RU2768761C1 - Use of substituted esters of (z)-2-(2-(diphenylmethylene)hydrazinyl)-5,5-dimethyl-4-oxohex-2-enoic acid as an antibacterial and antifungal agent - Google Patents

Abstract

FIELD: chemistry.SUBSTANCE: invention relates to use of substituted esters of (Z)-2-(2-(diphenylmethylene)hydrazinyl)-5,5-dimethyl-4-oxohex-2-enoic acid 1, 2 of structural formula I as an antibacterial and antifungal agent.(I).EFFECT: compounds having pronounced antibacterial and antifungal activity, as well as low toxicity are obtained.1 cl, 2 tbl, 4 ex

Description

The invention relates to the field of organic chemistry, to new biologically active substances of the class of derivatives of 2-hydrazono-5,5-dimethyl-4-oxobut-2-enoic acids, namely to substituted esters (2)-2-(2-(diphenylmethylene) hydrazinyl)-5,5-dimethyl-4-oxohex-2-enoic acid 1, 2 structural formulas:

possessing antibacterial and antifungal activity, which suggests their use in medicine as a drug with antibacterial and antifungal properties.

The structural analogue of the claimed compounds is 2-((9H-fluoren-9-ylidene)hydrazono)-5,5-dimethyl-4-oxo-N-(2-hydroxy-3-isopropyl-6-methylphenyl)hexanamide 3, which has antibacterial and antifungal activity [A.I. Syutkina, N.M. Igidov, M.V. Dmitriev, P.P. Makhmudov, V.V. Novikova // Journal of General Chemistry. - 2019. - V. 89, No. 7. - S. 1026-1032] formulas:

Data on the antibacterial and antifungal activity of compound 3 are given:

The comparison standards are phenyl salicylate with the structural formula:

and fluconazole of the structural formula:

which are widely used in medical practice and are analogues in action [Mashkovsky M.D. Medicines. - 16th ed., revised, corrected. and additional - M.: New wave, 2012. - p. 9505].

From the prior art, the structural formulas of compounds 1, 2 [A.I. Syutkina, N.M. Igidov, I.A. Kizimova // Journal of Organic Chemistry. - 2020. - V. 56, No. 4. - S. 613-618].

Compounds 1 and 2 were prepared by reacting 5-tert-butyl-3-((diphenylmethylene)hydrazono)furan-2(3H)-one with methanol and ethanol, respectively, in the presence of equimolar amounts of triethylamine, followed by isolation of the target product by known methods according to the scheme:

However, from the patent and scientific and technical literature, the use of compounds 1, 2 as agents with antimicrobial (antibacterial) and antifungal properties has not been revealed.

The objective of the invention is to search in the series of derivatives of 2-hydrazono-5,5-dimethyl-4-oxobut-2-enoic acids for substances with a pronounced antibacterial and antifungal effect, as well as low toxicity.

The task is achieved by obtaining methyl and ethyl esters of (Z)-2-(2-(diphenylmethylene)hydrazinyl)-5,5-dimethyl-4-oxohex-2-enoic acid 1, 2, which have antibacterial and antifungal effects.

EFFECT: obtained compounds with high yields, with pronounced antimicrobial (antibacterial) and antifungal activity, as well as low toxicity.

The invention is illustrated by examples of the study of pharmacological properties.

Example 1 Preparation of Compound 1 To 1 mmol of 5-tert-butyl-3-((diphenylmethylene)hydrazono)furan-2(3H)-one was added 10 ml of methanol and 1 mmol of triethylamine. The reaction mixture was refluxed with stirring for 5 min until a yellow solution formed, then cooled. The resulting precipitate was filtered off and recrystallized from methanol.

The resulting compound 1 is a white crystalline substance, soluble in chloroform, DMSO, acetone, sparingly soluble in methanol, insoluble in water and hexane.

Yield 0.35 g (96%), white crystals, mp. 152-154°C (ethanol). IR spectrum, ν, cm ^-1 : 1739 (C=O), 1619, 1560 (C=C, C=N, CO _chel ). 1H NMR spectrum _{(CDCl 3} ⁾ , δ, ppm: 1.10 s (9H, t-Bu), 4.00 s (3H, COOCH ₃ ), 5.50 s (1H, CH), 7.33 m (5H _arom ), 7.51 m (2H _arom ), 7.61 m (3H _arom ), 12.18 s (1H, NH). Spectrum of NMR ¹³ C (CDCl ₃ ), δ, ppm: 27.14, 42.51, 52.58, 90.96, 127, 129.67, 129.5, 129.6, 129.33, 129.67, 129.5, 129.6, 129.33 Mass spectrum, m/z (I _rel , %): 364 [M] ⁺ (12), 307 [M-t-Bu ⁺ ] (100), 305 [M-COOMe] ⁺ (6), 279 [M -t-BuCO] ⁺ (16), 181 [(Ph) ₂ C=N] ⁺ (9), 57 [t-Bu] ⁺ (23). Found, %: С 72.52; H 6.60; N 7.68. C ₂₂ H ₂₄ N ₂ O ₃ . Calculated, %: С 72.51; H 6.64; N 7.69. M 364.44.

Example 2 Compound 2 was obtained similarly, replacing methanol with ethanol.

The resulting compound 2 is a white crystalline substance, soluble in chloroform, DMSO, acetone, sparingly soluble in methanol, insoluble in water and hexane.

Yield 0.37 g (98%), white crystals, mp. 142-144°C (ethanol). IR spectrum, ν, cm ^-1 : 1739 (C=O), 1615, 1554 (C=C, C=N, CO _chel ). 1Н NMR spectrum (DMSO-d ⁶ ), δ, ^ppm : 1.00 s (9Н, t-Bu), 1.35 t (3Н, CH ₂ CH ₃ ), 4.39 q (2Н, CH ₂ CH ₃ ), 5.54 s (1H, CH), 7.45 m (10H _arom ), 12.04 br s (1H, NH) Found, %: C 72.95; H 6.90; N 7.43 C ₂₃ H ₂₆ N ₂ O ₃ Calculated, %: C 72.99, H 6.92, N 7.40, M 378.47.

Example 4. To determine the antibacterial and antifungal activity, the method of two-fold serial dilutions in a liquid nutrient medium was used in relation to three strains - S. aureus ATCC 6538-P and E. coli ATCC 25922, C. albicans ATCC 885-653 [Guidelines for conducting preclinical drug research. Part one. - M.: Grif and K, 2012. - 202 s]. To prepare the stock solution, the test compound in an amount of 0.05 g was dissolved in 5 ml of dimethyl sulfoxide, obtaining a solution concentration of 10 ⁴ μg/ml. Next, a working solution of the substance was prepared: 4 ml of the nutrient medium was placed in a sterile test tube, 1 ml of the stock solution of the test compound was added, obtaining a concentration of 2000 μg/ml. After that, a series of serial dilutions of compounds with a two-fold decreasing concentration (from 1000 μg/ml to 0.06 μg/ml) was prepared. An intact nutrient medium was used as a negative control, and a culture medium without the test compound was used as a positive control.

Cultures were grown in test tubes on slant agar medium (nutrient agar for bacteria, Sabouraud agar for fungi), 24-hour cultures were used. To prepare a working suspension of microorganisms, the grown culture was washed with an isotonic sterile sodium chloride solution and the density of the microbial suspension was set according to the turbidity standard of 5 units. Further, a working solution with a concentration of 5⋅10 ⁶ CFU/ml was prepared from the resulting microbial suspension. A microbial suspension was added to each test tube in an amount of 0.1 ml. In this case, the microbial load was about 2.5×10 ⁵ CFU/ml for bacteria and 2.5×10 ⁴ CFU/ml for fungi. The results were recorded after 18-24 hours of thermostating at 35-37°C. The minimum inhibitory concentration (MIC) was determined by the absence of visual signs of microbial growth on the nutrient medium.

Example 5. Acute toxicity was determined in experiments on white non-linear male mice weighing 20-22 g, contained on a normal vivarium diet. For calculations, the method of V.B. Prozorovsky was used. [Prozorovsky V.B. Statistical processing of the results of pharmacological studies. Psychopharmacology and biological narcology. - 2007. - No. 7. - S. 2090-2120.] The test compound was administered orally as a suspension in a 2% starch solution. Each dose was administered to two animals. The animals were observed for 14 days. The general condition and behavior of the animals, the condition of the coat were recorded, and the body weight was determined. In each group of animals, the number of deaths for the entire observation period was taken into account.

For test compounds 1, 2, LD50 is >5000 mg/kg.

According to the toxicity classification of drugs, compounds 1, 2 belong to the V class of practically non-toxic drugs [Izmerov N.F., Sanotsky I.V., Sidorov K.K. Parameters of toxicometry of industrial poisons at a single exposure: a Handbook. M., 1977. - p. 196].

The test results are presented in the table:

As can be seen from the table, the antibacterial effect of the claimed compounds 1, 2 exceeds that of the reference drug phenyl salicylate by 3 times in relation to S. aureus in the presence of an effect on E. coli as well, in addition, the claimed compounds 1, 2 have an antifungal effect comparable to the effect the reference drug fluconazole, while having more than 3.5 times less toxicity. Therefore, the claimed compounds 1, 2 can be used in medical practice as antibacterial and antifungal drugs.

Claims (3)

Application of 2-(2-(diphenylmethylene)hydrazinyl)-5,5-dimethyl-4-oxohex-2-enoates 1, 2 structural formula:

as an antibacterial and antifungal agent.