RU2675806C1 - N-(indolyl)trifluoroacetamides with antimicrobial activity production method - Google Patents

Abstract

FIELD: chemistry.SUBSTANCE: invention relates to the field of chemistry, namely to the N-(2,3-dimethyl-1H-indole-7-yl)-2,2,2-trifluoroacetamide and N-(1,2,3-trimethyl-1H-indol-7-yl)-2,2,2-trifluoroacetamide production method. Method consists in the corresponding aminoindole interaction with trifluoroacetic acid ethyl ether.EFFECT: invention can be used for the drugs with antimicrobial activity production.2 cl, 2 tbl, 4 ex

Description

The invention relates to chemistry, namely to N- (2,3-dimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide and N- (1,2,3-trimethyl-1H-indole-7 -yl) -2,2,2-trifluoroacetamide, which can be used in medical practice as antimicrobial agents.

The need to search for new highly effective and safe antimicrobial compounds is enshrined in the Russian Federation at the legislative level [1]. One of the main modern directions in the development of antimicrobial compounds is the synthesis of analogues and derivatives of already studied drugs. Currently, there are more than fifteen million chemical compounds with antimicrobial activity isolated from natural sources and obtained by chemical synthesis, but only dozens satisfy the requirements for antimicrobial action and biological safety [2-4]. The problem of resistance to both antibiotics and disinfectants is becoming increasingly relevant and alarming in the 21st century. Since the uncontrolled use of antimicrobials in the food industry as preservatives, in animal husbandry and in agriculture as disinfectants and chemotherapeutic agents inevitably entails the formation of resistance of microorganisms to the most commonly used antimicrobial agents. In most regions of the world and in Russia, including antibiotic-resistant strains, both gram-positive (staphylococci, enterococci) and gram-negative bacteria (enterobacteria, pseudomonads, etc.) are spreading [5-9]. About twenty million people die every year from infectious diseases in the world. The annual incidence of infectious diseases amounts to hundreds of millions of cases. Difficulties in the treatment and prevention of infectious diseases caused by the variety of biological forms of pathogens, the colossal adaptive ability of microorganisms and the constant emergence of forms with multiple resistance, the emergence of new types of microorganisms, determine the urgency of the problem of creating new antimicrobial agents [10-12]. And the prospects for the development and implementation of new antimicrobial agents look rather depressing [1]. When conducting microbiological monitoring in 2013-2014. an increased specific weight of polyresistant methicylin-resistant strains of S.aureus, P.aeruginosa, multidrug resistance of the Enterobacteriaceae family to three or more antimicrobial drugs was noted [11-14]. In modern conditions, studies based on one of the fundamental principles of overcoming the resistance of microorganisms - the search for new compounds with antimicrobial activity and, possibly, with a different mechanism of action - are undoubtedly of interest [1, 13].

Substituted aminoindoles with an amino group in the benzene ring are known as starting compounds for the preparation of indolylamides, indolenamino ketones, pyrroloindoles, pyrroloquinolines. Many of the resulting products show various kinds of biological activity. So, in amides, pyrroloquinolines based on 4,7-aminoindoles and trifluoroacetoacetic ester, a rather high antimicrobial activity was found [14, 15]. In this regard, it is important to study the reaction of 2,3-dimethyl- and 1,2,3-trimethyl-7-aminoindoles with trifluoroacetic acid ethyl ester in order to develop a method for the preparation of the corresponding (indolyl) trifluoroacetamides and their laboratory study on antimicrobial activity.

The closest technical solution to the claimed invention is a method for producing trifluoromethyl-containing N- (indolyl) acylamides by acylation of trifluoroacetoacetic acid ethyl ester of 2,3-dimethyl- and 1,2,3-trimethyl-7-aminoindoles [16].

The disadvantage of this method is the formation in the reaction along with indolylamides of side enaminoketones, when purified from which the yield of the target product decreases.

The claimed compounds, their antimicrobial properties and method of preparation are not described in the literature.

The technical result consists in obtaining new compounds of N- (indolyl) acylamides in high yield, having an effective antimicrobial effect.

The specified technical result is achieved due to the use of a more accessible compound, ethyl trifluoroacetic acid, as the acylating agent.

The invention consists in the fact that in the process for producing N- (indolyl) trifluoroacetamides general formula (1), which by chromatographic and spectral analysis, may be in solution in two tautomeric forms: in DMSO-d ₆ N- (2,3 -dimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide (a) and N- (1,2,3-trimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide (mainly b):

(1)

(one)

where R = H, CH ₃ , aminoindoles of the general formula (2)

(2)

(2)

where R = H, CH ₃ , in boiling benzene with catalytic amounts of glacial acetic acid is reacted with an acylating agent.

Ethyl trifluoroacetic acid of general formula (3) is used as the acylating agent;

(3)

(3)

The invention also consists in the use of N- (indolyl) trifluoroacetamides of the general formula (1) for the preparation of synthetic drugs with an effective antimicrobial effect.

In the table. 1 shows the determination of the minimum inhibitory concentrations (MPC) of the studied compounds by the method of serial dilutions in a liquid nutrient medium; in table 2 - determination of the IPC of the studied compounds by the method of serial dilutions in a liquid nutrient medium.

Information confirming the achievement of the technical result is presented in the following examples.

Example 1. N- (2,3-dimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide

To a solution of 0.32 g (2 mmol) of 2,3-dimethyl-7-aminoindole in 150 ml of absolute benzene, add 5 ml (excess) of ethyl trifluoroacetic acid, a catalytic amount of glacial acetic acid and boil under effective reflux until until the starting amine disappears (chromatographic control). At the end of the reaction (boiling for 20-30 hours) benzene, excess ether, traces of acetic acid are distilled off to dryness from the reaction mixture. The solid residue is dissolved in chloroform and passed through a layer of alumina. The yield of indolyl trifluoroacetamide is 57%, mp. 119-121 ° C, Rf = 0.12, 0.83 (benzene-ethyl acetate 5: 1). Found,%: C 56.03; H 4.19; M 256 (mass spectrometric). C ₁₂ H ₁₁ F ₃ N ₂ O. Calculated,%: C 56.25; H 4.33; M 256. UV spectrum (ethanol) λ _max (log έ): 213 (4.47), 231 (4.37), 298 (3.95); ¹ H NMR spectrum (DMSO-d ₆ ): 212 (3H, s, 3-CH ₃ ), 2.30 (3H, s, 2-CH ₃ ), 6.92 (1H, t, J = 8 Hz, H-5), 7.00 (1H, d, J = 8 Hz, H-6), 7, 27 (1H, d, J = 8 Hz, H-4), 10.57 (1H, s, 1 -H), 10.91 (1H. S, H-N) ppm. Mass spectrum Jm / z (% to J _max ): 256 (100), 255 (34.13), 241 (10.61), 237 (15.82), 223 (10.81), 185 (5, 61), 159 (7.71), 69 (7.11), 28 (7.41), 18 (17.22).

Example 2. N- (1,2,3-trimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide

Obtain analogously to example 1 from 0.35 g (2 mmol) of 1,2,3-trimethyl-7-aminoindole, but to complete the reaction, the reaction mixture is heated for 40-50 hours. The resulting amide is purified on a column of aluminum oxide in a chloroform-system petroleum ether 5: 1. The yield is 65%, mp. 126-128 ° C, Rf = 0.21, 0.87 (benzene-ethyl acetate 5: 1). Found,%: C 57.32; H 4.70; M 270 (mass spectrometric). C ₁₃ H ₁₃ F ₃ N ₂ O. Calculated,%: C 57.48, H 4.85; M 270. UV spectrum (ethanol) λ _max (log έ)): 212 _pl (4.40), 232 (4.73), 290 (4.15), ¹ H NMR spectrum (DMSO-d ₆ ) (a) tautomeric forms: 2.18 (3H, s, 3-CH ₃ ), 2.30 (3H, s, 2-CH ₃ ), 3.69 (3H, s, 1-CH ₃ ), 6.88 (1H, d, J = 8 Hz, H-6), 6.99 (1H, t, J = 8 Hz, H-5), 7.42 (1H, d, J = 8 Hz, H-4), 11, 35 (1H, C, H-N) ppm; (b) the tautomeric forms: 2.18 (3H, s, _3-CH3), 2.31 (3H, s, _2-CH3), 3.94 (3H, s, _1-CH3), 6, 99 (1H, t, J = 8 Hz, H-5), 7.17 (1H, d, J = 8 Hz, H-6), 7.36 (1H, s, H-O), 7.42 (1H, d, J = 8 Hz, H-4) ppm. The ratio of (a) to (b) according to the integrated intensity of the characteristic signals in the ¹ H NMR spectrum is 1: 3. Mass spectrum Jm / z (% to J _max ): 270 (100), 269 (18.62), 255 (16.52), 239 (6.71), 214 (24.52), 201 (8, 0), 199 (15.02), 185 (6.51), 184 (10.31), 183 (8.81), 69 (10.51), 28 (10.71).

1H NMR spectra were recorded on a Joel JNM-ECX400 multicore nuclear magnetic resonance spectrometer (400 MHz) in DMSO-d6. Electronic spectra were obtained on a LEKI SS2109UV instrument in ethanol. Mass spectra were recorded on a Finnigan MAT INCOS-50 mass spectrometer with direct input of the sample into an ion source at an ionization energy of 70 eV. The names of amines, amides are given according to the rules of the computer program ACD / LABS IUPAC Name Generator. Structural formulas of compounds are drawn in the ISIS Draw 2.4 computer program. Purification of the reaction products was carried out by column chromatography. As the sorbent used aluminum oxide (neutral, I and II art. Act. According to Brockman). Monitoring the progress of reactions, the purity of the compounds obtained, R _f determination was performed using TLC plates Silufol UV-254 (contains the system specific for each compound in the experimental method).

Example 3. The study of the antimicrobial activity of N- (2,3-dimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide and N- (1,2,3-trimethyl-1H-indol-7-yl ) -2,2,2-trifluoroacetamide

When conducting a microbiological experiment, the studied compounds were used in the form of a solution (Dimexide was used as a solvent for the preparation of solutions for external use, manufactured by Marbiopharm OJSC). The following museum strains were used as test microorganisms in the study of the antimicrobial activity of the compounds of the general formula (1): Staphylococcus aureus 6538-P ATCC Escherichia coli 25922 ATCC, Pseudomonas aeruginosa 27853 ATCC, Streptococcus pyogenes 19615 ATCC, Klebsiococcalcephonia pneumonia. The museum strains used in this work were obtained from the collection of the Museum of Living Cultures of FSBI “NTsESMP” of the Ministry of Health of Russia, Becton Dickinson France S.A.S. The antimicrobial activity of amides of the general formula (1) was determined by the method of serial dilutions in broth (macroscopic method "test tube") (MUK 4.2.1980-04). The antimicrobial preparation dioxidin (a derivative of di-N-hydroxyquinoxaline) (production of Biosynthesis, solution for topical application, endotracheal and intravenous administration, 10 mg / ml), widely used in medical practice, was used as a comparison drug. This drug has high in vivo chemotherapeutic activity on model infections similar in pathogenesis to pathological processes in humans (purulent meningitis, pyelonephritis, septicopyemia) and caused by strains of anaerobic bacteria that are resistant (including multiresistant) to drugs of other classes, including Pseudomonas aeruginosa strains sticks and methicillin-resistant staphylococci. Dioxidin is characterized by a wide antibacterial spectrum with a bactericidal effect, and is also active against gram-positive and gram-negative aerobic conditionally pathogenic bacteria. The activity of dioxidine against tuberculosis mycobacteria is shown. For the preparation of comparison of dioxidine IPC relative to strains of Staphylococcus spp. is 125.0-1000.0 μg / ml, Escherichia coli 8.0-250.0 μg / ml, Pseudomonas spp. 125.0-1000.0 μg / ml, Streptococcus spp. 64.0-1000.0 μg / ml [17].

To assess the sensitivity of microorganisms, Muller-Hinton Bouillon (ICB) was used, which is approved for use in the Russian Federation in the prescribed manner and satisfies the requirements in terms of its characteristics. Intralaboratory quality control of the environment was carried out using all the media approved for use in the Russian Federation in the established manner. The concentration of the suspension of the studied microorganism was 1.5 × 10 ⁸ CFU / ml The optical density of the bacterial suspension with a concentration of 1.5 × 10 ⁸ CFU / ml upon visual inspection corresponded to the turbidity standard of 0.5 according to McFarland. A commercial turbidity standard was used in the work. A bacterial suspension was prepared from agar cultures. To prepare the inoculum, a pure daily culture of microorganisms grown on solid nutrient media was used. Selected several of the same type, clearly isolated colonies grown on non-selective solid nutrient media. Loops transferred a small amount of material from the tops of the colonies into a test tube with sterile saline, adjusting the inoculum density to exactly 0.5 according to the McFarland standard. Inoculum was used for 15 minutes after preparation.

Example 4. The method of serial dilutions in the broth - macrometer (test)

Testing was carried out in a volume of 1 ml of each dilution of the test compound with a final concentration of the studied microorganism of about 5 × 10 ⁵ CFU / ml. For the determination of sensitivity, MBC was poured into 0.5 ml in each tube. The number of tubes was nine, plus one for setting a “negative” control, that is, ten. A working solution of the test compound was prepared from the main solution using a liquid nutrient medium - MBB. Then a working solution in an amount of 0.5 ml using a micropipette with a sterile tip was introduced into the first tube containing 0.5 ml of broth. The mixture was thoroughly mixed and a new sterile tip transferred 0.5 ml of a solution of the test compound in the broth to a second tube containing initially 0.5 ml of broth. This procedure was repeated until all the necessary series of dilutions was prepared. 0.5 ml of broth was removed from the last tube. Thus, a number of test tubes with solutions of the test compound were obtained, the concentrations of which differed by a factor of 2 in neighboring tubes. For inoculation, a standard microbial suspension equivalent to 0.5 according to the McFarland standard, diluted 100 times in ICB, was used, after which the concentration of the microorganism in it was approximately 10 ⁶ CFU / ml. 0.5 ml of inoculum was added to each tube containing 0.5 ml of the appropriate dilution of the test compound, and to one tube with 0.5 ml of MCB without antibiotic (“negative” control). The final concentration of the microorganism in each tube was approximately 5 × 10 ⁵ CFU / ml. The inoculum was introduced into test tubes with dilutions of the test compound no later than 15-30 minutes from the time of preparation. The tubes were closed with sterile cotton-gauze plugs and all but the “negative” control tubes were incubated in a normal atmosphere at 37 ° C for 16–20 or 20–24 h (depending on the type of microorganism being tested). The negative control tube was placed in a refrigerator at 4 ° C, where it was stored until results were taken into account. To determine the presence of microorganism growth, test tubes with inoculations were examined in transmitted light. The culture growth in the presence of the test compound was compared with a reference tube (“negative” control) containing the original inoculum and stored in the refrigerator. MIC was determined by the lowest concentration of the test compound, which inhibits the visible growth of the microorganism.

Relative to test strains of microorganisms, N- (2,3-dimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide shows the following activity: for S. aureus 6538-P, ATCC MPC of the test compound was 31.3 μg / ml; E. coli 25922 ATCC - 125 μg / ml, P. aeruginosa 27853 ATCC - 125 μg / ml, S. pyogenes 19615 ATCC - 31.3 μg / ml, Klebsiella pneumoniae 13883 ATCC - more than 250 μg / ml., N- ( 1,2,3-trimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide: for S. aureus 6538-P, the ATCC MPC of the test compound was 62.3 μg / ml; E. coli 25922 ATCC - 200 μg / ml, P. aeruginosa 27853 ATCC - 125 μg / ml, S. pyogenes 19615 ATCC - 31.3 μg / ml, Klebsiella pneumoniae 13883 ATCC - 125 μg / ml, Enterococcus faecalis 19433 ATCC - 125 μg / ml, which is comparable with the antimicrobial activity of the preparation for comparison of dioxidine. Thus, the claimed compounds have antimicrobial activity comparable to the activity of the preparation of comparison dioxidine.

The invention allows to obtain new compounds of N- (indolyl) acylamides in high yield, with an effective antimicrobial effect.

Information sources:

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8. Sukhorukova M.V. Antibiotic resistance of nosocomial Enterobacteriaceae strains in Russian hospitals: results of a multicenter epidemiological study “Marathon” 2013-2014 / M.V. Sukhorukova [et al.] // Clinical Microbiology and Antimicrobial Chemotherapy. - 2017. - T. 19. - No. 1. - S. 49-56.

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10. Kozlov R.S. Antibiotic resistance of gram-positive pathogens of complicated intra-abdominal infections in Russia / R.S. Kozlov [et al.] // Clinical Microbiology and Antimicrobial Chemotherapy. - 2015. - T. 17. - No. 3. - S. 227-234.

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Note: - ¹ titer of activity, "+++" - abundant growth; "++" - the deep or surface growth of strains is less abundant; "+" - weak growth - inactive less than 50-30%; "+/-, 0" - more than 70% delay or lack of culture growth compared to control

Note: - ¹ titer of activity, "+++" - abundant growth; "++" - the deep or surface growth of strains is less abundant; "+" - weak growth - inactive less than 50-30%; "+/-, 0" - more than 70% delay or lack of culture growth compared to control

Claims (7)

1. The method of obtaining N- (indolyl) trifluoroacetamides of the general formula (1) in solution in two tautomeric forms of N- (2,3-dimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide and N - (1,2,3-trimethyl-1H-indol-7-yl) -2,2,2-trifluoroacetamide

where R = H, CH ₃ in which aminoindoles of the general formula (2)

where R = H, CH ₃ , in boiling benzene with catalytic amounts of glacial acetic acid is reacted with an acylating agent, ethyl trifluoroacetic acid, of the general formula (3)

2. The use of N- (indolyl) trifluoroacetamides of the general formula (1), which are obtained by the method according to claim 1 of the claims, for the preparation of drugs having an antimicrobial effect.