COMPARATIVE INFLUENCE RADIATION SCALE ON PHYSICAL AND CHEMICAL PROPERTIES OF VENOM OF SNAKE VIPERA LEBETINA OBTUSA Sh.A.TOPCHIYEVA, H.A. ABIYEV, E.M.BABAYEV Azerbaijan National Academy of Sciences Instutute of Zoology Az1073 Baku, Pass.1128, block 504 AZERBAIJAN REPUBLIC Shtopchiyeva@mail.ru Abstract: -The purpose of research studying of influence of small dozes scale of radiation on spectral characteristics and stability of venom vipera. Results of chemical analyses shown, that in structure of venom (Vipera lebetina ) is found out significant concentration of heavy metals (Cd, Ni, Zn, Cr, Pb). As a result of cooling samples up to nitric temperature (77К) appears new narrow maxima at 440 and 470 nm which is connected by metals taking place in structure of venom of Vipera lebetina obtusa. As a result of the lead experimental researches it is established, that under influence of small dozes γ- radiations at dozes 2.7, 4.05, 5.4 kgy within 1.0, 1.5 and 2.0 hours results in gradual decrease of toxicity and it is possible pharmacological activity of enzymes of snake venom. Key-Words: venom, snake, Vipera lebetina obtusa, γ-radiations, heavy metals, atom-absorbtion spectrophotometer, ecology. 1 Introduction The problem of pollution of biosphere of Apsheron of Azerbaijan by toxic, heavy metals like Pb, Hg, As, Cd, Ni, Cu, Va, Zn, Co, Mo, Sr and other metals, has arisen with emission in an atmosphere of technogens emissions of the industrial enterprises. They are the most dangerous toxic elements for fauna and animals and the people by entering in the basic biological circulation of substances. Studying of the given problem is the actual problem, having important value for medicine, biology and ecology. Snake venoms are unique on a chemical compound and physiological action group of biologically active connections of a natural origin. In territory of Azerbaijan from 23 kinds of snakes 4 kinds venomous. Insignificant degree of a level of scrutiny of venom Transcaucasian viper Vipera lebetina obtusa as representative of fauna of Azerbaijan being the large supplier of this invaluable medicinal raw material for a farmaceutical industry has served as the precondition of its research. In the literature there are not numerous and rather inconsistent data on dependence of properties zootoxins from various ecological factors (biotics and abiotics). On the basis of literary given and our experimental researches it is revealed, that poison гюрз contains ions of metals of Cd, Cu, Ni, Hg, Zn and Cr quantities which varies depending on places of dwelling of snakes [1, 2, 3, 4]. The purpose of research was studying influence of ecological factors (biotics, abiotics), including pollution of biosphere texnogens emissions of the industrial enterprises on elementary structure of venom Transcaucasian vipers. The method atom-absorbtion spectrophotometric (AAS-300, Perkin-Elmer) in venom viper, taking place in a serpentarium in rather non-polluting conditions and in venom of the snakes caught from ecologically polluted sites Apsheron, determines the maintenance of heavy metals for a dirtyng. [5,6]. Essential fluctuations in quantitative and a chemical compound of poison of the snakes caught from various sites of Azerbaijan, and in venom of the snakes who are taking place in a serpentarium are established. Maintenances of heavy metals in structure of biologically active polymer (what snake venom is) - zootoxin, which in the polluted zones fluctuation in limits for Cr, Pb, Cd, Zn, Ni 29-101.1, 7.0-134.8, 1.8-24.2, 23.0, 600.89-863.6 mg / kg accordingly are revealed. For venom of the snakes who are taking place in a serpentarium, quantity indicators make for Pb, Ni, Cd, Zn-15.12, 0.28, 0.31, 180.0 mg / kg, accordingly. Thus, essential quantitative fluctuations of a chemical ISBN: 978-1-61804-001-5 45
compound of poison of snakes taking place in bondage and on the nature in the various ecologically polluted sites of Apsheron of Azerbaijan are established. Essential changes physical and chemical, pharmacological and toxicological properties of biologically active biopolymer are revealed depending on conditions of their dwelling, from biotics and abiotics factors [5,6]. From references it is known, that water solutions of venoms are unstable and lose toxicity in some day. Snake poisons in the physiological solution containing 50 % of glycerin, on given to Poguda A.A. [7], at storage in a refrigerator within 6 months have not lowered toxicity. More proof to influence of factors of an environment there are venoms after drying (above chloride calcium) or liophylisation. According to Pavlovsk Е.Н [8], venom of a cobra at storage more than 20 years on a cold in the sealed - in ampoule is kept with toxicity. Snake venoms termostabilisation and in the sour environment maintain heating up to 1200 With without loss of activity [7]. Destroying chemical agents operate on venoms: permanganate kalium, chloroform, ethanol, methyl dark blue [5, 6, 8, 9]. Snake venoms inactivated also under action of some physical factors: a ultra-violet irradiation, X-rays. On data Salafranca [10], in 7 days after an irradiation of poison филипийской cobras Со60 in dozes 0.25, 0.5, 1 mrad its toxicity accordingly made 83, 66 and 43 % from not irradiated venom. Proceeding from above stated, studying of influence of small dozes the scale of radiation on spectral characteristics of poison and accordingly on toxicity and pharmacological activity zootoxini is represented rather actual.. 2 The purpose of research The purpose of research studying of influence of small dozes scale of radiation on spectral characteristics and stability of venom vipera. 2 Material and research method As material of research venom vipera, dried up in exicator above pairs chloride calcium and the venom irradiated with small dozes served γ-radiations. For experiment tests of dry venom, and also their water solutions have been taken. Water solutions of venom have been subjected γto radiation up to dozes 1.35, 2.7, 4.05, 5.4 kgy during 0.5, 1.0, 1.5, 2.0 hours. Simultaneously dry venom vipera was exposed γ-radiations in continuation continuation 2.0 hours a doze 5.4 kgy. In comparative aspect action scale of radiation on spectral characteristics of venom Caucasian viper Vipera lebetina obtusа was studied. Irradiation of venom vipera small dozes scale of radiation (γ-radiation D=0.75kG/sec) carried out on K-25 to isotope installation with application 60 Со. 3 Results of researches Ultra-violet spectra passing of venom have been removed on device Specord UV-VIS. On rice 1 it is resulted UV-VIS a spectrum of absorption of a water solution of venom vipera. Apparently from figure to 1 venom vipera characteristic maxima of absorption are peculiar in the field of 285 nm and 800 nm. Absorption of group in these molecules represents the big resounding structures with the big number of the connected communications Electrons, carrying out chemical communication are not located, but are the general for all structure. Therefore it is possible to speak about characteristic for zootoxins spectra of absorption. With the help of measurement of a spectrum of absorption of venom vipera, after an irradiation, it is possible to define places of attack and to establish chemical changes zootoxin, both in the irradiated solutions, and in a firm phase. On rice 2 spectra of absorption of samples of venom vipera, irradiated are resulted γradiation (water solutions and dry venom). Apparently from figure 2 at increase in a doze of an irradiation γ-radiation of solutions of venom vipera observes characteristic changes of optical density of researched samples of snake venom. The revealed reduction of intensity of absorption at 260 and 300 nm testifies to course of biochemical reactions in a firm phase of separate enzymes zootoxins ISBN: 978-1-61804-001-5 46
Recent Researches in Energy, Environment, Entrepreneurship, Innovation As a result of the lead experimental researches it is established, that under influence of small dozes γradiations at dozes 2.7, 4.05, 5.4 kgy within 1.0, 1.5 and 2.0 hours results in gradual decrease of toxicity and it is possible pharmacological activity of enzymes of snake venom. It is necessary to note, that at comparing intensity of absorption of control samples venom with samples zootoxin irradiated up to dozes 1.35 kgy it has not been revealed essential changes. Wave number in sm - 1 Fig. 1. UV-VIS a spectrum of absorption of venom vipera. It is necessary to note, that at сравнивании intensity of absorption of control samples (not irradiated) poison with samples zootoxin irradiated up to dozes 1.35 kgy it has not been revealed essential changes. From above-stated follows, that influence on venom vipera small dozes γ-radiation up to dozes 1.35 kgy within 0.5 hours does not result in reduction of intensity of absorption and according to toxicity, and including pharmacological activity. The subsequent increase in a doze scale of radiation up to 2.7, 4.05, 5.4 kgy within 1.0, 1.5, 2.0 hours promotes proportional reduction of intensity of absorption of snake venom and according to reduction, both toxicity, and pharmacological activity. At low temperatures researches and at revealing postrations effects in organic and biological systems of an organism from effective methods is radiotermoluminence. The method radiotermoluminence will allow to receive data on structural properties of system, on the centers of stabilization of charges of initial products radiolis venom, about ways of migration of energy absorbed at an irradiation and so on. Wave number in sm - 1 Fig. 2. Spectra of absorption of samples of venom vipera, irradiated γ-radiation (1,2,3,4-water solutions, 5-dry venom): 1-control; 2,3,4-irradiated (D =1.35, 2.7, 4.05 kgy), 5 (D=5.4kGy) The sample were excited a Nitrogen Laser diode emitting at 337.1 nm with a pulse width of 0.6 ns. The spectra were analyzed with a Jobin-Yvon HR 360 monochromator coupled with cooled CCD detector. The visible emitted light was collected from the same side. Luminescent properties at liquid N2 temperature measured using a cold finger cryostat under vacuum. Results of chemical analyses shown, that in structure of venom (Vipera lebetina obtusa) is found out significant concentration of heavy metals (Cd, Ni, Zn, Cr, Pb). In Fig. 2 and 3 emission spectra of Vipera lebetina obtusa under λexc= =337.1nm at room and 77 K temperature are present. It is visible, that PL specter of ISBN: 978-1-61804-001-5 47
venom covers the area 350-800 nm and consists of the wide maximum at 520 nm and poorly appreciable maxima covering wavelengths of 400-500 nm at 300K. As a result of cooling samples up to nitric temperature (77К) appears new narrow maxima at 440 and 470 nm which is connected by metals taking place in structure of venom of Vipera lebetina obtusa Fig.2. Photoluminescence spectra of venom at 300 0 K. Fig.3. Photoluminescence spectra of venom at 77 0 K Samples of venom were irradiated in special a ditch with scales - beams at 77К up to dozes of % 5 кgr. Before an irradiation samples were cleared of traces of oxygen. The irradiation was spent on air and in vacuum. Curves lighting registered with a speed ~ 5 0 /l.min. It is shown, that in an interval of temperatures 77-330К the curve lighting radiotermoluminence venom of viperas irradiated at 77К up to dozes 3 кgr is characterized not by a symmetric maximum at temperature 172К. Warming up up to temperatures 320К results in monotonous decrease of intensity of a luminescence. It is established, that intensity of peak at 172К increases depending on doze Д=0-15 кgr gamma irradiation under the linear law. The irradiation in air results in additional weak peak at 185К and to downturn of intensity at 172К. It is shown, that the peak of a luminescence at 172К can be dosimetric for an estimation of the absorbed doze. The opportunity of change of medicinal properties of venom of Viperas is discussed at influence of small dozes of scale - radiation. 4 Conclusion Thus, as a result of the lead researches it is revealed that under influence of small dozes γ-radiations up to dozes 2.7, 4.05, 5.4 kgy during 1.0, 1.5 and 2.0 hours results in reduction of intensity of absorption at 260 and 300 nm that testifies to course of biochemical reactions in a firm phase of separate enzymes zootoxin, and in turn results in decrease of toxicity and it is possible pharmacological activity of enzymes of snake venom. However influence on a solution of snake venom γ-radiations up to dozes 1.35 kgy within 30 minutes promotes stabilization as to toxicity, and pharmacological activity with simultaneous increase accordingly a period of storage of water solutions of snake venom that can be used at sterilization of water solutions of snake poison, and in turn is important for a farmaceutical industry by manufacture of injections on a basis zootoxins. Reference: [1] Sh.A.Topchiyeva.To the physics-chemical properties, application and some ecological aspects of changability of venom properties Journal "Knowledge", Baku, 2, 2000, pp. 48-52. [2] Sh.A.Topchiyeva et al. Influence of the ecological factors to number Vipera lebetina obtuse and chemical composition of venom. Proceedings of the First International Conference on Enviromental Research and Assessment, Bucharest, Romaniya, 2003, pp.34-37. [3] Sh.A.Topchiyeva, G.A.Abiyev. Ecological factor and venom Transcaucasian viper. Prospects of using of natural resources and economic development of southern region of Azerbaijan, Respublican conference, Lenkoran, 2004, pp.120-125 ISBN: 978-1-61804-001-5 48
[4] Sh.A.Topchiyeva, H.Ə.Abiyev. Comparative influence of impurity soils Apsheron of Azerbaijan on a chemical compound of venom Transcaucasian viper Vipera lebetina obtusa. J. Knowledge, Baku, 4, 2004, pp.74-78. [5] T.M. Iskenderov, Sh.A.Topchiyeva. The influence of ecologicfl factors on chemical composition of venom and adaptation of Caucasian viper to the captivity conditions. J. Knowledge Education Society of Azerbaijan Republic (Chemistry, Biology, Medicine), Baku, 4, 2003, pp.62-66. [6] Sh. A.Topchiyeva, T.M.Isgenderov, O.G.Mamedov. The Influence of biospher pollution on chemical content of the Vipera venom in Azerbaijan. J.Medical. Baku, Vol.1, 2003, pp. 219-223. [7] Pavlovskiy E.N. Poisonous the animal USSR, Moscow, 1990, 271 p. [8] Pokuda A.A.Stability of solutions of venoms of snakes vipera, halis, cobras at storage. Scientific reports of the higher school, biological sciences, 3, 1972, pp. 52-55. [9] Sh.A.Topchiyeva, A.M.Magerramov, N.N.Musayeva, L.Z. Allaxverdiyeva, R.Z.Allax-verdiyeva, Abiyev H.A. Influence of small dozes γ- radiations on molecular mobility and pharmacological properties of venom of Vipera Lebetina Obtusa, 40 th IUPOC International Symposium on Macromol, World Polymer Congress Macro, Juli 4-9, Paris-France, 2004, pp.53-120. [10] Salafranca E.S. Irradiated cobra (Naja naja philippinensis) venom. Int.J. Appl. Radiatand isotop., Vol.24, 1, 1973, pp.60. ISBN: 978-1-61804-001-5 49