SYNTHESIS AND EVALUATION OF THE ANTIOXIDANT ACTIVITY OF {[1-ARYL-4-CHLORO-1 H -IMIDAZOLE-5-YL) METHYL]THIO}ALKANE CARBOXYLIC ACIDS

This study is devoted to development of the optimal conditions for synthesis and the study of some “structure – antioxidant activity” regularities of [(1-arylimidazole-5-yl)methylthio]alkane carboxylic acids, which structural analogues have found an application as medicinal products with a wide range of biological activities. The methodology of interaction between 4-cloro-5-chloromethylimidazoles with thioglycolic and thiopropionic acids has been used to obtain these compounds. Selection of the optimal reaction conditions has allowed to obtain target compounds in a dry dimethylformamide in the presence of potash at 50 о C with yields of 75-82%. The compounds synthesized are high-melting crystalline substances that dissolve well in polar organic solvents and aqueous alkaline solutions. Their composition and structure have been confirmed by the results of elemental analysis and mea surement data of IR-, 1 H NMR- and chromatography mass-spectra. The study of the compounds synthesized has been conducted in vitro on biological samples. The antioxidant activity has been determined by the inhibition value of the ascorbate-dependent endogenous lipid peroxidation rate in rats’ liver found by the concentration of one of the final products of free-radical lipid oxidation processes – malonaldehyde in the test sample. The results of the biological activity screening of the compounds synthesized show that all imidazole derivatives studied in the final concentration ranges of 10 -3 -10 -1 M exhibit a high antioxidant action in the system in vitro. It has been found that the value of the antioxidant activity is influenced by the nature and position of the substituent in position 1 of imidazole . In particular, the presence of electron-acceptor substituents in the aryl fragment decreases the molecule activity in comparison with electron-donor substituents, wherein increase of the methylene groups quantity in the carboxyalkylthiol fragment does not significantly impact the antioxidant effect of the compounds synthesized.

The process of free-radical lipid oxidation (FRLO) plays a significant role in development of the most diseases of the liver, cardiovascular, respiratory and nervous systems [1,16]. Antioxidants are widely used to normalize the basic organism functions as a part of complex therapy of such diseases [7]. It should be noted that at present there is insufficient amount of medicines with the antioxidant mechanism of action offered for clinical use, and those that are in use have many adverse effects and high toxicity [4]. Taking this into account, the search of compounds that are able to inhibit the FRLO processes effectively is of high interest nowadays.
Design and synthesis of new compounds with antioxidant properties is a subject of many studies as they are of high interest in prophylactics and therapy of many diseases that have FRLO in their pathogenesis [3,9]. From this aspect the derivatives of imidazole are not left aside too; according to the literature data they are characterized by a wide spectrum of pharmacological properties, among which the antioxidant effect is particularly noteworthy [10,11,15].
Earlier we found the antioxidant activity in the series of [(1-phenyl-5-formyl-1H-imidazole-4-yl)thio]acetic acids and their derivatives [5,6,8,13]. It was to be expected that the change in the position of the thioalkanecarboxylic acid fragment in the structure of the imidazole cycle would allow a much bigger quantity of potentially active compounds with the antioxidant action, and also give an opportunity to approach to the solution of the topical problem of pharmaceutical chemistry -establishing of the structure -activity relationship. With this purpose we have synthesized [(1-arylimidazole-5-yl)methylthio]alkanecarboxylic acids (2а-g) and carried out the screening of their antioxidant properties.
The reaction of easily available 4-chloro-5-chlorometylimidazoles (1а-d) [12] with thioglycolic and thiopropionic acids, which occurs selectively at 50 о С in a dry DMFA in the presence of potash, has been used to obtain compounds of such kind. The reaction gives the target compounds (2а-g) with yields of 75-82%.
The compounds (2а-g) synthesized (Tab. 1, 2) are light-yellow, high-melting crystalline compounds, readily soluble in polar organic solvents and aqueous alkaline solutions. Their composition and structure were confirmed by elemental analysis and by the results of IR-, 1 H NMR and chromatography mass-spectra measurements. IR-spectra, in particular, are characterized by intensive absorption bands of carbonyl groups at 1675-1685 cm -1 and by a wide absorption range (2430-2850 cm -1 ) of carboxyl groups, and it indicates a dimeric character of the acids obtained in their solid state. In 1 H NMR spectra of all compounds the most illustrative are H 2 proton singlets of the imidazole cycle at 7.83-7.95 ppm and the singlets of methylene groups bound with the imidazole cycle at 3.72-3.85 ppm. In their turn, the methylene protons of thioacetic acid fragments are visible as singlets at 3.17 ppm, and those of thiopropionic acid -as triplets in the range of 2.30-2.35 and 2.49-2.54 ppm.

Experimental Part (Biology)
The research of the antioxidant activity of the compounds (2а-g) synthesized was performed in vitro [14] and determined by the inhibition value of the ascorbate-dependent endogenous lipid peroxidation rate in rats' liver found by the concentration of one of the final products of free-radical lipid oxidation processesmalonaldehyde (MA) in the test sample. The content of MA was determined by the reaction with thiobarbituric acid (TBA) and was calculated in μmoles/g of the tissue. Statistical analysis of the results obtained was performed using the parametric Student's t-test [2]. The value of ascorbate-induced FRLO inhibition was calculated in percents, with MA concentration in the control samples being equal to 100%.
The range of concentrations of the compounds synthesized was chosen within the limits of concentrations already researched for their structural analogue, thiotriazoline (Т), (the manufacturer is "Arterium" corporation, Ukraine, solution for injections, 25 mg/ml), which has the proven antioxidant activity [7].

Results and Discussion
The results of the antioxidant activity screening of the compounds synthesized in vitro (Tab. 3) show that all compounds are able to inhibit Fe 2+ -ascorbate initiated FRLO in these conditions in the final concentration ranges of 10 -3 -10 -1 M studied. Thus, the degree of The highest antioxidant effect in vitro was recorded for compound 2d in the range of the final concentrations of 5·10 -3 -10 -2 M: the degree of FRLO inhibition was 76.47-76.25%. On average, it is 43% higher than the results shown by thiotriazoline in the same range of the final concentrations, and 37% higher than the maximum effect of thiotriazoline recorded in vitro.
Analysis of the data obtained shows that the value of the antioxidant activity is influenced by the nature and position of the substituent in position 1 of imidazole. In particular, the presence of electron-acceptor substituents in the aryl fragment decreases the molecule activity in comparison with electron-donor substituents, wherein increase of the methylene groups quantity in the carboxyalkylthiol fragment does not significantly impact the antioxidant effect of the compounds synthesized. CONCLUSIONS 1. By interaction of 5-chlorometylimidazoles with thioglycolic and thiopropionic acids new {[(1-Aryl-4- Table 3 The chloro-1Н-imidazole-5-yl)methyl]thio}alkanecarboxylic acids have been synthesized. 2. All derivatives of imidazole studied in the range of concentrations of 10 -3 -10 -1 М show a high antioxidant activity in the system in vitro. The highest activity was recorded for compound 2d in the final concentration of 5·10 -3 M.