Influence of Cu L-Histidinate Schiff base derivatives on structural features of irradiated rat’s DNA

Influence of Cu L-Histidinate Schiff base derivatives on structural features of irradiated rat’s DNA | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Influence of Cu L-Histidinate Schiff base derivatives on structural features of irradiated rat’s DNA Nelli H. Karapetyan, Samvel G. Haroutiunian, Gayane V. Ananyan This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4387030/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 12 You are reading this latest preprint version Abstract A study of rats liver DNA damages under the influence of X-ray radiation at a dose of 6.5 Gy(LD60) was carried out. The radioprotective properties of newly synthesized Cu(II) L-Schiff Histidinate complexes were also studied. The survival of rats was determined over a 30-day period after exposure to X-rays without pretreatment and also after preadministration of Cu(II) L-Histidinate-Schiff base complexes. The structural defects of rat's liver DNA were detected at 3, 7, 14, and 30 days post-irradiationxtracted. The results obtained revealed that irradiation with a 6.5Gy dose in the control group degraded the characteristics of rat liver DNA in comparison to healthy DNA. On all investigated experimental days, a decrease in the melting temperature (T m ), a widening of the melting interval (ΔT), and a decrease in hypochromicity (Δh) were observed in the DNA samples of irradiated animals compared to the norm. The rat's pretreatment by Cu(II) L-Histidinate complexes 1 or 24 hours prior to irradiation improved DNA characteristics. Electrophoretic studies of DNA were in good agreement with the melting data. Based on the study results, it can be concluded that Cu(II) L-Histidinate complexes exhibit radioprotective properties under the studied conditions and can protect DNA from damage. Irradiation DNA melting Electrophoresis Schiff base complexes. Figures Figure 1 Figure 2 Figure 3 Figure 4 INTRODUCTION Ionizing radiation, being high-energy, generated ions that can destroy the covalent bonds of molecules. It can cause various types of damage to DNA, RNA, proteins, and other biomolecules. Among these, DNA is the major target of radiation-induced damage within the cell. DNA molecule damage occurs either through direct or indirect ionization through the production of water radicals, which affect and damage DNA ( 1 ). In general, the sugar-phosphate backbone lesions generate DNA single-strand breaks ( 2 , 3 ). If damage occurs on opposite strands of DNA within the same helical turn, then double-strand DNA breaks are formed. The radiolysis of water results the formation of reactive oxygen species like superoxide radicals, hydrogen peroxide, singlet oxygen, and highly reactive hydroxyl radicals ( 3 ). As a result, these species oxidize proteins and lipids, and also induce various types of damage to DNA, including modifications in purine and pyrimidine bases, sugar damage, and apurinic or apyrimidinic sites damage, removal of bases, cross-linking of DNA with DNA or adjacent proteins ( 4 ). These changes induce cell death and mitotic failure ( 5 ). The extent of radiation damage to the body depends on cellular antioxidant statuses, such as antioxidant compounds and enzyme systems (e.g. superoxide dismutase, catalase, peroxidases) that reduce intracellular reactive oxygen species levels. Currently, a special area of radiobiology is the search for means of chemical protection against radiation. These substances block the development of chain radiation-chemical reactions by intercepting active radicals, antioxidants, and agents that create tissue hypoxia, reduce the intensity of oxidative processes by binding metal ions with variable valence, and catalyze oxygen transfer. The protection of sulfhydryl groups of proteins by stimulating cellular recovery systems has been demonstrated ( 6 ). The possibility of introducing agents required for DNA repair is being studied ( 7 , 8 ). The purpose of this study was to identify DNA damage in the liver of X-ray irradiated rats and DNA repair in rats pretreated with Cu(II) L-Histidinate Schiff bases complexes before irradiation. In this study, various exposure schemes were used to identify the most optimal one. Promising results were obtained in the study of the radioprotective activity of the Cu(II) and Mn(II) complexes with Schiff bases derived from isomeric pyridincarboxaldehydes with tryptophan and tyrosine cyclic amino acids ( 9 , 10 ). Metals, such as copper, cobalt, iron, manganese, and zinc are elements required by all cells for normal metabolic processes. Ionic forms of these metals have particularly high affinities for organic ligands found in biological systems and rapidly undergo bonding interactions to form complexes, or more precisely, coordinate-covalent bonded chelate structures (11). Chelated forms of metals exist in tissues in the form of metal-dependent enzymes, proteins, and various low molecular-weight peptide chelates. Transition metal ions can coordinate the ligand in a precise 3D configuration, thus adapting the molecule to recognize and interact with the target molecule. The synthesis of metal element-dependent enzymes, required for oxygen utilization and superoxide accumulation prevention, as well as tissue repair processes, including metal element-dependent DNA and RNA repair, is key to the hypothesis that small molecular mass essential metal chelates such as Schiff bases facilitate recovery from radiation-induced pathology. MATERIALS AND METHODS All used chemicals and reagents of Analytical Reagent grade were obtained from Sigma-Aldrich. Cu(II) Schiff Base complexes with L-Histidine and 2-, 3- and 4-pyridinecarboxaldehydes were used for animal studies (Fig. 1 ). These new complexes were synthesized and kindly presented by V. Matosyan and V. Tonoyan at the Scientific Centre of Radiation Medicine and Burns (Yerevan, Armenia) ( 12 ). The toxicity of synthesized Schiff Bases was characterized based on a calculation of the value of LD 50 , which is the dose of a compound at which the lethality of 50% of animals was observed in 7 days after subcutaneous administration of the compound to the organism. With the use of the Behrens integration method in experiments in mice, the toxicity of compounds was calculated ( 12 ). According to the test results, Cu(II) complexes with Schiff Bases proved to be low toxic. Male albino rats of the Wistar strain with 180-200g were used for experimental studies. The rats were bred at the vivarium of the Scientific Centre of Radiation Medicine and Burns (Yerevan, Armenia) under conventional laboratory conditions. To undergo whole-body irrigation X-ray, 10 rats were placed in a well-ventilated box, located 50cm from the source of radiation. The RUM-17 (RF) therapeutic X-ray was used for irradiation. The radiation conditions were the following: voltage 180kV, current 15mA, without filter, dose rate 1.78Gy/min. The studies were carried out under various schemes of exposure in order to choose the most optimal of them. The design of the experiment is as follows: X-ray exposure of white rats at 6.5Gy (LD 60 ). Treatments: 1 or 24 hours prior to irradiation; Drug administration mode: subcutaneous and oral; Dose levels: 10 mg/kg or 40 mg/kg. This article presents experimental data for rats administrated subcutaneously by investigating compounds 1 hour prior to irradiation at a dose of 6.5Gy. The rats’ liver DNA was isolated from the following 6 groups of animals: I group: intact animals served as control II group: rats obtained 10mg/kg Cu(II)(Picolinyl-L-Histidinate) 2 , Cu(II)(Nicotinyl-L- Histidinate) 2 and Cu(II)(Isonicotinyl-L-Histidinate) 2 (the pure effect) on 1st and 30th days. III group: animals exposed to X-ray irradiation at 6.5Gy dose level (Irradiated control group) IV group: rats pre-treated subcutaneously administration of 10mg/kg Cu(II)(Picolinyl-L-Histidinate) 2 1 hour before exposure to X-ray irradiation at 6.5Gy dose level V group: rats obtained 10mg/kg Cu(II)(Nicotinyl-L-Histidinate) 2 subcutaneously 1 hour prior to irradiation at 6.5Gy VI group: rats obtained 10mg/kg Cu(II)(Isonicotinyl-L-Histidinate) 2 subcutaneously 1 hour prior to irradiation at 6.5Gy Cu(II) complexes used in this study were administrated to the rats in the form of suspension using deionizing water as solvent. The animals were injected subcutaneously with 2mg Cu(II) chelates in 0.5ml solution on 200mg body weight 1 hour before irradiation. In order to perform analyses, rats were sacrificed under anesthesia on 3, 7, 14, and 30 days after irradiation (per 5 rats at each point). Data are presented as the average of three measurements. DNA isolation. DNA was isolated from animal's liver using the standard chloroform method ( 9 ). The absorbance of DNA solution at 230 nm, 260 nm and 280 nm was used to evaluate DNA purity. The DNA samples had А 260 /А 280 = 1.8 and А 260 /А 230 = 2 and were free from contaminations. Spectroscopy. The absorption spectra and ultraviolet melting curves of DNA samples were recorded on a Lambda 800 UV/VIS spectrometer (Perkin-Elmer). DNA concentrations were determined spectrophotometrically with ε 260nm = 1.31x10 4 M − 1 cm − 1 and calculated in base pairs ( 13 ). All spectroscopic measurements were performed in biphosphate buffer solution (0.6mM Na 2 HPO 4 , 0.2mM NaH 2 PO 4 , 18.5mM NaCl, 0.01mM EDTA), [Na + ] = 0.02, pH 7.2. \(\Delta h=({A_{{{95}^o}C}}/{A_{{{25}^o}C}} - 1) \cdot 100\%\) Melting experiments were carried out at a 35-95 o C temperature interval, using 10mm thermostatic quartz cuvettes. The heating rate was 0.5 o C/min, while absorbance at 260 nm was recorded. Melting characteristics of DNA - the melting temperature (T m ), the width of the melting interval ΔT=( ∂θ/∂T ) −1 T=Tm and hypochromic effect were calculated from melting curves (14). T m is the temperature at which half of all base pairs “melted”, i.e. 1-θ = 0.5, and ΔT is the width of the melting interval, equal to the temperature difference at which the tangent at the inflection point crosses the levels θ = 0 and θ = 1 ( 15 ). Electrophoresis. DNA electrophoresis was performed using 1% agarose gel in Tris-boric acid - EDTA buffer, pH 8.0, under condition of 5 V/cm. The 1kb DNA Ladder (Promega Product, USA) was used as a marker for determining the size of double-stranded DNA from 250 − 10 000 base pairs ( 16 ). The gel was stained with 0.5 ng/ml ethidium bromide solution, viewed and photographed under UV-trans illuminator. RESULTS The effectiveness of the studied Cu(II) L-Histidinate Schiff base compounds, as radioprotectors was assessed by the survival of animals for 30 days (Table 1). On the 30th day of the experiment, the survival rate of rats for all investigated Cu(II) L-Histidinate Schiff base compounds was 100%. When rats were irradiated with a dose of 6.5Gy, their survival rate on 30 day was 40%. However, pre-treatment with Cu(II)L-Histidinate complexes before irradiation increased the survival rate of animals by 1.5-2.5 times. All studied Schiff base complexes exhibited a pronounced radioprotective effect in all cases of use. However, the effectiveness of the compounds depends on the scheme of their administration. According to our results (Fig. 2 and Fig. 3), the most effective radioprotective effect was observed with subcutaneous administration of drugs. Good results (80-100% survival) were obtained when 10 mg/kg Cu(II) complexes were administered to rats 1 hour before irradiation. At oral use, a good radioprotective effect (75-100% survival) was observed in the case of administration at 40 mg/kg dose 24 hours prior to irradiation. Probably this can be explained by the fact that, upon oral administration, the complexes are absorbed into the blood more slowly and effectiveness of compounds was observed later. Administration of Cu(II)(Picolinyl-L-Histidinate) 2 exhibited 80% survival by subcutaneous administration at 10mg/kg dose 1 hour before irradiation, and 100% survival by oral administration at 40mg/kg dose 24 hours prior to exposure. Table 1. Survival of animals 30 days after irradiation exposure to X-rays at 6.5Gy on the background of subcutaneous or oral administration of 10 mg/kg and 40 mg/kg Cu(II)(Picolinyl-L-Histidinate) 2 , Cu(II)(Nicotinyl-L-Histidinate) 2 ,and Cu(II)(Isonicotinyl-L-Histidinate) 2 1or 24 hours prior to irradiation. At all experiments Norm (intact) rats survival was 100%, Irradiated rats Control survival was 40 %. p ≤ 0.02 Substance Mode Dose, mg/kg Survival,% 6.5Gy 1h prior 24h prior Cu(II)(Picolinyl-L-Histidinate) 2 subcutaneous 10 80 60 40 75 60 oral 10 80 90 40 70 100 Cu(II)(Nicotinyl-L-Histidinate) 2 subcutaneous 10 90 70 40 70 60 oral 10 80 80 40 50 85 Cu(II)( Isonicotinyl-L-Histidinate) 2 subcutaneous 10 100 60 40 90 60 oral 10 75 75 40 70 80 Pretreatment with 10mg/kg Cu(II)(Nicotinyl-L-Histidinate) 2 subcutaneously 1 hour before irradiation resulted in a 90% survival rate. In the case of oral administration at a 40mg/kg dose 24 hours prior to irradiation, an 85% survival rate was observed. Rats administered with Cu(II)(Isonicotinyl-L-Histidinate) 2 exhibited 100% survival when given a subcutaneous dose of 10mg/kg one hour before irradiation. With oral administration at a 40mg/kg dose 24 hours prior to irradiation, 80% survival was observed at 6.5Gy irradiation dose. The main cause of death in animals at radiation is damages of DNA, which is very sensitive to ionizing irradiation. The next stage of research was the identification of structural features of DNA during irradiation. For this purpose DNA was isolated from the liver of rats on 3, 7, 14 and 30 days after irradiations. Structural changes in DNA also were detected in animals pre-treated subcutaneously with Cu(II) L-Histidinate Schiff bases. Damage to the DNA molecule was assessed by the melting method. The melting temperature (Т m ), melting interval (DТ) and hypochromic effect (Δh) provide information on DNA stability and defects in the DNA secondary structure. The pure effect of Cu(II)L-Histidinate complexes on the stability of rats liver DNA was examined on 1st and 30th days after administration of the compounds. The structural features of DNA did not change when the compounds was administered to rats. The DNA melting parameters were the same as those of normal DNA. Since Cu(II)L-Histidinate complexes do not affect DNA melting parameters on both 1st and 30th days, the stability of the DNA molecule is preserved (Table 2). The obtained results showed that irradiation with a dose of 6.5Gy of the control group of animals promote deviation of the characteristics of the rat's liver DNA from the norm. The DNA melting parameters of irradiated rats on days 3, 7, 14 and 30 of the experiment differed from the norm. In particular a decrease in T m and Δh, an increase DТ were observed. On 30 days after irradiation the DNA of the irradiated rat's destabilized (T m decreased from 71.2 o C to 62.2 o C), the hypochromicity decreased (from 33% to 14%), and the ΔT increased from 7.3 o C to 27.1 o C, in comparison to normal DNA. These changes in the melting parameters indicate the destabilization of the DNA molecule, and the presence of partially melted, different fragments of DNA molecules (15, 16). The results suggested that pre-treatment of the irradiated rats with Cu(II)(Picolinyl-L- Histidinate) 2 , Cu(II)(Nicotinyl-L-Histidinate) 2 and Cu(II)(Isonicotinyl-L-Histidinate) 2 compounds improve the liver DNA characteristics. Table 2. The melting parameters of DNA isolated from I and II groups of rats on 1st and 30th day and from III, IV, V and VI groups of rats on 3, 7, 14, 30 days after irradiation at 6.5Gy (10mg/kg subcutaneously 1 hour prior to irradiation) Study groups T m , С о D T, С о D h, % Norm Control 71.2±0.10 7.3±0.15 33±0.15 1 day Cu(II) (Picolinyl-L-Histidinate) 2 Cu(II) (Nicotinyl- L-Histidinate) 2 Cu(II) (Isonicotinyl-L-Histidinate) 2 70.9±0.2 71.2±0.1 71.5±0.15 7.4±0.15 7.3±0.1 7.5±0.15 31.0±0.1 32.5±0.15 31.5±0.2 30 days Cu(II) (Picolinyl-L-Histidinate) 2 Cu(II) (Nicotinyl- L-Histidinate) 2 Cu(II) (Isonicotinyl-L-Histidinate) 2 71.0±0.2 71.1±0.15 71.3±0.2 7.3±0.10 7.4±0.2 7.3±0.25 32.0±0.15 33.3±0.2 32.5±0.1 3 days Irradiated Control, 6.5Gy Cu(II) (Picolinyl-L-Histidinate) 2 +6.5Gy Cu(II) (Nicotinyl-L-Histidinate) 2 +6.5Gy Cu(II) (Isonicotinyl-L-Histidinate) 2 +6.5Gy 68.5±0.15 70.1±0.2 70 ±0.1 70.1±0.15 17±0.20 7.6±0.15 8±0.2 7.9±0.15 22±0.15 29.5±0.1 29±0.15 29.6±0.2 7 Days Irradiated Control, 6.5Gy Cu(II) (Picolinyl-L-Histidinate) 2 +6.5Gy Cu(II) (Nicotinyl-L-Histidinate) 2 + 6.5Gy Cu(II) (Isonicotinyl-L-Histidinate) 2 +6.5Gy 67.5±0.2 70±0.15 69.5±0.1 70±0.2 23±0.2 8.1±0.1 8.2±0.15 7.6±0.2 16±0.15 28.3±0.15 28±0.1 27.5±0.2 14 Days Irradiated Control, 6.5Gy Cu(II)(Picolinyl-L-Histidinate) 2 +6. 5Gy Cu(II)(Nicotinyl-L-Histidinate) 2 + 6.5Gy Cu(II)(Isonicotinyl-L-Histidinate) 2 +6.5Gy 64.3±0.2 69±0.15 68.3±0.1 70.4 ±0.1 25±0.15 10.5±0.2 11.4 ±0.15 8.2±0.15 15±0.15 27.7±0.1 26±0.15 27.3±0.1 30 Days Irradiated Control, 6.5Gy Cu(II) (Picolinyl-L-Histidinate) 2 +6.5Gy Cu(II) (Nicotinyl-L-Histidinate) 2 +6.5Gy Cu(II) (Isonicotinyl-L-Histidinate) 2 +6.5Gy 62.2±0.2 67.8±0.2 68.2±0.1 70.1±0.15 27.1±0.2 12.5±0.15 11.8±0.1 8.4±0.2 14±0.2 26.3±0.15 26.5±0.1 27.8.±0.15 The melting parameters of rat DNA treated before irradiation at a dose of 6.5Gy are approaching the norm (Table 2). Thus, increasing the T m and Δh, and decreasing the ΔT in comparison to the irradiated rat's DNA was observed on 3, 7, 14 and 30 days. The studied compounds improved the melting parameters of DNA isolated from the rat liver, indicating the preservation of DNA's secondary structure stability (10, 15, 17). Hence, it can be assumed that the investigated Cu(II)-L-Histidinate complexes had radioprotective properties. For the visualization of the obtained data, the agarose gel electrophoresis method was used to separate DNA fragments based on their sizes. This technique allows for the identification and estimation of post-irradiation DNA damage. Figure 4 depicts the corresponding agarose gel electrophoretic patterns of DNA isolated from the rat's liver, which was irradiated and pre-treated with Cu(II)-L-Histidinate complexes before irradiation at 7, 14, 30 days. As seen, the DNA from the healthy rat's liver (norm, N) shows a homogeneous stain, corresponding to high molecular DNA with a size of 15kbp. In contrast, the DNA patterns of irradiated rats on days 7, 14, and 30 exhibit high electrophoretic mobility, indicating DNA fragmentation. The data clearly reveal the absence of DNA with high molecular weight on the electrophoresis tracks of irradiated rat DNA (Figure 4A, B, C marked 1). DNA fragments with sizes of 1-9 kb, 0.5-8 kb, and 0.2-4 kb were detected at 7, 14, and 30 days of irradiation, respectively. DNA samples obtained from the liver of rats pretreated with Cu(II)-L-Histidinate complexes exhibit electrophoretic bands similar to healthy DNA. Thus, DNA with sizes of 11-14kbp, 11-15kbp, and 11-15kbp were detected in the livers of rats pre-treated with Cu(II)(Picolinyl-L-Histidinate)₂ and Cu(II)(Nicotinyl-L-Histidinate)₂ on days 7, 14, and 30, respectively (Figure 4A, B, marked 2). For rats pre-treated with Cu(II)(Isonicotinyl-L-Histidinate)₂ (Figure 4C, marked 2), DNA bands with a size of 15kbp were detected on all investigated days. Thus, the chelating agents may protect DNA from hydroxyl radicals and prevent enzyme damage, which is responsible for the DNA enzymatic repair system. The efficiency of the chemical pathway of DNA repair has been studied in the works of Morozova (18, 19). In this works, amino acids were considered as electron donors, providing the reduction of radicals of the guanosine purine base. DISCUSSION Ionizing radiation has sufficient energy to induce free radicals in biological molecules, DNA being a particularly important target. Free radicals lead to the formation of potentially lethal DNA damages ( 20 , 21 ). The development of novel and effective approaches using non-toxic radioprotectors is of considerable interest for defense of nuclear industries, radiation accidents, and especially in medicine, in tumor radiotherapy ( 7 , 22 ). It is known that compounds containing heterocycles have high chemotherapeutic properties and serve as the basis for the development of new drugs. Many heterocyclic compounds are used in clinical practice for the treatment of infectious diseases. For example, nitrogen compounds containing an imidazole fragment easily interact with active centers in living organisms and have a unique position in the chemistry of heterocycles ( 23 , 24 ). The radioprotective effects of the Cu(II)(Picolinyl-L-Histidinate) 2 , Cu(II)(Nicotinyl-L-Histidinate) 2 and Cu(II)(Isonicotinyl-L-Histidinate) 2 compounds were assessed by determining the survival of rats, pre-treated with investigated complexes and irradiated at 6.5Gy dose of X-rays. According to the results obtained, the Cu(II) L-Histidinate Schiff base complexes had a strong radioprotective effect when administered orally and subcutaneously to rats under X-ray exposure at a dose of 6.5Gy. The used radiation dose corresponded to LD 60 , i.e. the survival rate of rats was 40%. Pre-treatment of animals with the studied complexes increased the survival rate of rats during the 30 days after irradiation to 70–100%. The previous report ( 25 ) presents the evaluation of the cytogenetic state of bone marrow cells in rats surviving to day 30 after irradiation at 6.5Gy. In irradiated rats, compared with the norm, there was an increase in the number of chromosomal aberrations to 7.7% and the number of tetraploid chromosomes to 2.1% (3.1% and 0.2% in the norm). However, in rats pre-treated with the Cu(II) Histidinate Schiff base complexes, there was a tendency toward the normalization of these parameters. Structural changes of the DNA molecule were identified using the DNA melting method, because since this method is widely used in identifying damages in DNA molecule ( 26 , 27 ). The melting parameters of DNA isolated from the liver of irradiated rats on the 3, 7, 14 and 30 days showed deviations from normal values, which indicated structural changes in the DNA molecule. Based on the DNA melting parameters, we can conclude that damages of the DNA molecule increased during the studied periods of the experiment. The presence of oxygen radicals induced by ionizing radiation increased DNA damages on 30 day, which contributed to the death of animals. The melting parameters of DNA isolated from rats pre-treated with Cu(II) L-histidinate-Schiff base complexes on the same (i.e. 3, 7, 14 and 30) days of the experiment significantly improved and approached to normal. The studied compounds demonstrated radiotherapeutic effects and ensured the stability of the secondary structure of rat liver DNA from damages. Our electrophoretic data correspond well to the literature, which showed the detection of various fragments with strand breaks in DNA samples obtained from irradiated animals ( 28 ). Free radical processes appear to be the primary cause of radiation damage to DNA. The activation level of free-radical processes significantly increases over time. As observed, DNA damage in irradiated samples also increases over time. At the same time, there are published data indicating that the activation of endogenous nucleases occurs during X-ray irradiation, leading to the accumulation of single- and double-strand breaks in DNA ( 29 ). The results clearly show that DNA isolated from animals pre-treated with the Cu(II) chelates exhibits electrophoretic bands similar to healthy DNA. Our results show that the presence of a metal in the investigated complexes plays an important role in ensuring their radioprotective properties. Preliminary studies of the radioprotective properties of Schiff bases derived from L-Histidine and 2-, 3-, and 4-pyridine carboxaldehyde showed weak radioprotective activity at irradiation dose of 6.5Gy ( 25 ). However, at complexation with Cu(II) Histidinate Schiff base metal complexes became effective radiation-protective agents capable of reducing or preventing radiation-induced lethality in rats. Perhaps this is due to the fact that the main enzymes involved in reactions that prevent the accumulation of toxic metabolic products are metal-dependent. They promote biochemical, cellular and tissue restoration, and also provide recovery after radiation injury. In this regard, copper cations are probably the most promising ( 30 , 31 ). Trace elements such as Cu(II) in Schiff base form have a protective effect against cellular toxicity caused by oxidative stress, especially with superoxide dismutase mimetic activity. It can inhibit oxidative stress and protect the DNA against peroxide radicals. In previous work, the activity of the antioxidant enzymes superoxide dismutase and catalase was determined on 3, 7, 14, and 30 days after irradiation at a dose of 6.5Gy in the blood of animals. The results of the studies revealed superoxide dismutase and catalase mimetic activity of Cu(II)L-Histidinate Schiff base complexes ( 25 ). The superoxide dismutase and catalase mimetic activity of the copper organic complexes have an important effect on the body's defense against radiation-induced oxidative stress. Conclusion Newly synthesized compounds Cu(II)(Picolinyl-L-Histidinate) 2 , Cu(II)(Nicotinyl-L-Histidinate) 2 and Cu(II)(Isonicotinyl-L-Histidinate) 2 are effective radiation protective agents capable to reduce or prevent radiation-induced lethality in rats. All three metal complexes demonstrate significant radiation protection through both subcutaneous and oral administration to rats. Moreover, 100% survival was registered in rats irradiated on the background oral administration of 40mg/kg Cu(II)(Picolinyl-L-Histidinate) 2 24 hours prior to irradiation. The same result, i.e. 100% survival, was achieved in the case of subcutaneous injection of 10mg/kg Cu(II)(Isonicotinyl-L-Histidinate) 2 to rats one hour before exposure at the dose of 6.5Gy. A study of DNA melting curves and electrophoresis data revealed improved characteristics of rat liver DNA in animals pretreated with Cu(II) Schiff base complexes before to X-ray exposure. Declarations Conflict of interest. The authors declare that they have no conflict of interest. Statement of Human and Animal Rights Experiments were fulfilled according to the "International Recommendations on carrying out of biomedical Research with use of Animals" (CIOMS, 1985), to the "Human Rights and Biomedicine, Oviedo Convention" (CE, 1997), to the European Convention for the Protection of Vertebral Animals Used for Experimental and Other Scientific Purposes (CE, 2005) and approved by the National Center of Bioethics (Armenia). Author Contribution N. K. contributed in the execution of the experiments, data analysis, investigation, design, , writing and drafting the original manuscript. All experimental data was analyzed by N. K., G. A. and S. H. and contributed to the article review and editing. Reagent preparation and experimentation were carried out by N. K. and G. A., analysed and reviewed the manuscript. The manuscript was critically revised for accurate knowledgeable content by S. H. Finally, all authors approved the final version of the manuscript for publication. 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Structural Chemistry. 2011. 22, 475-482. https://doi.org/10.1007/s11224-011-9765-4 Karapetyan NH, Malakyan MH, Bajinyan SA, Torosyan AL Grigoryan IE, Haroutiunian SG. (2013) Influence of Amino Acids Shiff Bases on Irradiated DNA Stability In Vivo. Cell Biochem Biophys. 2013. 67, 1137–1145. https://doi.org/10.1007/s12013-013-9617-5 Karapetyan NH, Torosyan AL, Malakyan MH, Bajinyan SA, Haroutiunian SG. (2016) Investigation of irradiated rats DNA in the presence of Cu(II) chelates of amino acids Schiff bases. Journal of Biomolecular Structure and Dynamics. 2016. 34, 177-183. http://dx.doi.org/10.1080/07391102.2015.1020876 Jomova K, Makova M, Alomar SY, Alwasel SH, Nepovimova E, Kuca K et al. (2022) Essential metals in health and disease. Chemico-Biological Interactions. 2022. 367, 110173. https://doi.org/10.1016/j.cbi.2022.110173 Malakyan МН, Bajinyan SA, Matosyan VH, Tonoyan VJ, Babayan KN. (2016) Synthesis, characterization and toxicity studies of pyridinecarboxaldehydes and L-tryptophan derived Schiff bases and corresponding copper (II) complexes. F1000 Research, 2016. 5, 1921. doi:10.12688/f1000research.9226.1 Lando DY, Teif VB. (2002) Modeling of DNA Condensation and Decondensation Caused by Ligand Binding. Journal of Biomolecular Srtucture and Dynamics. 2002. 20, 215-222. https://doi.org/10.1080/07391102.2002.10506837 Wells RD, Larson JE, Grant RC, Shortle BE, Cantor CR. (1970) Physicochemical studies on polydeoxyribonucleotides containing defined repeating nucleotide sequences. Journal of Molecular Biology. 1970. 54, 465-497. doi:10.1016/0022-2836(70)90121-x. Wartell RM, Benight AS. (1985) Thermal denaturation of DNA molecules: A comparison of theory with experiment. Physics Reports. 1985. 126, 67–107. https://doi.org/10.1016/0(370-1573(85)90060-2 Lee PY, Costumbrado J, Hsu C-Y, Kim YH. (2012) Agarose Gel Electrophoresis for the Separation of DNA Fragments. J Vis Exp. 2012. 62, 3923. doi: 10.3791/3923 Vologodskii A, Frank-Kamenetskii MD. (2018) DNA melting and energetics of the double helix. Physics of Life Reviews. 2018. 25, 1-21. https://doi.org/10.1016/j.plrev.2017.11.012 Morozova OB, Kiryutin AS, Sagdeev RZ, Yurkovskaya AV. (2007) Electron Transfer between Guanosine Radical and Amino Acids in Aqueous Solution. 1. Reduction of Guanosine Radical by Tyrosinev J. Phys. Chem. B, 2007. 111, 7439–7448. DOI: 10.1021/jp067722i Morozova OB, Kiryutin AS, Yurkovskaya AV. (2008) Electron Transfer between Guanosine Radicals and Amino Acids in Aqueous Solution. II. Reduction of Guanosine Radicals by Tryptophan. Journal of Physical Chemistry 2008. B 112, 2747-2754. DOI: 10.1021/jp0752318 Ward JF . ( 1988) DNA damage produced by ionizing radiation in mammalian cells: iden­tities, mechanisms of formation, and reparability. Prog. Nucleic Acid Res. Mol. Biol. 1988. 35 , 95–125. https://doi.org/10.1016/S0079-6603(08)60611-X Nikjoo H, O'Neill P, Terrissol M, Goodhead DT. (1994) Modelling of radiation -induced DNA damage: the early physical and chemical event. Int. J. Radiat. Biol . 1994. 66 , 453–457.https://doi.org/10.1080/09553009414551451 Johnke RM, Sattler JA, Allison RR. (2014) Radioprotective agents for radiation therapy: future trends. Future Oncol. 2014. 10(15), 2345-2357. https://doi.org/10.2217/fon.14.175 Reyes-Arellano A, Gómez-García O, Torres-Jaramillo J. (2016). Synthesis of azolines and imidazoles and their use in drug design. Med Chem. 2016; 6:561–70. DOI: 10.4172/2161-0444.1000400. Verma A, Joshi S, Singh D. (2013) Imidazole: having versatile biological activities. New J Chem. 2013. 1-12. https://doi.org/10.1155/2013/329412 Malakyan MH, Dallakyan A, Bajinyan SA, Tonoyan V, Ayvazyan V, Karapetyan NH. (2017) Development of potential radioprotective agents for use in field exposure situations. International Conference. BRITE “Biomarkers of Radiation In The Environment: Robust tools for risk assessment” 2017, Yerevan, Armenia, p. 22. Kalfas CA, Loukakis GK, Georgakilas AG, Sideris E.G, Anagnostopoulou -Konsta A. (1996) Flexibility and thermal denaturation (melting) of irradiated DNA. Journal of Biological Systems. 1996. 4, 405-423. https://doi.org/10.1142/S0218339096000272 Tankovskaia SA, Kotb OM, Dommes OA, Paston SV. (2018) DNA Damage Induced by Gamma-Radiation Revealed from UV Absorption Spectroscopy J. Phys.:Conf. Ser. 2018. 1038, 1-6. doi:10.1088/1742-6596/1038/1/012027 Torudd J, Protopopova M, Sarimov R, Nygren J, Eriksson S, Marková E et al. (2005) Dose-response for radiation-induced apoptosis, residual 53BP1 foci and DNA-loop relaxation in human lymphocytes. International Journal of Radiation Biology. 2005. 8, 125-138. DOI: 10.1080/09553000500077211 Bakayev VV, Yugai AA, Luchnik AN. (1985). Effect of X-ray induced DNA damage on DNAase I hypersensitivity of SV40 chromatin: relation to elastic torsional strain in DNA. Nucleic Acids Research, 1985; 13, 7079–7093. DOI: 10.1093/nar/13.19.7079 Zoroddu MA, Aaseth J, Crisponi G, Medici S, Peana M, Nurchi VM. (2019) The essential metals for humans: a brief overview. J. Inorg Biochem. 2019. 195, 120-129. https://doi.org/10.1016/j.jinorgbio.2019.03.013 Sorenson JRJ. (2002). Cu, Fe, Mn, and Zn chelates offer a medicinal chemistry approach to overcoming radiation injury. Current Med. Chem. 2002. 9, 639-662. doi: 10.2174/0929867023370725 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 28 May, 2024 Reviews received at journal 28 May, 2024 Reviews received at journal 13 May, 2024 Reviewers agreed at journal 11 May, 2024 Reviewers agreed at journal 11 May, 2024 Reviewers agreed at journal 08 May, 2024 Reviewers agreed at journal 08 May, 2024 Reviewers agreed at journal 08 May, 2024 Reviewers invited by journal 08 May, 2024 Submission checks completed at journal 08 May, 2024 Editor assigned by journal 08 May, 2024 First submitted to journal 08 May, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4387030","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":302038727,"identity":"2b764ad6-c16f-4429-8123-1a7cc313cf17","order_by":0,"name":"Nelli H. Karapetyan","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAn0lEQVRIiWNgGAWjYFAC5mZmhgrStDACtZwhWQtjGykazNkbm40L5x22NzjA/HQDUVosew42J8/cdjhxwwE2sxtEaTG4kdh8mHfb4QSDAwzEarn/EKhlDshh7N+ItYWxOZm34TDjhgM8xNpyJrHZmOdYeuLMwzxlRGo5fviwNE+NtT3f8fZtxGmBgmZgKiBFPRDUkah+FIyCUTAKRhQAAJFIMsJ+JzlPAAAAAElFTkSuQmCC","orcid":"","institution":"Yerevan State University","correspondingAuthor":true,"prefix":"","firstName":"Nelli","middleName":"H.","lastName":"Karapetyan","suffix":""},{"id":302038728,"identity":"9db5ebd2-24bb-4efb-be8e-132a6417fd8a","order_by":1,"name":"Samvel G. Haroutiunian","email":"","orcid":"","institution":"Yerevan State University","correspondingAuthor":false,"prefix":"","firstName":"Samvel","middleName":"G.","lastName":"Haroutiunian","suffix":""},{"id":302038729,"identity":"f4923ea0-8918-44b7-867a-b4383fb025ca","order_by":2,"name":"Gayane V. Ananyan","email":"","orcid":"","institution":"Yerevan State University","correspondingAuthor":false,"prefix":"","firstName":"Gayane","middleName":"V.","lastName":"Ananyan","suffix":""}],"badges":[],"createdAt":"2024-05-08 06:42:06","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4387030/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4387030/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":56621545,"identity":"85eb4d0f-7ffd-43c6-a135-d0921daa30f7","added_by":"auto","created_at":"2024-05-16 18:09:04","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1638547,"visible":true,"origin":"","legend":"\u003cp\u003eThe chemical structure of Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2 \u003c/sub\u003e(1), Cu(II)(Nicotinyl-L- Histidinate)\u003csub\u003e2\u003c/sub\u003e (2) and Cu(II)( Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e (3), which differ from each other by the position of the nitrogen atom in the pyridine ring.\u003c/p\u003e","description":"","filename":"Figure1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4387030/v1/1369e08b16518c20f90ba415.jpg"},{"id":56621543,"identity":"5b5b221b-e58c-4fa3-a7c7-cf4ffcc29291","added_by":"auto","created_at":"2024-05-16 18:09:03","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":494159,"visible":true,"origin":"","legend":"\u003cp\u003eRats survival during 30 days post X-ray irradiation on the background of subcutaneous administration of 10 mg/kg (a) and 40mg/kg (b) Cu(II) complexes 1 h before the exposure at 6.5Gy.\u003cstrong\u003e ■ – \u003c/strong\u003eIrradiated Control,\u003cstrong\u003e ● – \u003c/strong\u003eCu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e, ▲\u003cstrong\u003e–\u003c/strong\u003eCu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e,\u003csub\u003e \u003c/sub\u003e▼\u003cstrong\u003e– \u003c/strong\u003eCu(II)( Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u003cstrong\u003e.\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Figure2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4387030/v1/c9b882eee16f657919db326d.jpg"},{"id":56621546,"identity":"b70c17bd-0969-4ab0-b8c7-d21d35cc76fd","added_by":"auto","created_at":"2024-05-16 18:09:04","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":495559,"visible":true,"origin":"","legend":"\u003cp\u003eRats survival during 30 days post X-ray irradiation on the background of oral administration of 10 mg/kg (a) and 40mg/kg (b) Cu(II) complexes 24 h before the exposure at 6.5Gy. \u003cstrong\u003e■ – \u003c/strong\u003eIrradiated Control,\u003cstrong\u003e ● – \u003c/strong\u003eCu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e, ▲ \u003cstrong\u003e– \u003c/strong\u003eCu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e,\u003csub\u003e \u003c/sub\u003e▼\u003cstrong\u003e– \u003c/strong\u003eCu(II)( Isonicotinyl-L-Histidinate.\u003c/p\u003e","description":"","filename":"Figure3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4387030/v1/d5e5f3d9892e466c929617f2.jpg"},{"id":56621544,"identity":"2e909375-be0b-4829-b30d-8ced3f8780bc","added_by":"auto","created_at":"2024-05-16 18:09:04","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":371385,"visible":true,"origin":"","legend":"\u003cp\u003eElectrophoresis of isolated from investigated rats groups on 7, 14 and 30 days after irradiation.\u0026nbsp; M-Marker, N-Norm, Control. 1-Irradiated Control, 2-pretreated by Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e (A), Cu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e (B), and Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e (C) on 7, 14 and 30 days.\u003c/p\u003e\n\u003cp\u003eDNA\u003c/p\u003e","description":"","filename":"Figure4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4387030/v1/275345a6b28293e619006f5c.jpg"},{"id":56622209,"identity":"1dc47e13-a7d3-4c24-80a8-202cba824edf","added_by":"auto","created_at":"2024-05-16 18:17:05","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3487508,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4387030/v1/e2a897e5-784e-4350-b98f-35813a2c1184.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Influence of Cu L-Histidinate Schiff base derivatives on structural features of irradiated rat’s DNA","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eIonizing radiation, being high-energy, generated ions that can destroy the covalent bonds of molecules. It can cause various types of damage to DNA, RNA, proteins, and other biomolecules. Among these, DNA is the major target of radiation-induced damage within the cell. DNA molecule damage occurs either through direct or indirect ionization through the production of water radicals, which affect and damage DNA (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn general, the sugar-phosphate backbone lesions generate DNA single-strand breaks (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). If damage occurs on opposite strands of DNA within the same helical turn, then double-strand DNA breaks are formed. The radiolysis of water results the formation of reactive oxygen species like superoxide radicals, hydrogen peroxide, singlet oxygen, and highly reactive hydroxyl radicals (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e). As a result, these species oxidize proteins and lipids, and also induce various types of damage to DNA, including modifications in purine and pyrimidine bases, sugar damage, and apurinic or apyrimidinic sites damage, removal of bases, cross-linking of DNA with DNA or adjacent proteins (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). These changes induce cell death and mitotic failure (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e). The extent of radiation damage to the body depends on cellular antioxidant statuses, such as antioxidant compounds and enzyme systems (e.g. superoxide dismutase, catalase, peroxidases) that reduce intracellular reactive oxygen species levels.\u003c/p\u003e \u003cp\u003eCurrently, a special area of radiobiology is the search for means of chemical protection against radiation. These substances block the development of chain radiation-chemical reactions by intercepting active radicals, antioxidants, and agents that create tissue hypoxia, reduce the intensity of oxidative processes by binding metal ions with variable valence, and catalyze oxygen transfer. The protection of sulfhydryl groups of proteins by stimulating cellular recovery systems has been demonstrated (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e). The possibility of introducing agents required for DNA repair is being studied (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe purpose of this study was to identify DNA damage in the liver of X-ray irradiated rats and DNA repair in rats pretreated with Cu(II) L-Histidinate Schiff bases complexes before irradiation. In this study, various exposure schemes were used to identify the most optimal one.\u003c/p\u003e \u003cp\u003ePromising results were obtained in the study of the radioprotective activity of the Cu(II) and Mn(II) complexes with Schiff bases derived from isomeric pyridincarboxaldehydes with tryptophan and tyrosine cyclic amino acids (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e). Metals, such as copper, cobalt, iron, manganese, and zinc are elements required by all cells for normal metabolic processes. Ionic forms of these metals have particularly high affinities for organic ligands found in biological systems and rapidly undergo bonding interactions to form complexes, or more precisely, coordinate-covalent bonded chelate structures \u003cem\u003e(11).\u003c/em\u003e Chelated forms of metals exist in tissues in the form of metal-dependent enzymes, proteins, and various low molecular-weight peptide chelates. Transition metal ions can coordinate the ligand in a precise 3D configuration, thus adapting the molecule to recognize and interact with the target molecule.\u003c/p\u003e \u003cp\u003eThe synthesis of metal element-dependent enzymes, required for oxygen utilization and superoxide accumulation prevention, as well as tissue repair processes, including metal element-dependent DNA and RNA repair, is key to the hypothesis that small molecular mass essential metal chelates such as Schiff bases facilitate recovery from radiation-induced pathology.\u003c/p\u003e"},{"header":"MATERIALS AND METHODS","content":"\u003cp\u003eAll used chemicals and reagents of Analytical Reagent grade were obtained from Sigma-Aldrich. Cu(II) Schiff Base complexes with L-Histidine and 2-, 3- and 4-pyridinecarboxaldehydes were used for animal studies (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). These new complexes were synthesized and kindly presented by V. Matosyan and V. Tonoyan at the Scientific Centre of Radiation Medicine and Burns (Yerevan, Armenia) (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe toxicity of synthesized Schiff Bases was characterized based on a calculation of the value of LD\u003csub\u003e50\u003c/sub\u003e, which is the dose of a compound at which the lethality of 50% of animals was observed in 7 days after subcutaneous administration of the compound to the organism. With the use of the Behrens integration method in experiments in mice, the toxicity of compounds was calculated (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e). According to the test results, Cu(II) complexes with Schiff Bases proved to be low toxic.\u003c/p\u003e \u003cp\u003eMale albino rats of the Wistar strain with 180-200g were used for experimental studies. The rats were bred at the vivarium of the Scientific Centre of Radiation Medicine and Burns (Yerevan, Armenia) under conventional laboratory conditions.\u003c/p\u003e \u003cp\u003eTo undergo whole-body irrigation X-ray, 10 rats were placed in a well-ventilated box, located 50cm from the source of radiation. The RUM-17 (RF) therapeutic X-ray was used for irradiation. The radiation conditions were the following: voltage 180kV, current 15mA, without filter, dose rate 1.78Gy/min.\u003c/p\u003e \u003cp\u003eThe studies were carried out under various schemes of exposure in order to choose the most optimal of them. The design of the experiment is as follows:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003eX-ray exposure of white rats at 6.5Gy (LD\u003csub\u003e60\u003c/sub\u003e).\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003eTreatments: 1 or 24 hours prior to irradiation;\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003eDrug administration mode: subcutaneous and oral;\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003eDose levels: 10 mg/kg or 40 mg/kg.\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eThis article presents experimental data for rats administrated subcutaneously by investigating compounds 1 hour prior to irradiation at a dose of 6.5Gy. The rats\u0026rsquo; liver DNA was isolated from the following 6 groups of animals:\u003c/p\u003e \u003cp\u003eI group: intact animals served as control\u003c/p\u003e \u003cp\u003eII group: rats obtained 10mg/kg Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e, Cu(II)(Nicotinyl-L- Histidinate)\u003csub\u003e2\u003c/sub\u003e and Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e (the pure effect) on 1st and 30th days.\u003c/p\u003e \u003cp\u003eIII group: animals exposed to X-ray irradiation at 6.5Gy dose level (Irradiated control group)\u003c/p\u003e \u003cp\u003eIV group: rats pre-treated subcutaneously administration of 10mg/kg Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e 1 hour before exposure to X-ray irradiation at 6.5Gy dose level\u003c/p\u003e \u003cp\u003eV group: rats obtained 10mg/kg Cu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e subcutaneously 1 hour prior to irradiation at 6.5Gy\u003c/p\u003e \u003cp\u003eVI group: rats obtained 10mg/kg Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e subcutaneously 1 hour prior to irradiation at 6.5Gy\u003c/p\u003e \u003cp\u003eCu(II) complexes used in this study were administrated to the rats in the form of suspension using deionizing water as solvent. The animals were injected subcutaneously with 2mg Cu(II) chelates in 0.5ml solution on 200mg body weight 1 hour before irradiation.\u003c/p\u003e \u003cp\u003eIn order to perform analyses, rats were sacrificed under anesthesia on 3, 7, 14, and 30 days after irradiation (per 5 rats at each point). Data are presented as the average of three measurements.\u003c/p\u003e \u003cp\u003e \u003cb\u003eDNA isolation.\u003c/b\u003e DNA was isolated from animal's liver using the standard chloroform method (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e). The absorbance of DNA solution at 230 nm, 260 nm and 280 nm was used to evaluate DNA purity. The DNA samples had А\u003csub\u003e260\u003c/sub\u003e/А\u003csub\u003e280\u003c/sub\u003e = 1.8 and А\u003csub\u003e260\u003c/sub\u003e/А\u003csub\u003e230\u003c/sub\u003e = 2 and were free from contaminations.\u003c/p\u003e \u003cp\u003e \u003cb\u003eSpectroscopy.\u003c/b\u003e The absorption spectra and ultraviolet melting curves of DNA samples were recorded on a Lambda 800 UV/VIS spectrometer (Perkin-Elmer). DNA concentrations were determined spectrophotometrically with ε\u003csub\u003e260nm\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;1.31x10\u003csup\u003e4\u003c/sup\u003e M\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003ecm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e and calculated in base pairs (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e). All spectroscopic measurements were performed in biphosphate buffer solution (0.6mM Na\u003csub\u003e2\u003c/sub\u003eHPO\u003csub\u003e4\u003c/sub\u003e, 0.2mM NaH\u003csub\u003e2\u003c/sub\u003ePO\u003csub\u003e4\u003c/sub\u003e, 18.5mM NaCl, 0.01mM EDTA), [Na\u003csup\u003e+\u003c/sup\u003e]\u0026thinsp;=\u0026thinsp;0.02, pH 7.2.\u003c/p\u003e \u003cp\u003e \u003cspan class=\"InlineEquation\"\u003e \u003cspan class=\"mathinline\"\u003e\\(\\Delta h=({A_{{{95}^o}C}}/{A_{{{25}^o}C}} - 1) \\cdot 100\\%\\)\u003c/span\u003e \u003c/span\u003eMelting experiments were carried out at a 35-95\u003csup\u003eo\u003c/sup\u003eC temperature interval, using 10mm thermostatic quartz cuvettes. The heating rate was 0.5\u003csup\u003eo\u003c/sup\u003eC/min, while absorbance at 260 nm was recorded. Melting characteristics of DNA - the melting temperature (T\u003csub\u003em\u003c/sub\u003e), the width of the melting interval ΔT=(\u003cem\u003e\u0026part;θ/\u0026part;T\u003c/em\u003e)\u003csup\u003e\u0026minus;1\u003c/sup\u003e\u003csub\u003e\u003cem\u003eT=Tm\u003c/em\u003e\u003c/sub\u003e and hypochromic effect were calculated from melting curves \u003cem\u003e(14).\u003c/em\u003e\u003c/p\u003e \u003cp\u003eT\u003csub\u003em\u003c/sub\u003e is the temperature at which half of all base pairs \u0026ldquo;melted\u0026rdquo;, i.e. 1-θ\u0026thinsp;=\u0026thinsp;0.5, and ΔT is the width of the melting interval, equal to the temperature difference at which the tangent at the inflection point crosses the levels θ\u0026thinsp;=\u0026thinsp;0 and θ\u0026thinsp;=\u0026thinsp;1 (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cb\u003eElectrophoresis.\u003c/b\u003e DNA electrophoresis was performed using 1% agarose gel in Tris-boric acid - EDTA buffer, pH 8.0, under condition of 5 V/cm. The 1kb DNA Ladder (Promega Product, USA) was used as a marker for determining the size of double-stranded DNA from 250\u0026thinsp;\u0026minus;\u0026thinsp;10 000 base pairs (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e). The gel was stained with 0.5 ng/ml ethidium bromide solution, viewed and photographed under UV-trans illuminator.\u003c/p\u003e"},{"header":"RESULTS","content":"\u003cp\u003eThe effectiveness of the studied Cu(II) L-Histidinate Schiff base compounds, as radioprotectors was assessed by the survival of animals for 30 days (Table 1). \u0026nbsp;On the 30th day of the experiment, the survival rate of rats for all investigated Cu(II) L-Histidinate Schiff base compounds \u0026nbsp;was 100%. When rats were irradiated with a dose of 6.5Gy, their survival rate on 30 day was 40%. However, pre-treatment with Cu(II)L-Histidinate complexes before irradiation increased the survival rate of animals by 1.5-2.5 times.\u003c/p\u003e\n\u003cp\u003eAll studied Schiff base complexes exhibited a pronounced radioprotective effect in all cases of use. However, the effectiveness of the compounds depends on the scheme of their administration. According to our results (Fig. 2 and Fig. 3), the most effective radioprotective effect was observed with subcutaneous administration of drugs. Good results (80-100% survival) were obtained when 10 mg/kg Cu(II) complexes were administered to rats 1 hour before irradiation. At oral use, a good radioprotective effect (75-100% survival) was observed in the case of administration at 40 mg/kg dose 24 hours prior to irradiation.\u003c/p\u003e\n\u003cp\u003eProbably this can be explained by the fact that, upon oral administration, the complexes are absorbed into the blood more slowly and effectiveness of compounds was observed later. Administration of Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e exhibited 80% survival by subcutaneous administration at 10mg/kg dose 1 hour before irradiation, and 100% survival by oral administration at 40mg/kg dose 24 hours prior to exposure.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1.\u003c/strong\u003e Survival of animals 30 days after irradiation exposure to X-rays at 6.5Gy on the background of subcutaneous or oral administration of 10 mg/kg and 40 mg/kg Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e, \u0026nbsp;Cu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e,and Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u0026nbsp;\u003c/sub\u003e1or \u0026nbsp;24 hours prior to irradiation. \u0026nbsp;At all experiments Norm (intact) rats survival was 100%, Irradiated rats Control survival was 40 %. \u0026nbsp;p \u0026le; 0.02\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003eSubstance\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003eMode\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003eDose, mg/kg\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\"\u003e\n \u003cp\u003eSurvival,%\u0026nbsp;6.5Gy\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e1h prior\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e24h prior\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\"\u003e\n \u003cp\u003eCu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003esubcutaneous\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e80\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e60\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e75\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e60\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003eoral\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e90\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e70\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\"\u003e\n \u003cp\u003eCu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003esubcutaneous\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e70\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e70\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e60\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003eoral\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e80\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e85\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\"\u003e\n \u003cp\u003eCu(II)(\u0026nbsp;Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003esubcutaneous\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e60\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e60\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\"\u003e\n \u003cp\u003eoral\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e75\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e75\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e70\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e80\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003ePretreatment with 10mg/kg Cu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e subcutaneously 1 hour before irradiation resulted in a 90% survival rate. In the case of oral administration at a 40mg/kg dose 24 hours prior to irradiation, an 85% survival rate was observed. Rats administered with Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e exhibited 100% survival when given a subcutaneous dose of 10mg/kg one hour before \u0026nbsp;irradiation. With oral administration at a 40mg/kg dose 24 hours prior to irradiation, 80% survival was observed at 6.5Gy irradiation dose.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main cause of death in animals at radiation is damages of DNA, which is very sensitive to ionizing irradiation. The next stage of research was the identification of structural features of DNA during irradiation. \u0026nbsp;For this purpose DNA was isolated from the liver of rats on 3, 7, 14 and 30 days after irradiations. Structural changes in DNA also were detected in animals pre-treated subcutaneously with Cu(II) L-Histidinate Schiff bases. Damage to the DNA molecule was assessed by the melting method. The melting temperature (Т\u003csub\u003em\u003c/sub\u003e), melting interval (DТ) and hypochromic effect (\u0026Delta;h) provide information on DNA stability and defects \u0026nbsp;in the DNA secondary structure.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe pure effect of Cu(II)L-Histidinate complexes on the stability of rats liver DNA was examined on 1st and 30th days after administration of the compounds. The structural features of DNA did not change when the compounds was administered to rats. The DNA melting parameters were the same as those of normal DNA. Since Cu(II)L-Histidinate complexes do not affect DNA melting parameters on both 1st and 30th days, the stability of the DNA molecule is preserved (Table 2). \u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe obtained results showed that irradiation with a dose of 6.5Gy of the control group of animals promote deviation of the characteristics of the rat\u0026apos;s liver DNA from the norm. The DNA melting parameters of irradiated rats on days 3, 7, 14 and 30 of the experiment differed from the norm. In particular a decrease in T\u003csub\u003em\u003c/sub\u003e and \u0026Delta;h, an increase DТ were observed. On 30 days after irradiation the DNA of the irradiated rat\u0026apos;s destabilized (T\u003csub\u003em\u003c/sub\u003e decreased from 71.2\u003csup\u003eo\u003c/sup\u003eC to 62.2\u003csup\u003eo\u003c/sup\u003eC), the hypochromicity decreased (from 33% to 14%), and the \u0026Delta;T increased from 7.3\u003csup\u003eo\u003c/sup\u003eC to 27.1\u003csup\u003eo\u003c/sup\u003eC, in comparison to normal DNA. These changes in the melting parameters indicate the destabilization of the DNA molecule, and the presence of partially melted, different fragments of DNA molecules \u003cem\u003e(15, 16).\u003c/em\u003e The results suggested that pre-treatment of the irradiated rats with Cu(II)(Picolinyl-L- Histidinate)\u003csub\u003e2\u003c/sub\u003e, Cu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e and Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e compounds improve the liver DNA characteristics.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2.\u0026nbsp;\u003c/strong\u003eThe melting parameters of DNA isolated from I and II groups of rats on 1st and 30th day and from III, IV, V and VI groups of rats on 3, 7, 14, 30 days after irradiation at 6.5Gy (10mg/kg subcutaneously 1 hour \u0026nbsp;prior to irradiation)\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"297\"\u003e\n \u003cp align=\"center\"\u003e\u003cstrong\u003eStudy groups\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"91\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp; \u0026nbsp;T\u003csub\u003em\u003c/sub\u003e, С\u003csup\u003eо\u003c/sup\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"94\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eD\u003c/strong\u003e\u003cstrong\u003eT, С\u003csup\u003eо\u003c/sup\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"85\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eD\u003c/strong\u003e\u003cstrong\u003eh, %\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"297\"\u003e\n \u003cp\u003eNorm Control\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"91\" valign=\"top\"\u003e\n \u003cp\u003e71.2\u0026plusmn;0.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"94\" valign=\"top\"\u003e\n \u003cp\u003e7.3\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"85\" valign=\"top\"\u003e\n \u003cp\u003e33\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"567\" colspan=\"4\"\u003e\n \u003cp align=\"center\"\u003e\u003cstrong\u003e1 day\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"297\"\u003e\n \u003cp\u003eCu(II) (Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCu(II) (Nicotinyl- L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCu(II) (Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"91\"\u003e\n \u003cp\u003e70.9\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e71.2\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e71.5\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"94\"\u003e\n \u003cp\u003e7.4\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e7.3\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e7.5\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"85\"\u003e\n \u003cp\u003e31.0\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e32.5\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e31.5\u0026plusmn;0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"567\" colspan=\"4\"\u003e\n \u003cp align=\"center\"\u003e\u003cstrong\u003e\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; 30 days\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"297\"\u003e\n \u003cp\u003eCu(II) (Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCu(II) (Nicotinyl- L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCu(II) (Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"91\"\u003e\n \u003cp\u003e71.0\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e71.1\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e71.3\u0026plusmn;0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"94\"\u003e\n \u003cp\u003e7.3\u0026plusmn;0.10\u003c/p\u003e\n \u003cp\u003e7.4\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e7.3\u0026plusmn;0.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"85\"\u003e\n \u003cp\u003e32.0\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e33.3\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e32.5\u0026plusmn;0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"567\" colspan=\"4\"\u003e\n \u003cp align=\"center\"\u003e\u003cstrong\u003e3 days\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"297\"\u003e\n \u003cp\u003eIrradiated Control, 6.5Gy\u003c/p\u003e\n \u003cp\u003eCu(II) (Picolinyl-L-Histidinate)\u003csub\u003e2\u0026nbsp;\u003c/sub\u003e+6.5Gy\u0026nbsp;Cu(II) (Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e +6.5Gy\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCu(II) (Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e+6.5Gy\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"91\"\u003e\n \u003cp\u003e68.5\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e70.1\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e70 \u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e70.1\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"94\"\u003e\n \u003cp\u003e17\u0026plusmn;0.20\u003c/p\u003e\n \u003cp\u003e7.6\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e8\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e7.9\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"85\"\u003e\n \u003cp\u003e22\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e29.5\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e29\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e29.6\u0026plusmn;0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"567\" colspan=\"4\"\u003e\n \u003cp align=\"center\"\u003e\u003cstrong\u003e7 Days\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"297\"\u003e\n \u003cp\u003eIrradiated Control, 6.5Gy\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCu(II) (Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e +6.5Gy\u0026nbsp;Cu(II) (Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e + 6.5Gy\u003c/p\u003e\n \u003cp\u003eCu(II) (Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e+6.5Gy\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"91\"\u003e\n \u003cp\u003e67.5\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e70\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e69.5\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e70\u0026plusmn;0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"94\"\u003e\n \u003cp\u003e23\u0026plusmn;0.2\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e8.1\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e8.2\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e7.6\u0026plusmn;0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"85\"\u003e\n \u003cp\u003e16\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e28.3\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e28\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e27.5\u0026plusmn;0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"567\" colspan=\"4\"\u003e\n \u003cp align=\"center\"\u003e\u003cstrong\u003e14 Days\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"297\"\u003e\n \u003cp\u003eIrradiated Control, 6.5Gy\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e +6. 5Gy\u0026nbsp;Cu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e + 6.5Gy\u0026nbsp;Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e+6.5Gy\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"91\"\u003e\n \u003cp\u003e64.3\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e69\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e68.3\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e70.4 \u0026plusmn;0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"94\"\u003e\n \u003cp\u003e25\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e10.5\u0026plusmn;0.2 11.4 \u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e8.2\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"85\"\u003e\n \u003cp\u003e15\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e27.7\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e26\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e27.3\u0026plusmn;0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"567\" colspan=\"4\"\u003e\n \u003cp align=\"center\"\u003e\u003cstrong\u003e30 Days\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"297\"\u003e\n \u003cp\u003eIrradiated Control, 6.5Gy\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eCu(II) (Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e+6.5Gy\u0026nbsp;Cu(II) (Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e +6.5Gy\u0026nbsp;Cu(II) (Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e+6.5Gy\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"91\"\u003e\n \u003cp\u003e62.2\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e67.8\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e68.2\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e70.1\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"94\"\u003e\n \u003cp\u003e27.1\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e12.5\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e11.8\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e8.4\u0026plusmn;0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"85\"\u003e\n \u003cp\u003e14\u0026plusmn;0.2\u003c/p\u003e\n \u003cp\u003e26.3\u0026plusmn;0.15\u003c/p\u003e\n \u003cp\u003e26.5\u0026plusmn;0.1\u003c/p\u003e\n \u003cp\u003e27.8.\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eThe melting parameters of rat DNA treated before irradiation at a dose of 6.5Gy are approaching the norm (Table 2). Thus, increasing the T\u003csub\u003em\u003c/sub\u003e and \u0026Delta;h, and decreasing the \u0026Delta;T in comparison to the irradiated rat\u0026apos;s DNA was observed on 3, 7, 14 and 30 days. The studied compounds improved the melting parameters of DNA isolated from the rat liver, indicating the preservation of DNA\u0026apos;s secondary structure stability \u003cem\u003e(10, 15, 17).\u003c/em\u003e Hence, it can be assumed that the investigated Cu(II)-L-Histidinate complexes had radioprotective properties.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFor the\u0026nbsp;visualization\u0026nbsp;of the obtained data, the\u0026nbsp;agarose gel electrophoresis\u0026nbsp;method\u0026nbsp;was used to\u0026nbsp;separate DNA fragments based on their sizes. This technique allows for the identification and estimation of post-irradiation DNA damage. \u0026nbsp;Figure 4 depicts the corresponding agarose gel electrophoretic patterns of DNA isolated from the rat\u0026apos;s liver, which was irradiated and pre-treated with Cu(II)-L-Histidinate complexes before irradiation at 7, 14, 30 days.\u003c/p\u003e\n\u003cp\u003eAs seen, the DNA from the healthy rat\u0026apos;s liver (norm, N) shows a homogeneous stain, corresponding to high molecular DNA with a size of 15kbp. In contrast, the DNA patterns of irradiated rats on days 7, 14, and 30 exhibit high electrophoretic mobility, indicating DNA fragmentation. The data clearly reveal the absence of DNA with high molecular weight on the electrophoresis tracks of irradiated rat DNA (Figure 4A, B, C marked 1).\u003c/p\u003e\n\u003cp\u003eDNA fragments with sizes of 1-9 kb, 0.5-8 kb, and 0.2-4 kb were detected at 7, 14, and 30 days of irradiation, respectively. DNA samples obtained from the liver of rats pretreated with Cu(II)-L-Histidinate complexes exhibit electrophoretic bands similar to healthy DNA. Thus, DNA with sizes of 11-14kbp, 11-15kbp, and 11-15kbp were detected in the livers of rats pre-treated with Cu(II)(Picolinyl-L-Histidinate)₂ and Cu(II)(Nicotinyl-L-Histidinate)₂ on days 7, 14, and 30, respectively (Figure 4A, B, marked 2).\u0026nbsp;For rats pre-treated with Cu(II)(Isonicotinyl-L-Histidinate)₂ (Figure 4C, marked 2), DNA bands with a size of 15kbp were detected on all investigated days.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThus, the chelating agents may protect DNA from hydroxyl radicals and prevent enzyme damage, which is responsible for the DNA enzymatic repair system. The efficiency of the chemical pathway of DNA repair has been studied in the works of Morozova \u003cem\u003e(18, 19).\u003c/em\u003e In this works, amino acids were considered as electron donors, providing the reduction of radicals of the guanosine purine base.\u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eIonizing radiation has sufficient energy to induce free radicals in biological molecules, DNA being a particularly important target. Free radicals lead to the formation of potentially lethal DNA damages (\u003cspan class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan class=\"CitationRef\"\u003e21\u003c/span\u003e). The development of novel and effective approaches using non-toxic radioprotectors is of considerable interest for defense of nuclear industries, radiation accidents, and especially in medicine, in tumor radiotherapy (\u003cspan class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan class=\"CitationRef\"\u003e22\u003c/span\u003e).\u003c/p\u003e\n\u003cp\u003eIt is known that compounds containing heterocycles have high chemotherapeutic properties and serve as the basis for the development of new drugs. Many heterocyclic compounds are used in clinical practice for the treatment of infectious diseases. For example, nitrogen compounds containing an imidazole fragment easily interact with active centers in living organisms and have a unique position in the chemistry of heterocycles (\u003cspan class=\"CitationRef\"\u003e23\u003c/span\u003e, \u003cspan class=\"CitationRef\"\u003e24\u003c/span\u003e).\u003c/p\u003e\n\u003cp\u003eThe radioprotective effects of the Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e, Cu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e and Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e compounds were assessed by determining the survival of rats, pre-treated with investigated complexes and irradiated at 6.5Gy dose of X-rays. According to the results obtained, the Cu(II) L-Histidinate Schiff base complexes had a strong radioprotective effect when administered orally and subcutaneously to rats under X-ray exposure at a dose of 6.5Gy.\u003c/p\u003e\n\u003cp\u003eThe used radiation dose corresponded to LD\u003csub\u003e60\u003c/sub\u003e, i.e. the survival rate of rats was 40%. Pre-treatment of animals with the studied complexes increased the survival rate of rats during the 30 days after irradiation to 70\u0026ndash;100%.\u003c/p\u003e\n\u003cp\u003eThe previous report (\u003cspan class=\"CitationRef\"\u003e25\u003c/span\u003e) presents the evaluation of the cytogenetic state of bone marrow cells in rats surviving to day 30 after irradiation at 6.5Gy. In irradiated rats, compared with the norm, there was an increase in the number of chromosomal aberrations to 7.7% and the number of tetraploid chromosomes to 2.1% (3.1% and 0.2% in the norm). However, in rats pre-treated with the Cu(II) Histidinate Schiff base complexes, there was a tendency toward the normalization of these parameters.\u003c/p\u003e\n\u003cp\u003eStructural changes of the DNA molecule were identified using the DNA melting method, because since this method is widely used in identifying damages in DNA molecule (\u003cspan class=\"CitationRef\"\u003e26\u003c/span\u003e, \u003cspan class=\"CitationRef\"\u003e27\u003c/span\u003e). The melting parameters of DNA isolated from the liver of irradiated rats on the 3, 7, 14 and 30 days showed deviations from normal values, which indicated structural changes in the DNA molecule. Based on the DNA melting parameters, we can conclude that damages of the DNA molecule increased during the studied periods of the experiment. The presence of oxygen radicals induced by ionizing radiation increased DNA damages on 30 day, which contributed to the death of animals. The melting parameters of DNA isolated from rats pre-treated with Cu(II) L-histidinate-Schiff base complexes on the same (i.e. 3, 7, 14 and 30) days of the experiment significantly improved and approached to normal. The studied compounds demonstrated radiotherapeutic effects and ensured the stability of the secondary structure of rat liver DNA from damages.\u003c/p\u003e\n\u003cp\u003eOur electrophoretic data correspond well to the literature, which showed the detection of various fragments with strand breaks in DNA samples obtained from irradiated animals (\u003cspan class=\"CitationRef\"\u003e28\u003c/span\u003e). Free radical processes appear to be the primary cause of radiation damage to DNA. The activation level of free-radical processes significantly increases over time. As observed, DNA damage in irradiated samples also increases over time. At the same time, there are published data indicating that the activation of endogenous nucleases occurs during X-ray irradiation, leading to the accumulation of single- and double-strand breaks in DNA (\u003cspan class=\"CitationRef\"\u003e29\u003c/span\u003e). The results clearly show that DNA isolated from animals pre-treated with the Cu(II) chelates exhibits electrophoretic bands similar to healthy DNA.\u003c/p\u003e\n\u003cp\u003eOur results show that the presence of a metal in the investigated complexes plays an important role in ensuring their radioprotective properties. Preliminary studies of the radioprotective properties of Schiff bases derived from L-Histidine and 2-, 3-, and 4-pyridine carboxaldehyde showed weak radioprotective activity at irradiation dose of 6.5Gy (\u003cspan class=\"CitationRef\"\u003e25\u003c/span\u003e). However, at complexation with Cu(II) Histidinate Schiff base metal complexes became effective radiation-protective agents capable of reducing or preventing radiation-induced lethality in rats. Perhaps this is due to the fact that the main enzymes involved in reactions that prevent the accumulation of toxic metabolic products are metal-dependent. They promote biochemical, cellular and tissue restoration, and also provide recovery after radiation injury. In this regard, copper cations are probably the most promising (\u003cspan class=\"CitationRef\"\u003e30\u003c/span\u003e, \u003cspan class=\"CitationRef\"\u003e31\u003c/span\u003e).\u003c/p\u003e\n\u003cp\u003eTrace elements such as Cu(II) in Schiff base form have a protective effect against cellular toxicity caused by oxidative stress, especially with superoxide dismutase mimetic activity. It can inhibit oxidative stress and protect the DNA against peroxide radicals. In previous work, the activity of the antioxidant enzymes superoxide dismutase and catalase was determined on 3, 7, 14, and 30 days after irradiation at a dose of 6.5Gy in the blood of animals. The results of the studies revealed superoxide dismutase and catalase mimetic activity of Cu(II)L-Histidinate Schiff base complexes (\u003cspan class=\"CitationRef\"\u003e25\u003c/span\u003e). The superoxide dismutase and catalase mimetic activity of the copper organic complexes have an important effect on the body\u0026apos;s defense against radiation-induced oxidative stress.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eNewly synthesized compounds Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e, Cu(II)(Nicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e and Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e are effective radiation protective agents capable to reduce or prevent radiation-induced lethality in rats.\u003c/p\u003e\n\u003cp\u003eAll three metal complexes demonstrate significant radiation protection through both subcutaneous and oral administration to rats. Moreover, 100% survival was registered in rats irradiated on the background oral administration of 40mg/kg Cu(II)(Picolinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e 24 hours prior to irradiation. The same result, i.e. 100% survival, was achieved in the case of subcutaneous injection of 10mg/kg Cu(II)(Isonicotinyl-L-Histidinate)\u003csub\u003e2\u003c/sub\u003e to rats one hour before exposure at the dose of 6.5Gy. A study of DNA melting curves and electrophoresis data revealed improved characteristics of rat liver DNA in animals pretreated with Cu(II) Schiff base complexes before to X-ray exposure.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e \u003ch2\u003eConflict of interest.\u003c/h2\u003e \u003cp\u003eThe authors declare that they have no conflict of interest.\u003c/p\u003e \u003ch2\u003eStatement of Human and Animal Rights\u003c/h2\u003e \u003cp\u003e Experiments were fulfilled according to the \"International Recommendations on carrying out of biomedical Research with use of Animals\" (CIOMS, 1985), to the \"Human Rights and Biomedicine, Oviedo Convention\" (CE, 1997), to the European Convention for the Protection of Vertebral Animals Used for Experimental and Other Scientific Purposes (CE, 2005) and approved by the National Center of Bioethics (Armenia).\u003c/p\u003e \u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eN. K. contributed in the execution of the experiments, data analysis, investigation, design, , writing and drafting the original manuscript. All experimental data was analyzed by N. K., G. A. and S. H. and contributed to the article review and editing. Reagent preparation and experimentation were carried out by N. K. and G. A., analysed and reviewed the manuscript. The manuscript was critically revised for accurate knowledgeable content by S. H. Finally, all authors approved the final version of the manuscript for publication.\u003c/p\u003e\u003ch2\u003eAcknowledgement\u003c/h2\u003e\u003cp\u003eThe authors thank Dr. M.H Malakyan, Dr. V. Matosyan and Dr. V.Tonoyan for their assistance in the work\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eRastogi RP,Richa,Kumar A, Tyagi MB, Sinha RP\u003cstrong\u003e.\u003c/strong\u003e (2010) Molecular Mechanisms of Ultraviolet Radiation-Induced DNA Damage and Repair. J Nucleic Acids 2010. Published online, https://doi.org/10.4061/2010/592980\u003c/li\u003e\n\u003cli\u003eRiley PA. (1994) Free radicals in biology: Oxidative stress and the effects of ionizing radiation. International Journal of Radiation Biology 1994. 65, 27-33. https://doi.org/10.1080/09553009414550041\u003c/li\u003e\n\u003cli\u003eSingh SK, Wang M, Staud CT, Iliakis G. (2011) Post-irradiation chemical processing of DNA damage generates double-strand breaks in cells already engaged in repair. Nucleic Acids Res 2011. 39, 8416\u0026ndash;8429. https://doi.org/10.1093/nar/gkr463. \u003c/li\u003e\n\u003cli\u003eShikazono N, Noguchi M, Fujii K, Urushibara A, Yokoya A. (2009) The yield, processing, and biological consequences of clustered DNA damage induced by ionizing radiation. Journal of Radiation Research 2009. 50, 27\u0026ndash;36. https://doi.org/10.1269/jrr.08086 .\u003c/li\u003e\n\u003cli\u003eSutherland BM, Bennett PV, Sidorkina O, Lava J. 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Structural Chemistry. 2011. 22, 475-482. https://doi.org/10.1007/s11224-011-9765-4\u003c/li\u003e\n\u003cli\u003eKarapetyan NH, Malakyan MH, Bajinyan SA, Torosyan AL Grigoryan IE, Haroutiunian SG. (2013) Influence of Amino Acids Shiff Bases on Irradiated DNA Stability In Vivo. Cell Biochem Biophys. 2013. 67, 1137\u0026ndash;1145. https://doi.org/10.1007/s12013-013-9617-5\u003c/li\u003e\n\u003cli\u003eKarapetyan NH, Torosyan AL, Malakyan MH, Bajinyan SA, Haroutiunian SG. (2016) Investigation of irradiated rats DNA in the presence of Cu(II) chelates of amino acids Schiff bases. Journal of Biomolecular Structure and Dynamics. 2016. 34, 177-183. http://dx.doi.org/10.1080/07391102.2015.1020876\u003c/li\u003e\n\u003cli\u003eJomova K, Makova M, Alomar SY, Alwasel SH, Nepovimova E, Kuca K et al. (2022) Essential metals in health and disease. Chemico-Biological Interactions. 2022. 367, 110173. https://doi.org/10.1016/j.cbi.2022.110173\u003c/li\u003e\n\u003cli\u003eMalakyan МН, Bajinyan SA, Matosyan VH, Tonoyan VJ, Babayan KN. (2016) Synthesis, characterization and toxicity studies of pyridinecarboxaldehydes and L-tryptophan derived Schiff bases and corresponding copper (II) complexes. F1000 Research, 2016. 5, 1921. doi:10.12688/f1000research.9226.1\u003c/li\u003e\n\u003cli\u003eLando DY, Teif VB. (2002) Modeling of DNA Condensation and Decondensation Caused by Ligand Binding. Journal of Biomolecular Srtucture and Dynamics. 2002. 20, 215-222. https://doi.org/10.1080/07391102.2002.10506837 \u003c/li\u003e\n\u003cli\u003eWells RD, Larson JE, Grant RC, Shortle BE, Cantor CR. (1970) Physicochemical studies on polydeoxyribonucleotides containing defined repeating nucleotide sequences. Journal of Molecular Biology. 1970. 54, 465-497. doi:10.1016/0022-2836(70)90121-x.\u003c/li\u003e\n\u003cli\u003eWartell RM, Benight AS. (1985) Thermal denaturation of DNA molecules: A comparison of theory with experiment. Physics Reports. 1985. 126, 67\u0026ndash;107. https://doi.org/10.1016/0(370-1573(85)90060-2\u003c/li\u003e\n\u003cli\u003e Lee PY, Costumbrado J, Hsu C-Y, Kim YH. (2012) Agarose Gel Electrophoresis for the Separation of DNA Fragments. J Vis Exp. 2012. 62, 3923. doi: 10.3791/3923 \u003c/li\u003e\n\u003cli\u003eVologodskii A, Frank-Kamenetskii MD. (2018) DNA melting and energetics of the double helix. Physics of Life Reviews. 2018. 25, 1-21. https://doi.org/10.1016/j.plrev.2017.11.012\u003c/li\u003e\n\u003cli\u003eMorozova OB, Kiryutin AS, Sagdeev RZ, Yurkovskaya AV. (2007) Electron Transfer between Guanosine Radical and Amino Acids in Aqueous Solution. 1. Reduction of Guanosine Radical by Tyrosinev J. Phys. Chem. B, 2007. 111, 7439\u0026ndash;7448. DOI: 10.1021/jp067722i\u003c/li\u003e\n\u003cli\u003eMorozova OB, Kiryutin AS, Yurkovskaya AV. 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DOI: 10.1080/09553000500077211 \u003c/li\u003e\n\u003cli\u003eBakayev VV, Yugai AA, Luchnik AN. (1985). Effect of X-ray induced DNA damage on DNAase I hypersensitivity of SV40 chromatin: relation to elastic torsional strain in DNA. Nucleic Acids Research, 1985; 13, 7079\u0026ndash;7093. DOI: 10.1093/nar/13.19.7079 \u003c/li\u003e\n\u003cli\u003e Zoroddu MA, Aaseth J, Crisponi G, Medici S, Peana M, Nurchi VM. (2019) The essential metals for humans: a brief overview. J. Inorg Biochem. 2019. 195, 120-129. https://doi.org/10.1016/j.jinorgbio.2019.03.013 \u003c/li\u003e\n\u003cli\u003eSorenson JRJ. (2002). Cu, Fe, Mn, and Zn chelates offer a medicinal chemistry approach to overcoming radiation injury. Current Med. Chem. 2002. 9, 639-662. doi: 10.2174/0929867023370725\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"cell-biochemistry-and-biophysics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"cbbi","sideBox":"Learn more about [Cell Biochemistry and Biophysics](http://link.springer.com/journal/12013)","snPcode":"12013","submissionUrl":"https://submission.nature.com/new-submission/12013/3","title":"Cell Biochemistry and Biophysics","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Irradiation, DNA melting, Electrophoresis, Schiff base complexes.","lastPublishedDoi":"10.21203/rs.3.rs-4387030/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4387030/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eA study of rats liver DNA damages under the influence of X-ray radiation at a dose of 6.5 Gy(LD60) was carried out. The radioprotective properties of newly synthesized Cu(II) L-Schiff Histidinate\u0026nbsp; complexes\u0026nbsp; were also studied.\u0026nbsp; The survival of rats was determined over a 30-day period after exposure to X-rays without pretreatment and also after preadministration of Cu(II) L-Histidinate-Schiff base complexes. The structural defects of rat's liver DNA were detected at 3, 7, 14, and 30 days post-irradiationxtracted. The results obtained revealed that irradiation with a 6.5Gy dose in the control group degraded the characteristics of rat liver DNA in comparison to healthy DNA. On all investigated experimental days, a decrease in the melting temperature (T\u003csub\u003em\u003c/sub\u003e), a widening of the melting interval (ΔT), and a decrease in hypochromicity (Δh) were observed in the DNA samples of irradiated animals compared to the norm. The rat's pretreatment by Cu(II) L-Histidinate complexes 1 or 24 hours prior to irradiation improved DNA characteristics. Electrophoretic studies of DNA were in good agreement with the melting data. Based on the study results, it can be concluded that Cu(II) L-Histidinate complexes exhibit radioprotective properties\u0026nbsp; under the studied conditions and can protect DNA from damage.\u003c/p\u003e","manuscriptTitle":"Influence of Cu L-Histidinate Schiff base derivatives on structural features of irradiated rat’s DNA","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-05-16 18:08:58","doi":"10.21203/rs.3.rs-4387030/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-05-29T01:02:40+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-05-29T00:34:42+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-05-13T19:00:46+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"248063086826268558395162112072835857756","date":"2024-05-11T09:54:54+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"164217249833290325562058210502192160123","date":"2024-05-11T07:46:27+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"293872306412885040025720436913465786603","date":"2024-05-09T03:56:05+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"15598406218695782766542681213357037928","date":"2024-05-09T01:44:15+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"118680529306993029811522543386331665692","date":"2024-05-08T20:00:34+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-05-08T19:31:32+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-05-08T14:06:39+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-05-08T14:06:39+00:00","index":"","fulltext":""},{"type":"submitted","content":"Cell Biochemistry and Biophysics","date":"2024-05-08T06:40:14+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"cell-biochemistry-and-biophysics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"cbbi","sideBox":"Learn more about [Cell Biochemistry and Biophysics](http://link.springer.com/journal/12013)","snPcode":"12013","submissionUrl":"https://submission.nature.com/new-submission/12013/3","title":"Cell Biochemistry and Biophysics","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"d7cbe9c6-eddd-4323-ad20-7124c8792a57","owner":[],"postedDate":"May 16th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2024-06-13T18:38:24+00:00","versionOfRecord":[],"versionCreatedAt":"2024-05-16 18:08:58","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4387030","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4387030","identity":"rs-4387030","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}