibe Mbachu Department of Chemistry, Faculty of Science, University of Ibadan Department of Chemistry, Faculty of Science, University of Ibadan Nigeria Kolade Ibrahim Department of Zoology, Faculty of Science, University of Ibadan Department of Zoology, Faculty of Science, University of Ibadan Nigeria Paul Matthew Osamudiamen Department of Chemistry, Faculty of Science, University of IbadanDepartment of Chemical Sciences, Bells University of Technology Department of Chemistry, Faculty of Science, University of IbadanDepartment of Chemical Sciences, Bells University of Technology NigeriaNigeria Olapeju Oluyemisi Aiyelaagbe Department of Chemistry, Faculty of Science, University of Ibadan Department of Chemistry, Faculty of Science, University of Ibadan Nigeria Sunday Olakunle Idowu Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ibadan Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ibadan Nigeria Olajire Aremu Adegoke Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ibadan, Nigeria Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Ibadan, Nigeria Nigeria AnophelesCitrusGas chromatography-mass spectrometryInsect repellents 60603.pdf Adeniran OI, Fabiyi E. A cream formulation of an effective mosquito repellent: a topical product from lemongrass oil (Cymbopogon citratus) Stapf.##World Health Organization (1999). Malarial fact sheet. No. 94. Geneva (accessed May 3, 2002 at http:www.who.int/inf.Fs/en/fact 094.html).##World Health Organisation (2019). World Malaria Report 2019.##World Health Organisation. (1996). Report of the WHO informal consultation on the evaluation and testing of insecticides CTD/WHO PES/IC/96.1. Geneva 69.##Toure Y. (2002). Malaria vector control strategies and challenges in Africa, Mali. In: Epidemiology of Parasitic diseases. University of Mali Press.##Thorsell W, Mikiver A, Malander I, Tunón H. Efficacy of plant extracts and oils as mosquito repellents. Phytomedicine 1998 Aug;5(4):311-23.##Govere J, Durrheim DN, Du Toit N, Hunt RH, Coetzee M. Local plants as repellents against Anopheles arabiensis, in Mpumalanga Province, South Africa. Cent Afr J Med 2000 Aug;46(8):213-6.##Isman MB, Miresmailli S, Machial C. Commercial opportunities for pesticides based on plant essential oils in agriculture, industry and consumer products. Phytochemistry Reviews 2011 Jun;10:197-204.##Ahmad MM, Iqbal Z, Anjum FM, Sultan JI. Genetic variability to essential oil composition in four citrus fruit species. Pakistan Journal of Botany 2006 Feb;38(2):319.##Tomotake H, Koga T, Yamato M, Kassu A, Ota F. Antibacterial activity of citrus fruit juices against Vibrio species. Journal of Nutritional Science and Vitaminology 2006;52(2):157-60.##Anyanwu GI, Amaefule EC, Ngurukwem C. Larvicidal effects of lemon peels on mosquito larvae. Journal of Aquatic Sciences 2001;16(2):111-4.##Hofrichter M. Industrial applications of lemon. Chemistry and Biochemistry 2010;6:224.##Yang P, Ma Y, Zheng S. Adulticidal activity of five essential oils against Culex pipiens quinquefasciatus. Journal of Pesticide Science. 2005;30(2):84-9.##El‐Adawy TA, Rahma EH, El‐Bedawy AA, Gafar AM. Properties of some citrus seeds. Part 1. Physico‐chemical characteristics of proteins. Food/Nahrung. 1999 Dec 1;43(6):374-8.##Sharma K, Mahato N, Cho MH, Lee YR. Converting citrus wastes into value-added products: Economic and environmently friendly approaches. Nutrition 2017 Feb;34:29-46. ##Yilmaz E, Güneşer BA. Cold pressed versus solvent extracted lemon (Citrus limon L.) seed oils: yield and properties. J Food Sci Technol 2017 Jun;54(7):1891-1900.##Penniston KL, Nakada SY, Holmes RP, Assimos DG. Quantitative assessment of citric acid in lemon juice, lime juice, and commercially-available fruit juice products. J Endourol 2008 Mar;22(3):567-70.##Anwar F, Naseer R, Bhanger MI, Ashraf S, Talpur FN, Aladedunye FA. Physico‐chemical characteristics of citrus seeds and seed oils from Pakistan. Journal of the American Oil Chemists' Society 2008 Apr;85(4):321-30.##Reda SY, Leal ES, Batista EA, Barana AC, Schnitzel E, Carneiro PI. Characterization of rangpur lime (Citrus limonia Osbeck) and" sicilian" lemon (Citrus limon) seed oils, an agroindustrial waste. Food Science and Technology. 2005;25:672-6.##Zhang Y, Liu C, Zhang Z, Qi Y, Wu G, Li S. Solvent gradient elution for comprehensive separation of constituents with wide range of polarity in Apocynum venetum leaves by high-speed counter-current chromatography. J Sep Sci 2010 Sep;33(17-18):2743-8.##Dua VK, Gupta NC, Pandey AC, Sharma VP. Repellency of Lantana camara (Verbenaceae) flowers against Aedes mosquitoes. J Am Mosq Control Assoc 1996 Sep;12(3 Pt 1):406-8.##Murugan K, Vahitha R, Baruah I, Das SC. Integration of botanical and microbial pesticides for the control of filarial vector, Culex quinquefasciatus Say (Diptera: Culicidae).##Venkatachalam MR, Jebanesan A. Repellent activity of Ferronia elephantum Corr. (Rutaceae) leaf extract against Aedes aegypti (L.). Bioresour Technol 2001 Feb;76(3):287-8.##Yoon JK, Kim KC, Cho YD, Cho HS, Lee YW, Choi BK, et al. Development and evaluation of a semifield test for repellent efficacy testing. J Med Entomol 2014 Jan;51(1):182-8.##Mann A, Ibrahim K, Oyewale AO, Amupitan JO, Fatope MO, Okogun JI. Brine shrimp toxicity evaluation of the root bark extract of Terminalia avicennioides. Afr J Sci Res 2011;4(1):221-8.##Aboaba SA, Aiyelaagbe OO, Ekundayo O. Chemical composition, toxicity and larvicidal activity of the essential oil from the whole plant of Acalypha segetalis from south-west Nigeria. Nat Prod Commun 2010 Mar;5(3):481-3.##Mbachu KA, Ibok MG, Adeniyi-Akee MA, Ajala OE. Chemical compositions and antioxidant activity of leaf and stem essential oils of Bryophyllum pinnatum (lam.) Kurz. GSC Biological and Pharmaceutical Sciences 2019 Nov 30;9(2):057-64.##Adams RP, 2007. Identification of Essential Oil Components by GC-MS. Allured Publication Corp., Carlo Stream, II. USA.##Zen S, Sutanto A, Rohmah A. The Effect of Lemon (Citrus Limon L.) Extract On Mortality Of Aedes Sp. and Potential As Repellent. PENBIOS: JURNAL PENDIDIKAN BIOLOGI DAN SAINS 2020 Nov 12;5(02):39-45.##Hue PT, Van Loc T, Vien LT, Hieu HV, Khoa PT. Optimization of citrus limon peel extraction, determination of main chemical components and effectiveness in repellency against aedes mosquito dengue fever vector. Vietnam Journal of Biotechnology 2022 Jun 30;20(2):329-38.##Girgenti P, Suss L. Attività repellente nei confronti di Aedes aegypti (L.) di formulati a base di estratti vegetali naturali o principi attivi di sintesi [Repellent activity against Aedes aegypti (L.) of formulas based on natural vegetable extracts or synthetic active agents]. Ann Ig. 2002 May-Jun;14(3):205-10. Italian.##Effiom OE, Avoaja DA, Ohaeri CC. Mosquito repellent activity of phytochemical extracts from peels of citrus fruit species. Global Journal of Science Frontier Research. 2012;12(1):1-8.##Jacobson M. Plant derived Insect Deterents glossary. CRC Press, Inc., Boca Raton, Florida; (1990).##Foster S, Duke JA. Naptalactose, a mild sedative compound in Catnip, also possesses herbicidal and insect-repellent properties. Peterson Field Guides, Eastern/Central Medicinal plants. Houghton Mifflin Co. New York. 1990.##Aboaba SA, Oladosu IA, Ekundayo O. Chemical composition, acetyl cholinesterase inhibitory properties and brine shrimp toxicity of essential oils from Ficus capensis Thunb from Nigeria. International Journal of Biological and Chemical Sciences 2010;4(5).##Eliwa AA, Mubark AE, Dakroury GA, El-Shazly EA, El-Azony KM. Polyacryl-dimethyl-heptadecanamine-mullite as a promising sorbent for chromium and vanadium sorption from ilmenite. Journal of Environmental Chemical Engineering 2022 Dec 1;10(6):108886.##Li B, Zang S, Zhou H, An S, Qiu H. Study on the Removal of ROX from Wastewater by Modified Schwertmannite with Hexadecyl Trimethyl Ammonia Bromide. Russian Journal of Physical Chemistry A 2023 Apr;97(4):672-8.##Jager P, Rentzea CN, Kieczka H. Carbamates and carbamoyl chloride. Ullmann’s Encyclopedia of industrial Chemistry Weinheim: Wiley-VCH. 2000.##Elmquist DC, Landolt PJ, Cooper WR, Reed H, Foutz J, Clepper T, et al. The Venom Compound N-(3-methylbutyl)acetamide Attracts Several Polistes (Fuscopolistes) Species (Hymenoptera: Vespidae). J Econ Entomol 2020 Jun 6;113(3):1073-1079.##Lopez-Avila V, Zorio M. Identification of methylhexaneamine by GC high-resolution TOFMS and soft ionization. Forensic Sci Int 2013 Sep 10;231(1-3):113-9.##Eliason MJ, Eichner A, Cancio A, Bestervelt L, Adams BD, Deuster PA. Case reports: Death of active duty soldiers following ingestion of dietary supplements containing 1,3-dimethylamylamine (DMAA). Mil Med 2012 Dec;177(12):1455-9.##Catalani V, Prilutskaya M, Al-Imam A, Marrinan S, Elgharably Y, Zloh M, et al. Octodrine: New Questions and Challenges in Sport Supplements. Brain Sci 2018 Feb 20;8(2):34.##

Isolation and Preliminary Characterization of a Novel scFv against SARS-CoV-2: an Experimental and Computational Analysis 2 2 Background: Since the initial outbreak, the SARS-CoV-2 virus has continued to circulate and mutate, resulting in the emergence of new viral sublineages. Due to the lack of effective protection and therapeutic measures against these new variants, the virus is able to further evolve and diversify. This study aimed to screen a phage antibody library to identify monoclonal antibodies in single-chain variable fragment (scFv) format that target the Receptor Binding Domain (RBD) of different SARS-CoV-2 strains. The newly discovered scFv has the potential for use as a diagnostic or therapeutic option against SARS-CoV-2. Methods: The RBD protein was produced, purified, and used as an antigen during biopanning. Six rounds of panning enriched RBD-specific phages and the binding affinity of binders were monitored by polyclonal phage ELISA. Subsequently, monoclonal phage ELISA was employed to identify specific binders. After sequence confirmation, the reactivity of the isolated anti-RBD scFv was evaluated. Additionally, bioinformatics tools determined the interaction between selected scFv and SARS-CoV-2 strains. Results: The ELISA analysis demonstrated that the expressed RBD retains its structural integrity and effectively interacts with antibodies present in the sera of COVID-19 patients. Through screening a phage display library, a strong-binding scFv for RBD was discovered, which can effectively neutralize SARS-CoV-2 and its novel variants. Conclusion: The findings of this study have led to the discovery of a novel scFv that effectively neutralizes SARS-CoV-2 strains, offering immense potential for research and therapy purposes. 64 79 Samaneh Jahandar-Lashaki Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences Iran Safar Farajnia Biotechnology Research Center, Tabriz University of Medical Sciences Biotechnology Research Center, Tabriz University of Medical Sciences Iran Effat Alizadeh Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences Department of Medical Biotechnology, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences Iran Farzin Seirafi Biotechnology Research Center, Tabriz University of Medical Sciences Biotechnology Research Center, Tabriz University of Medical Sciences Iran Asghar Tanoumand Department of Microbiology, Maragheh University of Medical Sciences, Maragheh Department of Microbiology, Maragheh University of Medical Sciences, Maragheh Iran Mohammad Kazem Hosseini Faculty of Sciences, Molecular Biology and Genetic, Istanbul University Faculty of Sciences, Molecular Biology and Genetic, Istanbul University Turkey BioprospectingCOVID-19Phage display libraryReceptor binding domainSingle-chain antibodies 60604.pdf Zhou H, Ni WJ, Huang W, Wang Z, Cai M, Sun YC. Advances in Pathogenesis, Progression, Potential Targets and Targeted Therapeutic Strategies in SARS-CoV-2-Induced COVID-19. Front Immunol 2022;13:834942.##WHO, COVID-19 epidemiological update, https://www.who.int/publications/m/item/covid-19-epidemiological-update-edition-166 (accessed 12 April 2024) 2024.##Yuan Y, Jiao B, Qu L, Yang D, Liu R. The development of COVID-19 treatment. Front Immunol 2023;14:1125246.##Planas D, Staropoli I, Michel V, Lemoine F, Donati F, Prot M, et al. Distinct evolution of SARS-CoV-2 Omicron XBB and BA.2.86/JN.1 lineages combining increased fitness and antibody evasion. Nat Commun 2024 Mar 13;15(1):2254.##Brant AC, Tian W, Majerciak V, Yang W, Zheng ZM. SARS-CoV-2: from its discovery to genome structure, transcription, and replication. Cell Biosci 2021;11(1):136.##Takeda M. Proteolytic activation of SARS-CoV-2 spike protein. Microbiol Immunol 2022;66(1):15-23.##Naqvi AAT, Fatima K, Mohammad T, Fatima U, Singh IK, Singh A, et al. Insights into SARS-CoV-2 genome, structure, evolution, pathogenesis and therapies: Structural genomics approach. Biochim Biophys Acta Mol Basis Dis 2020;1866(10):165878.##Shahin K, Zhang L, Mehraban MH, Collard JM, Hedayatkhah A, Mansoorianfar M, et al. Clinical and experimental bacteriophage studies: Recommendations for possible approaches for standing against SARS-CoV-2. Microb Pathog 2022;164:105442.##Niknam Z, Jafari A, Golchin A, Danesh Pouya F, Nemati M, Rezaei-Tavirani M, et al. Potential therapeutic options for COVID-19: an update on current evidence. Eur J Med Res 2022;27(1):6.##Salian VS, Wright JA, Vedell PT, Nair S, Li C, Kandimalla M, et al. COVID-19 Transmission, Current Treatment, and Future Therapeutic Strategies. Mol Pharm 2021;18(3):754-71.##Hwang YC, Lu RM, Su SC, Chiang PY, Ko SH, Ke FY, et al. Monoclonal antibodies for COVID-19 therapy and SARS-CoV-2 detection. J Biomed Sci 2022;29(1):1.##Iversen PL, Kane CD, Zeng X, Panchal RG, Warren TK, Radoshitzky SR, et al. Recent successes in therapeutics for Ebola virus disease: no time for complacency. Lancet Infect Dis 2020;20(9):e231-e7.##Taylor PC, Adams AC, Hufford MM, de la Torre I, Winthrop K, Gottlieb RL. Neutralizing monoclonal antibodies for treatment of COVID-19. Nat Rev Immunol 2021;21(6):382-93.##Antoine D, Mohammadi M, McDermott CE, Walsh E, Johnson PA, Wawrousek KE, et al. Isolation of SARS-CoV-2-blocking recombinant antibody fragments and characterisation of their binding to variant spike proteins. Frontiers in Nanotechnology 2022;4:1028186.##Planas D, Veyer D, Baidaliuk A, Staropoli I, Guivel-Benhassine F, Rajah MM, et al. Reduced sensitivity of SARS-CoV-2 variant Delta to antibody neutralization. Nature 2021;596(7871):276-80.##Dong Y, Meng F, Wang Z, Yu T, Chen A, Xu S, et al. Construction and application of a human scFv phage display library based on Cre‑LoxP recombination for anti‑PCSK9 antibody selection. Int J Mol Med 2021;47(2):708-18.##Liu ZX, Yi GH, Qi YP, Liu YL, Yan JP, Qian J, et al. Identification of single-chain antibody fragments specific against SARS-associated coronavirus from phage-displayed antibody library. Biochem Biophys Res Commun 2005;329(2):437-44.##Nagano K, Tsutsumi Y. Phage Display Technology as a Powerful Platform for Antibody Drug Discovery. Viruses 2021;13(2).##Ahmad ZA, Yeap SK, Ali AM, Ho WY, Alitheen NB, Hamid M. scFv antibody: principles and clinical application. Clin Dev Immunol 2012;2012:980250.##Ma Z, Zhu M, Zhang S, Qian K, Wang C, Fu W, et al. Therapeutic antibodies under development for SARS-CoV-2. View (Beijing) 2022;3(2):20200178.##Shadman Z, Farajnia S, Pazhang M, Tohidkia M, Rahbarnia L, Najavand S, et al. Isolation and characterizations of a novel recombinant scFv antibody against exotoxin A of Pseudomonas aeruginosa. BMC Infect Dis 2021;21(1):300.##Rahbarnia L, Farajnia S, Babaei H, Majidi J, Dariushnejad H, Hosseini MK. Isolation and characterization of a novel human scFv inhibiting EGFR vIII expressing cancers. Immunol Lett 2016;180:31-8.##Chang AY, Chau V, Landas JA, Pang Y. Preparation of calcium competent Escherichia coli and heat-shock transformation. JEMI Methods 2017;1(22-25).##Campisi M, Kobe DH. Derivation of the Boltzmann principle. American Journal of Physics 2010 Jun 1;78(6):608-15.##Brenke R, Hall DR, Chuang GY, Comeau SR, Bohnuud T, Beglov D, et al. Application of asymmetric statistical potentials to antibody-protein docking. Bioinformatics 2012;28(20):2608-14.##Jian F, Feng L, Yang S, Yu Y, Wang L, Song W, et al. Convergent evolution of SARS-CoV-2 XBB lineages on receptor-binding domain 455-456 synergistically en-hances antibody evasion and ACE2 binding. PLoS Pathog. 2023;19(12):e1011868.##Nutalai R, Zhou D, Tuekprakhon A, Ginn HM, Supasa P, Liu C, et al. Potent cross-reactive antibodies following Omicron breakthrough in vaccinees. Cell 2022;185(12):2116-31.e18.##Han P, Su C, Zhang Y, Bai C, Zheng A, Qiao C, et al. Molecular insights into receptor binding of recent emerging SARS-CoV-2 variants. Nat Commun 2021;12(1):6103.##Starr TN, Greaney AJ, Hannon WW, Loes AN, Hauser K, Dillen JR, et al. Shifting mutational constraints in the SARS-CoV-2 receptor-binding domain during viral evolution. Science 2022;377(6604):420-4.##Dijokaite-Guraliuc A, Das R, Zhou D, Ginn HM, Liu C, Duyvesteyn HME, et al. Rapid escape of new SARS-CoV-2 Omicron variants from BA.2-directed antibody responses. Cell Rep 2023;42(4):112271.##McCallum M, Czudnochowski N, Rosen LE, Zepeda SK, Bowen JE, Walls AC, et al. Structural basis of SARS-CoV-2 Omicron immune evasion and receptor engagement. Science 2022;375(6583):864-8.##Du S, Cao Y, Zhu Q, Yu P, Qi F, Wang G, et al. Structurally Resolved SARS-CoV-2 Antibody Shows High Efficacy in Severely Infected Hamsters and Provides a Potent Cocktail Pairing Strategy. Cell 2020;183(4):1013-23.e13.##Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC, et al. UCSF Chimera--a visualization system for exploratory research and analysis. J Comput Chem 2004;25(13):1605-12.##Desoubeaux G, Pelegrin M. [Monoclonal antibodies in infectious diseases: new partners in the therapeutic arsenal]. Med Sci (Paris) 2019;35(12):1008-13.##Shukra AM, Sridevi NV, Dev C, Kapil M. Production of recombinant antibodies using bacteriophages. Eur J Microbiol Immunol (Bp) 2014;4(2):91-8.##Otsubo R, Yasui T. Monoclonal antibody therapeutics for infectious diseases: Beyond normal human immunoglobulin. Pharmacol Ther 2022;240:108233.##Shim H. Antibody Phage Display. Adv Exp Med Biol 2017;1053:21-34.##Du L, Yang Y, Zhang X. Neutralizing antibodies for the prevention and treatment of COVID-19. Cell Mol Immunol 2021;18(10):2293-306.##Verderese JP, Stepanova M, Lam B, Racila A, Kolacevski A, Allen D, et al. Neutralizing Monoclonal Antibody Treatment Reduces Hospitalization for Mild and Moderate Coronavirus Disease 2019 (COVID-19): A Real-World Experience. Clin Infect Dis 2022;74(6):1063-9.##Chen X, Li R, Pan Z, Qian C, Yang Y, You R, et al. Human monoclonal antibodies block the binding of SARS-CoV-2 spike protein to angiotensin converting enzyme 2 receptor. Cell Mol Immunol 2020;17(6):647-9.##Li F. Evidence for a common evolutionary origin of coronavirus spike protein receptor-binding subunits. J Virol 2012;86(5):2856-8.##Shilling PJ, Mirzadeh K, Cumming AJ, Widesheim M, Köck Z, Daley DO. Improved designs for pET expression plasmids increase protein production yield in Escherichia coli. Commun Biol 2020;3(1):214.##Rahbar Z, Nazarian S, Dorostkar R, Sotoodehnejadnematalahi F, Amani J. Recombinant expression of SARS-CoV-2 receptor binding domain (RBD) in Escherichia coli and its immunogenicity in mice. Iran J Basic Med Sci 2022;25(9):1110-6.##Gao X, Peng S, Mei S, Liang K, Khan MSI, Vong EG, et al. Expression and functional identification of recombinant SARS-CoV-2 receptor binding domain (RBD) from E. coli system. Prep Biochem Biotechnol 2022;52(3):318-24.##Leung KM, Feng DX, Lou J, Zhou Y, Fung KP, Waye MM, et al. Development of human single-chain antibodies against SARS-associated coronavirus. Intervirology 2008;51(3):173-81.##Tang XC, Agnihothram SS, Jiao Y, Stanhope J, Graham RL, Peterson EC, et al. Identification of human neutralizing antibodies against MERS-CoV and their role in virus adaptive evolution. Proc Natl Acad Sci USA 2014;111(19):E2018-26.##Jiang L, Wang N, Zuo T, Shi X, Poon KM, Wu Y, et al. Potent neutralization of MERS-CoV by human neutralizing monoclonal antibodies to the viral spike glycoprotein. Sci Transl Med 2014;6(234):234ra59.##Kim C, Ryu DK, Lee J, Kim YI, Seo JM, Kim YG, et al. A therapeutic neutralizing antibody targeting receptor binding domain of SARS-CoV-2 spike protein. Nat Commun 2021;12(1):288.##Li H, Zhu B, Li B, Chen L, Ning X, Dong H, et al. Isolation of a human SARS-CoV-2 neutralizing antibody from a synthetic phage library and its conversion to fluorescent biosensors. Sci Rep 2022;12(1):15496.##Parray HA, Chiranjivi AK, Asthana S, Yadav N, Shrivastava T, Mani S, et al. Identification of an anti-SARS-CoV-2 receptor-binding domain-directed human monoclonal antibody from a naïve semisynthetic library. J Biol Chem 2020;295(36):12814-21.##Shadman Z, Ghasemali S, Farajnia S, Mortazavi M, Biabangard A, Khalili S, et al. In silico Validation of Pseudomonas aeruginosa Exotoxin A Domain I Interaction with the Novel Human scFv Antibody. Infect Disord Drug Targets 2023;23(5):e290323215113.##Bandehpour M, Ahangarzadeh S, Yarian F, Lari A, Farnia P. In silico evaluation of the interactions among two selected single chain variable fragments (scFvs) and ESAT-6 antigen of Mycobacterium tuberculosis. Journal of Theoretical and Computational Chemistry 2017;16(08):1750069.##

Ethical Considerations Emerge from Artificial Intelligence (AI) in Biotechnology 2 2 The integration of Artificial intelligence (AI) in biotechnology presents significant ethical challenges that must be addressed to ensure responsible innovations. Key concerns include data privacy and security, as AI systems often handle sensitive genetic and health information, necessitating robust regulations to protect individuals' rights and maintain public trust. Algorithmic bias poses another critical issue; AI can reflect existing biases in training data, leading to inequitable healthcare outcomes. Transparency in AI decision-making is essential, as "black box" models hinder trust, especially in drug discovery and genetics. Ethical implications of genetic manipulation require careful scrutiny to define the limits of human intervention. Additionally, societal impacts must be considered to ensure equitable distribution of AI benefits, preventing the exacerbation of disparities. Engaging diverse stakeholders, including ethicists and policymakers, is vital in aligning these technologies with societal values. Ultimately, prioritizing ethics will allow us to harness AI and biotechnology's potential while safeguarding human rights and promoting equity. 80 81 Mahintaj Dara Negar Azarpira Transplant Research Center, Shiraz University of Medical Sciences Transplant Research Center, Shiraz University of Medical Sciences Iran Artificial intelligenceBiotechnologyEthics 60605.pdf Ashfi SA. Ethical Implications of Emerging Technologies: A Comparative Analysis of AI, Biotechnology, and Robotics. Journal of Research and Innovation in Technology and Management (JRITM)##Sheth D, Patel P, Pathak Y. Ethical Issues and Artificial Intelligence Technologies in Bioinformatics Concerning Behavioural and Mental Health Care. Ethical Issues in AI for Bioinformatics and Chemoinformatics: CRC Press; 2023. p. 72-86.##O'Brien JT, Nelson C. Assessing the risks posed by the convergence of artificial intelligence and biotechnology. Health Secur 2020;18(3):219-27.##Fu R, Huang Y, Singh PV. Ai and algorithmic bias: Source, detection, mitigation and implications. Detection, Mitigation and Implications (July 26, 2020). 2020.##Nagamarpalli RS. Biotechnology and AI. Issue 3 Int'l JL Mgmt & Human. 2021;4:4035.##Almeida M, Diogo R. Human enhancement: Genetic engineering and evolution. Evol Med Public Health 2019;2019(1):183-9.##Anomaly J, Johnson T. The ethics of genetic enhancement: Key concepts and future prospects. Routledge Handbook of the Ethics of Human Enhancement: Taylor & Francis; 2024.##Guan J. Artificial intelligence in healthcare and medicine: promises, ethical challenges and governance. Chin Med Sci J 2019;34(2):76-83.##Holzinger A, Keiblinger K, Holub P, Zatloukal K, Müller H. AI for life: Trends in artificial intelligence for biotechnology. N Biotechnol 2023;74:16-24.##Al-Delaimy A. Perspective of ethical issues and challenges of biotechnology in developing country. Malaysian Journal of Public Health Medicine 2022;22(1):242-51.##Ahmad SF, Han H, Alam MM, Rehmat M, Irshad M, Arraño-Muñoz M, et al. Impact of artificial intelligence on human loss in decision making, laziness and safety in education. Humanit Soc Sci Commun 2023;10(1):1-14.##Halina M. Insightful artificial intelligence. Mind & Language 2021;36(2):315-29.##Sakai H, Matsuda K, Kikkawa N, Kajita S. Human divergent exploration capacity for material design: A comparison with artificial intelligence. Computers in Human Behavior: Artificial Humans 2024;2(1):100064.##