Int J Pharm Pharm Sci, Vol 9, Issue 8, 154-160Original Article


SYNTHESIS AND ANTIDEPRESSANT ACTIVITY OF 2-BROMO-1-(THIETAN-3-YL) IMIDAZOLE-4, 5-DICARBOXYLIC ACID DERIVATIVES

FERKAT KHALIULLIN*1, IRINA NIKITINA2, ANFISA VALIEVA1, ALBINA MIFTAKHOVA2, EKATERINA MESCHERYAKOVA3, LEONARD KHALILOV3

1Department of Pharmaceutical Chemistry, Bashkir State Medical University, Lenina Str., 3, 450008, Ufa, Russia, 2Department of Pharmacology № 1, Bashkir State Medical University, Lenina Str., 3, 450008, Ufa, Russia, 3Laboratory of Structural Chemistry, Institute of Petrochemistry and Catalysis, Russian Academy of Science, Prospekt Oktyabrya, 141, 450075, Ufa, Russia
Email: khaliullin_ufa@yahoo.com

Received: 09 Feb 2017 Revised and Accepted: 30 Jun 2017

ABSTRACT

Objective: Synthesis of the salts and diylidenehydrazidеs of 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid to evaluate the antidepressant activities.

Methods: The structures of the synthesised compounds were confirmed by elemental analysis and 1Н NMR spectral data. The melting points of the compounds were determined on a Stuart SMP30 apparatus. The X-ray diffraction data for compound IIc were obtained at room temperature on a Xcalibur Gemini Еos. The antidepressant activity was investigated in the tail suspension and forced swimming tests. The locomotor activity and anxiety were studied in the open field test.

Results: All synthesised compounds showed antidepressant activity after single intraperitoneal injection to male mice at doses equimolar to 10 mg/kg of imipramine. One of the compounds, 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid di[(4-hydroxy-3-methoxyphenyl) methylidenehydrazide], reduced the anxiety and decreased the locomotor activity at statistically significant levels. Other compounds did not have sedative and/or stimulating effects.

Conclusion: Among the synthesised 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid derivatives, compounds with marked antidepressant activity were identified. An obvious advantage of these products is low toxicity.

Keywords: Imidazole-4,5-dicarboxylic acid, Thietane, Diylidenehydrazidеs, Antidepressant activity

© 2017 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)
DOI: http://dx.doi.org/10.22159/ijpps.2017v9i8.17613

INTRODUCTION

Imidazole derivatives represent a class of drugs widely used in medicine [1]. Nowadays, search for biologically active compounds among derivatives of imidazole, in particular, in a series of imidazole-4,5-dicarboxylic acids are in progress. The compounds with antiviral activity have been found among imidazole-4,5-dicarboxylic acid derivatives [2, 3]. The hydrazinium salt of imidazole-4,5-dicarboxylic acid shows an antibacterial effect [4]. 1-Methylimidazole-4,5-dicarbohydrazide is a monoamine oxidase inhibitor [5].

To find new biologic active compounds among 2-bromoimidazole-4, 5-dicarboxylic acid derivatives, the target activity forecast was performed with SARD-21 software. The study showed that 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylic acid salts and diylidenehydrazidеs may have an antidepressant effect. Depression refers to socially significant diseases, which is due to their high prevalence [6]. The study of the antidepressant activity of known drugs [7] and the search for new sources of antidepressant drugs are continuing [8]. The aim of this study was to synthesise and study the antidepressant activity of 2-bromo-1-(thietan-3-yl) imidazole-4, 5-dicarboxylicacid derivatives.

MATERIALS AND METHODS

Materials

1Н NMR spectra were recorded on a BrukerAM-300 instrument operating at 300 MHz. The residual solvent signals were used as internal standards. TLC was carried out on Sorbfil plates with a 4:1:2 1-butanol–acetic acid–water (IIb, IIc) or 1:1 dioxane–ethanol (IVb) mixture or with dioxane (IVа) as the mobile phase. The melting points of the compounds were determined on a Stuart SMP30 apparatus. The X-ray diffraction data for compound IIc were obtained at room temperature on a Xcalibur Gemini Еos diffractometer equipped with an EOS CCD three-dimensional array detector and a monochromated MoKα radiation (graphite monochromator, MoKα radiation, λ = 0.71073 Å, ω-scan mode, 2θmax = 62 °). The data were collected and treated using CrysAlisPr ° Oxford Diffraction Ltd. software, Version 1.171.36.20 [9]. The crystal of compound IIc (C8H6Br1K1N2O4S1, M = 345.21) is monoclinic. The unit cell parameters are a = 5.4769(4) Å, b = 16.8316(11) Å, c = 12.3235(10) Å, β = 101.291(7) °, V = 1114.05(14) Å3; space group P21/n (no. 14); Z = 4; Dcalcd. = 2.058 mg/mm3. The final R-factors are as follows: R1 = 0.0579 for 1914 collected unique reflections with I>2σ(I) and wR2 = 0.1709 for 2683 unique reflections. The structure was solved by the direct method and refined by the full-matrix least-squares method in the anisotropic approximation for non-hydrogen atoms. The hydrogen atoms were located in a difference Fourier synthesis and refined in the isotropic approximation. The calculations were carried out using the SHELX [10]. The CIF-file is deposited with the Cambridge crystallographic data centre, CCDC No 1452164. A copy of these data is available free of charge on request from CCDC, 12, Union Road, Cambridge, CB2 1EZ, UK (fax: +44 1223 336033, e-mail: deposit@ccdc. cam. ac. uk) or via http://www. ccdc. cam. ac. uk/data_request/cif.

Compounds I and III were synthesised by procedures described in [11].

Compounds IIa and IId-f were synthesised by procedures described in [12].

Dipotassium 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylate (IIb)

Potassium hydroxide (, 12 mmol) and compound I (, 3 mmol) were dissolved in a mixture of 10 ml of water and 25 ml of ethanol. The reaction mixture was stirred with a magnetic stirrer at room temperature for 5 h. The precipitate was filtered off, washed with ethanol, and dried. The product was purified by re-precipitation with acetone from an aqueous solution to give (78%) of compound IIb. Mp ( °C): 270 (decomp); Rf: 0.54; 1H NMR (D2О): 3.47-3.53 (m, 2Н, S(СН)2), 4.34-4.41 (m, 2Н, S(СН)2), 5.87-5.96 (m, 1Н, NCH) ppm; CHN analysis for C8H5N2O4Br1S1K2; C 25.07; H 1.31; N 7.31. Found C 25.41; H 1.30; N 7.28.

Potassium 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylate (IIc)

Compound IIb (, 1.6 mmol) was dissolved in 4 ml of water, and the solution was acidified with dilute HCl to рН=2-3. The precipitate was filtered off, washed once with water, and dried. The product was recrystallized from water to give (56%) of compound IIc. Mp ( °C): 250 (decomp); Rf: 0.54; 1H NMR (DMSO-d6): 3.29-3.34 (m, 2Н, S(СН)2), 4.30-4.36 (m, 2Н, S(СН)2), 7.10-7.22 (m, 1Н, NCH) ppm; CHN analysis for C8H6Br1N2O4S1K1; C 27.83; H 1.75; N 8.11. Found C 27.70; H 1.70; N 7.96.

2-Bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid di[(4-dimethylaminophenyl) ­methylidenehydrazide] (IVа)

Compound III (, 1.2 mmol) was dissolved in 2 ml of dilute HCl and 20 ml of ethanol, and 4-(dimethylamino) benzaldehyde (, 3.6 mmol) in 5 ml of ethanol was added. The reaction mixture was stirred for 10 min at room temperature. The precipitate was filtered off, washed with ethanol, and dried. The product was recrystallized from benzene to give (78%) of compound IVа. Mp ( °C): 171-173 (decomp); Rf: 0.62; 1H NMR (СDCl3): 3.38-3.44 (m, 2Н, S(СН)2), 4.44-4.50 (m, 2Н, S(СН)2), 7.15-7.25 (m, 1Н, NCH), 3.00 (s, 6Н, N(CH3)2), 3.02 (s, 6Н, N(CH3)2), 6.68 (d,J = 8.1 Hz, 4Н, Ar-Н), 7.66-7.70 (m, 4Н, Ar-Н), 8.09 (s, 1Н, =СН), 8.27 (s, 1Н, =СН), 10.36 (s, 1Н, NH), 14.23 (s, 1Н, NH) ppm; CHN analysis for C26H29Br1N8O2S1; C 52.26; H 4.89; N 18.75. Found C 52.10; H 4.87; N 18.78.

2-Bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid di[(4-hydroxy-3-methoxyphenyl)­methylidenehydrazide] (IVb)

Compound III (, 1.0 mmol) was dissolved in 2 ml of dilute HCl and 10 ml of ethanol, and 3-methoxy-4-hydroxybenzaldehyde (, 3.0 mmol) in 5 ml of ethanol was added. The reaction mixture was stirred for 10 min at room temperature. The precipitate was filtered off, washed with ethanol and acetonitrile, and dried. The product was recrystallized from a 2:1 dimethylformamide–water mixture to give (98%) of compound IVb. Mp ( °C): 243-244; Rf: 0.62; 1H NMR (DMSO-d6): 3.38-3.45 (m, 2Н, S(СН)2), 4.21-4.27 (m, 2Н, S(СН)2), 6.00-6.12 (m, 1Н, Z-NCH), 5.72-5.78 (m, 1Н, E-NCH), 3.81 (s, 3Н, Z-ОСН3), 3.66 (s, 3Н, Е-ОСН3), 3.86 (s, 3Н, Z-ОСН3), 3.79 (s, 3Н, Е-ОСН3), 6.82-6.88 (m, 2Н, Z-Ar-Н), 6.70-6.73 (m, 2Н, Е-Ar-Н), 7.04 (d,J = 8.2 Hz, 1Н, Z-Ar-Н), 7.16 (d,J = 8.1 Hz, 1Н, Z-Ar-Н), 7.38 (s, 1Н, Z-Ar-Н), 7.28 (s, 1Н, Z-Ar-Н), 7,21 (s, 1Н, Е-Ar-Н), 8.18 (s, 1Н, Z-=СН), 7.96 (s, 1Н, Е-=СН), 8.47 (s, 1Н, Z-=СН), 8.37 (s, 1Н, Е-=СН), 9.58 (s, 1H, Z-OH), 9.48 (s, 1Н, Е-ОН), 9.66 (s, 1H, Z-OH), 9.51 (s, 1Н, Е-ОН), 11.74 (s, 1Н, Z-NH), 11.59 (s, 1Н, Е-NH), 12.51 (s, 1Н, Z-NH), 12.22 (s, 1Н, Е-NH) ppm; CHN analysis for C24H23Br1N6O6S1; C 47.77; H 3.84; N 13.93. Found C 47.49; H 3.79; N 13.75.

Compound IVc was synthesised by the procedure described in [13].

Antidepressant activity

The present study evaluates the antidepressant-like activity of the compounds. The protocol was approved by the Committee on the Ethics of Animal Experiments of the Bashkir State Medical University. 173 white non-inbred male mice weighing 20 to were used in the study. The animals were randomly allocated into groups. Housing and handling of animals were performed according to the Guidelines for housing and breeding of laboratory animals in nurseries and experimental-biological clinics (vivariums) and their use in scientific, educational and production purposes, approved by the RAMS and the Ministry of Health of RF 03.04.2003. Ten thietane-containing derivatives of 2-bromoimidazole-4,5-dicarboxylic acid (I, IIa–f, IVа–c) were studied.

The antidepressant activity was investigated in the tail suspension test (TST) [14] and forced swimming test (FST) [15]. These animal tests are used more widely to predict the antidepressant action. The exploratory activity, locomotor activity, and anxiety were studied in the open field test (OF) [16], which evaluates the animal behavior in a novel environment.

The animal behavior analysis was performed with the Brain Test software [17]. The total immobility time was evaluated in the TST and FST. Additionally, the index of depression was calculated using the FST results. The index of depression is the ratio of short periods of immobilisation (less than 6 s) to the number of periods of active swimming. Individual behavior of animals in the OF test was considered as the set of discrete behavioral acts and postures: “moving”, “sniffing”, “rearing”, “grooming”, “movement on the spot”, “hole”, “recline against wall”, “sitting”, and “defecation”. The normality of behaviour, exploratory activity (the sum of the patterns “sniffing”, “moving”, and “hole”) and anxiety (the sum of the patterns “moving on the spot”, “recline against the wall”, and “rearing”) were considered as integral criteria.

The compounds were administered to the animals once intraperitoneally (i. p.) at doses equimolar to imipramine (Egis Pharmaceuticals PLC, Hungary) used as the reference drug (10 mg/kg) 30 min before the start of the test. Before administration, the compounds were suspended with 1-2 drops of TWIN- water for injections. Imipramine (the reference drug) was injected i. p. once 30 min before the test at the optimally effective animal dose of 10 mg/kg. The control animals received equivalent amounts of saline with TWIN-80.

Statistical analysis was performed with Statistica 7.0. Descriptive statistics included median (Me), 25 and 75 percentiles (Per). Kruskall-Wallis test and Wilcoxon-Mann-Whitney test were used for group comparison. The statistical analysis was performed at the significance level of 5% [18]. The acute toxicity (LD50) of the compounds was studied with adult non-inbred male mice, according to the method of Litchfield J. and Wilcoxon F. [19] in the modification of Prozorovskiy V. B. [20]. The compounds were suspended ex tempore and injected once i. p.

RESULTS AND DISCUSSION

Chemistry

The disodium (IIa) [12] and dipotassium (IIb) 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylates were synthesized by the reaction of dimethyl 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylate (I) with sodium or potassium hydroxide (Scheme 1). The monopotassium salt of 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid (IIc) was prepared by acidifying an aqueous solution of dipotassium salt IIb with dilute hydrochloric acid to рН = 2-3. The structure of IIc was proved by 1Н NMR spectroscopy. The 1Н NMR spectrum of monopotassium salt IIc exhibiting multiplets for the two S(CH)2 groups in the ranges of 3.29-3.34 and 4.30-4.36 ppm and a multiplet for the NCH group in the range of 7.10-7.22 ppm. The 1Н NMR spectrum does not specify the location of the potassium cation; therefore, the structure of salt IIc is confirmed by single-crystal X-ray diffraction. Fig. 1 shows the monomeric fragment of the monopotassium salt IIc (fig. 1а) and the arrangement of molecules in the crystal, which reflects its sheetlike structure (fig. 1b). This compound crystallises in the monoclinic system with space group P21/n. There are several types of potassium atoms with different coordination environments of potassium, namely KN1O4S1, KN1O3, KО3, KO1S1, and KО2.

A number of specific features can be distinguished in the ligand structure. The thietane ring has a folded conformation, the dihedral angle between the С10С8С9 and С10S2C9 planes is 16.52(6) °. Note the presence of an intermolecular hydrogen bond, Br1…H8, with a length of 2.624 Å. The hydrogen atom Н1 is hypervalent (fig. 1), the О4–Н1 and О1–Н1 bond lengths being 1.174 and 1.232 Å, respectively (О–О = 2.402Å) (table 1, 2).



Fig. 1: (A) Monomeric fragment of IIc, (B) Sheet-like structure of IIc projected along а axis

Table 1: Bond lengths for compound IIc

Atom Atom Length/Å Atom Atom Length/Å
Br1 C2 1.880(5) O2 C4 1.200(7)
K1 K11 3.745(3) O3 K12 2.947(4)
K1 K12 4.653(2) O3 K11 2.788(4)
K1 S23 3.3973(19) O3 C5 1.238(7)
K1 O23 2.684(4) O4 K12 2.767(4)
K1 O31 2.788(4) O4 C5 1.278(6)
K1 O3 2.828(4) N3 K11 3.401(5)
K1 O32 2.947(4) N3 C2 1.287(7)
K1 O42 2.767(4) N3 C3 1.374(6)
K1 N31 3.401(5) N4 C2 1.373(7)
K1 N3 2.929(5) N4 C7 1.409(7)
K1 C52 3.147(5) N4 C8 1.488(7)
K1 C5 3.530(5) C3 C5 1.492(7)
S2 K14 3.3973(19) C3 C7 1.367(7)
S2 C9 1.841(6) C4 C7 1.510(7)
S2 C10 1.838(6) C5 K12 3.147(5)
O1 C4 1.311(7) C8 C9 1.524(8)
O2 K14 2.684(4) C8 C10 1.524(8)

Symmetry code: 1-X,1-Y,1-Z; 21-X,1-Y,1-Z; 3-1/2+X,1/2-Y,-1/2+Z; 41/2+X,1/2-Y,1/2+Z

Table 2: Bond angles for compound IIc

Atom Atom Atom Angle/˚ Atom Atom Atom Angle/˚
K11 K1 K12 80.63(4) N3 K1 C52 134.23(13)
S23 K1 K12 148.49(6) N31 K1 C5 92.61(12)
S23 K1 K11 116.35(6) C52 K1 K12 49.31(10)
S23 K1 N31 89.11(8) C5 K1 K12 42.54(9)
S23 K1 C5 168.95(10) C52 K1 K11 110.48(11)
O23 K1 K11 132.11(11) C5 K1 K11 58.22(9)
O23 K1 K12 126.50(10) C52 K1 S23 99.18(10)
O23 K1 S23 63.08(9) C52 K1 N31 77.56(13)
O23 K1 O32 108.09(13) C52 K1 C5 91.85(13)
O23 K1 O31 103.67(13) C9 S2 K14 108.4(2)
O23 K1 O3 130.91(12) C10 S2 K14 86.8(2)
O23 K1 O42 109.67(13) C10 S2 C9 78.1(3)
O23 K1 N31 149.56(12) C4 O2 K14 154.4(4)
O23 K1 N3 81.73(13) K11 O3 K1 83.65(11)
O23 K1 C5 112.36(13) K11 O3 K12 145.51(15)
O23 K1 C52 116.92(14) K1 O3 K12 107.33(14)
O3 K1 K11 47.71(9) C5 O3 K12 87.5(3)
O32 K1 K11 112.87(9) C5 O3 K1 114.7(3)
O3 K1 K12 37.20(8) C5 O3 K11 118.1(3)
O31 K1 K12 126.11(9) C5 O4 K12 95.0(3)
O32 K1 K12 35.47(8) K1 N3 K11 72.10(11)
O31 K1 K11 48.64(9) C2 N3 K11 141.8(4)
O3 K1 S23 162.67(10) C2 N3 K1 126.1(3)
O31 K1 S23 68.14(9) C2 N3 C3 104.8(4)
O32 K1 S23 115.46(9) C3 N3 K11 99.1(3)
O3 K1 O32 72.67(14) C3 N3 K1 107.4(3)
O31 K1 O3 96.35(11) C2 N4 C7 104.0(4)
O31 K1 O32 145.51(15) C2 N4 C8 124.3(4)
O31 K1 N31 51.06(11) C7 N4 C8 131.3(4)
O3 K1 N31 74.64(11) N3 C2 Br1 122.5(4)
O31 K1 N3 83.40(13) N3 C2 N4 114.6(4)
O3 K1 N3 56.45(12) N4 C2 Br1 122.9(4)
O32 K1 N31 94.51(11) N3 C3 C5 117.7(4)
O32 K1 C5 75.30(12) C7 C3 N3 110.9(4)
O3 K1 C5 18.59(12) C7 C3 C5 131.2(5)
O31 K1 C5 104.72(13) O1 C4 C7 114.6(5)
O31 K1 C52 125.89(14) O2 C4 O1 123.1(5)
O3 K1 C52 83.43(13) O2 C4 C7 122.3(5)
O32 K1 C52 23.15(12) K12 C5 K1 88.15(13)
O42 K1 K12 72.64(8) O3 C5 K1 46.7(3)
O42 K1 K11 116.51(10) O3 C5 K12 69.3(3)
O42 K1 S23 75.94(8) O3 C5 O4 122.7(5)
O42 K1 O32 45.34(11) O3 C5 C3 118.4(4)
O42 K1 O3 103.98(12) O4 C5 K1 146.5(4)
O42 K1 O31 110.76(13) O4 C5 K12 61.2(3)
O42 K1 N3 158.03(12) O4 C5 C3 118.9(5)
O42 K1 N31 72.11(12) C3 C5 K1 80.1(3)
O42 K1 C52 23.87(12) C3 C5 K12 149.0(3)
O42 K1 C5 114.97(12) N4 C7 C4 122.6(5)
N31 K1 K11 48.10(8) C3 C7 N4 105.7(4)
N31 K1 K12 83.62(8) C3 C7 C4 131.7(5)
N3 K1 K12 85.46(9) N4 C8 C9 118.1(5)
N3 K1 K11 59.80(10) N4 C8 C10 117.8(5)
N3 K1 S23 125.83(9) C10 C8 C9 99.0(4)
N3 K1 O32 113.92(12) C8 C9 S2 90.2(4)
N3 K1 N31 107.90(11) C8 C10 S2 90.3(4)
N3 K1 C5 43.43(12)

Symmetry code: 1-X,1-Y,1-Z; 21-X,1-Y,1-Z; 3-1/2+X,1/2-Y,-1/2+Z; 41/2+X,1/2-Y,1/2+Z

The reaction of 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid obtained in situ from the disodium salt IIa with amines gave 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylic acid salts IId–f (Scheme 1) [12].

2-Bromo-1-(thietan-3-yl) imidazole-4,5-dicarbohydrazide (III) was prepared by the reaction of dimethyl 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylates (I) with hydrazine hydrate (Scheme 1) [11]. The condensation of compound III with 4-(dimethylamino) benzaldehyde, 3-methoxy-4-hydroxybenzaldehyde, and 4-bromoacetophenone gave the corresponding diylidenehydrazidеs of 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylic acids IVa-c (Scheme 1) [13]. The structure of compound IVb was proved by 1Н NMR spectroscopy. The 1Н NMR spectrum of IVb exhibits doubled signals as two singlets for NH groups at 12.51 (Z), 12.22 (Е) ppm and 11.74 (Z), 11.59 (Е) ppm. The spectrum shows characteristic signals for thietane ring protons, the NСH group resonating in the 6.00-6.12 (Z) and 5.72-5.78 (Е) ppm ranges, and the two S(CH)2 proton signals in the 4.21-4.27 and 3.38-3.45 ppm ranges. The spectrum also contains double proton signals for the 3-methoxy-4-hydroxybenzaldehyde residues.

Antidepressant activity

Screening for the presence of antidepressant activity revealed differently directed effects of the investigated derivatives on the immobilization parameters (the immobility time and the index of depression) in the TST and FST (table 3). It was demonstrated that only compound (IIb) significantly reduced the immobility time (by 59%, р=0,025) compared with the control in the TST. Other compounds, as well as the reference drug imipramine, caused a stable tendency of reducing this parameter. In addition, compound (IVa) caused a statistically significant reduction of the immobility time by 63% (p=0.0004) in the FST. The index of depression was significantly decreased under the action of I, IIb, IId-f, IVа, and IVc, in particular, by 21 (р= 0.0115), 19 (р= 0.027), 13 (р= 0.034), 24 (р= 0.002), 35 (р = 0.002), 17 (р= 0.048), and 19% (р= 0.004), respectively. Imipramine reduced the index of depression by 17% (p-0.004). A statistically significant reduction of both parameters (the immobility time and the index of depression) was observed only in the groups that received IIb and IVa (by 59%, 19% and 63%, 19%, respectively) (table 3). Therefore, the acute toxicity was studied for IIb and IVa. The compounds were injected with non-inbred male mice at a single i. p. dose of 200, 400, 600, 800, 1000, 1200, 1400, and 1600 mg/kg. The animal behavior was assessed constantly during the first day. The death of animals was registered during 14 d.


Scheme 1: Synthesis of 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid derivatives IIa-f, IVa-c

It was established that LD50 values of IIb and IVа were significantly lower than that of the reference drug imipramine. Compound IIb refers to hazard class V according to I. V. Berezovskaya's classification [21] (almost non-toxic compound). The LD50 of IVa was not reached, because there were no animal deaths, even after the injection of the maximal possible dose (1600 mg/kg i. p.). Imipramine is moderately toxic for the same route of administration (hazard class III) (table 4).

To exclude "false positive" results of the TST and FST, compounds I, IIa–f, and IVа-c were studied in the OF-test (fig. 2–6). The compounds were injected in male mice as single i. p. doses 30 min before the test. The doses were the same as the doses used in the TST and FST (table 3). It was discovered that IIb and IVа did not significantly modify the behavior of animals in the OF-test. Both integral criteria (exploratory activity and anxiety) and basic behavioral patterns were comparable with those for the control group, except “movement on the spot” and “defecation”. These two patterns significantly decreased in the group that received compound IIb, but in the control group, statistical significance was not observed (fig. 3). Compound IVb decreased the anxiety by 42% (p=0.018), the number of “moving” patterns by 24%, and “recline against the wall” by 62% and increased “movement on the spot” by 42% in comparison with the control group (fig. 5). A significant increase in the pattern “movement on the spot”, 50 and 58%, was induced by compounds I and IId, respectively (fig. 4–5). There was a decrease in the number of “defecations” and a tendency of “grooming” to increase in the group of IIe compound (fig. 2). The obtained results indicate that compound IVb reduces statistically significantly the anxiety and locomotor activity. The compounds I, IIa-f, and IVа,c do not have sedative and/or stimulative effects. This means that the TST and the FST results show an antidepressant effect of thietane-containing 2-bromoimidazole-4,5-dicarboxylic acid derivatives.

Thus, all the studied compounds I, IIa–IIf, and IVа-c, showed antidepressant-like activity after single i. p. injection. Compounds IIb and IVa have the most pronounced effect comparable with that of imipramine. These two compounds are safer (IV-V class of toxicity) than imipramine (III class of toxicity) and are promising for further studies. In addition, compound IVb significantly reduced the anxiety and decreased the locomotor activity of male mice.


Fig. 2: Study of the influence of IIa and IIe in the OF-test, *-The difference is significant in comparison with control for IIe (p<0.05 for Wilcoxon-Mann-Whitney U-test)

Table 3: The effect of thietane-containing derivatives of 2-bromoimidazole-4,5-dicarboxylic acid on the parameters of TST and FST after single injection

No. Compounds (dose) TST immobility time FST immobility time FST index of depression
Control 100.0 (62.0-105.0)n=14 101.5 (76.0-128.0)n=14 1.06 (0.94-1.12)n=13
Imipramine
(10 mg/kg)		 40.0(24.0-106.0)n=15 78.0*(26.0-82.0)n=13 0.85*(0.76-0.89)n=13
I
(13 mg/kg)		 77.0(34.5-141.5)n=8 122.5(96.0-147.0)n=8 0.80*(0.72-0.89)n=8
IIа
(13 mg/kg)		 84.0(32.0-159.5)n=8 104.5(56.5-145.5)n=8 0.92*(0.87-0.97)n=8
IIb
(10 mg/kg)		 41.0*(20.0-68.0)n=8 75.0(51.5-96.5)n=8 0.83*(0.73-0.91)n=7
IIc
(13 mg/kg)		 90.5(63.5-225.5)
n=8		 69.0(69.0-75.0)
n=5		 0.87*(0.84-1.0)n=6
IId
(15 mg/kg)		 68.0(48.0-157.5)n=8 99.0(65.0-122.0)n=8 0.78*(0.61-0.80)n=8
IIe
(15 mg/kg)		 62.0(35.0-212.0)n=8 72.5(55.0-112.0)n=8 0.90*(0.77-0.94)n=8
IIf
(16 mg/kg)		 64.5(38.0-110.0)n=8 126.0(102.5-154.0)n=8 0.67*(0.53-0.79)n=8
IVа
(21 mg/kg)		 81.5(35.0-149.5)n=8 37.5*(12.5-48.5)n=8 0.83*(0.71-0.89)n=8
IVb
(23 mg/kg)		 72.0(46.0-138.0)n=8 105.5(78.0-131.0)n=8 0.78*(0.63-0.88)n=8
IVc
(26 mg/kg)		 85.5(59.0-106.5)n=8 86.5(57.0-113.0)n=8 0.86*(0.83-0.96)n=7
Kruskal-Wallis test H (11. N= 109) =9.06 p =0.616 H (11. N= 104) =28.52 p =0.003 H (11. N= 102) =34.39 p =0.0003

Note: *-the difference is significant in comparison with control (p<0.05 for Wilcoxon-Mann-Whitney U-test)


Fig. 3: Study of the influence of IIb and IVa in the OF-test, *-The difference is significant in comparison with control for IIb (p<0.05 for Wilcoxon-Mann-Whitney U-test)


Fig. 4: Study of the influence of IId and IIf in the OF-test, *- The difference is significant in comparison with control for IId (p<0.05 for Wilcoxon-Mann-Whitney U-test)

Fig. 5: Study of the influence of I and IVb in the OF-test

*- The difference is significant in comparison with control for IVb (p<0.05 for Wilcoxon-Mann-Whitney U-test)

”- The difference is significant in comparison with control for I (p<0.05 for Wilcoxon-Mann-Whitney U-test)


Fig.6: The study of the influence of IIc and IVc in the OF-test

Table 4: The acute toxicity of compounds IIb, IVа, and imipramine for the single i. p. injection to male mice [21]

Compound LD50, mg/kg Class/Degree of toxicity
IIb 1170 IV/low toxicity
IVа >1600 V/Almost non-toxic
Imipramine 68,4 III/Moderately toxic

CONCLUSION

Salts and diylidenehydrazides of 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid were synthesized. The structure of synthesized compounds was confirmed by 1H NMR spectroscopy. The position of the potassium cation in the monopotassium salt of 2-bromo-1-(thiethan-3-yl) imidazole-4, 5-dicarboxylic acid was determined by X-ray diffraction analysis. The most pronounced antidepressant effect comparable to imipramine is characteristic of dipotassium 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylate and 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylic acid di[(4-dimethylaminophenyl) methylidenehydrazide], which is superior in safety and are promising for further investigation.

Thus, compounds with marked antidepressant activity were identified among the synthesized derivatives of 2-bromo-1-(thietan-3-yl) imidazole-4,5-dicarboxylic acid. An obvious advantage of these compounds is low toxicity.

AUTHORS CONTRIBUTION

Professor Ferkat Khaliullin is the scientific leader in the synthesis of biologically active derivatives of 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylic acid, Ms. Anfisa Valieva is the direct executor of the synthesis of new derivatives of 2-bromo-1-(thietan-3-yl)imidazole-4,5-dicarboxylic acid. Professor Irina Nikitina is the scientific leader in the study of antidepressant activity of newly synthesized compounds, Mrs. Albina Miftakhova played an important role in obtaining results on antidepressant activity. Professor Leonard Khalilov is the scientific leader in carrying out structural studies of new compounds, Mrs. Ekaterina Mescheryakova played an important role in conducting X-ray structural analysis with subsequent interpretation of the results.

CONFLICT OF INTERESTS

The authors declare that they have no conflict of interest

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