RU2720490C1 - 3-(2-oxoimidazolidinyl-5)indoles and a method for production thereof - Google Patents

Abstract

FIELD: chemistry.SUBSTANCE: invention relates to a method of producing 3-(2-oxoimidazolidinyl-5)indole of general formula I, in which R=H, C-Calkyl; R=H, C-Calkyl, CHX, where X=Me; R=phenyl, involving reaction of 5-hydroxyimidazolidin-2-one of general formula IIwhere R=phenyl, with equimolar amount of indole of general formula IIIwhere R=H, C-Calkyl; R=H, C-Calkyl, CHX, where X=Me, in presence of 1–10 mol. % of boron trifluoride etherate in tetrahydrofuran at room temperature.EFFECT: disclosed is a single-step method of producing compounds of formula I from available starting compounds, using a standard catalyst, carried out in mild conditions.1 cl, 3 dwg, 6 ex

Description

The invention relates to organic and pharmaceutical chemistry, in particular to derivatives of indoles and 2-imidazolidinones, specifically to 3- (2-oxoimidazolidinyl-5) indoles of the general formula I having anti-inflammatory activity, and to a method for their preparation.

where R ¹ = H, C ₁ -C ₆ alkyl;

R ² = H, C ₁ -C ₆ alkyl, C ₆ H ₄ X, where X = H, Me;

R ³ = aryl.

The inventive compounds of General formula I, their properties and methods of preparation are not described in the literature.

The inventive compounds of General formula I are bis-heterocyclic compounds containing in their composition the nucleus of 2-imidazolidinone and indole, specifically derivatives of 1- and / or 2-substituted indole, in which the third position is 1-aryl-2-oxoimidazolidin-5 Strong Deputy.

It is known that compounds containing 2,3-alkylindole scaffold exhibit a wide range of biological activity [Schulz E.E., Kartsev V.G., Tolstikov G.A. Chemistry and biological activity of indole alkaloids. M .: Scientific Partnership, 2018. - 880 p.]. In turn, compounds containing a 2-imidazolidinone fragment are promising for studying their biological activity: the imidazole ring plays a key role in the structure and functions of some important biological objects (histidine, histamine, purine precursors, etc.). The hydrogen analog acceptor containing a structural analog of imidazole, 2-imidazolidinone, can both enhance and completely change the biological properties of the studied objects. For example, 2-imidazolidinone is an integral part of the antihypertensive drug imidapril, which is also used to treat chronic heart failure [Demolis P. et al., Fundam. Clin. Pharmacol 8, 80, 1994]. In turn, a phenylindole derivative containing a 2-imidazolidinone core is known as the antipsychotic drug sertindole and is used in the treatment of schizophrenia [Lewis R., Bagnall A.M., Leitner M., Sertindole for schizophrenia. Cochrane Database of Systematic Reviews, 3, 2005 ,. Art. No .: CD001715].

The search for new therapeutic agents, including a number of substituted indoles, is necessary due to problems associated with intolerance to known drugs, the loss of their effectiveness in the treatment process, as well as the indiscriminate action. Bis-heterocyclic compounds containing an indole fragment are promising from the point of view of correcting existing biological activity, manifesting its new species, improving their pharmacological properties and creating new drugs. They represent a powerful structural tool with which you can modify the molecules of pharmacologically active compounds. Thus, it is known that compounds containing a pyrimidoindole scaffold have been used in the design of antitumor drugs [Reader J.C., Matthews TP, et al., J. Med. Chem. 2011, 54, 8328]. The idazol il indoles substituted by him also exhibit physiological activity, in particular as protein kinase C inhibitors [WO 99/32483, 1999, C07D 403/04], and can be used in the treatment of inflammatory, immunological, bronchopulmonary, cardiovascular and oncological diseases [ WO 00/78750 A1, 2000, C07D 403/04].

By introducing an additional heterocycle into bioactive molecules, one can change their pKa, construct conformational difficulties, vary the hydrophilic-hydrophobic balance, increase lipophilicity, and these parameters can be modulated individually or in various combinations.

It is known that the introduction of an additional heterocyclic nucleus often allows compounds to be obtained that are superior in activity to the active drugs [Archana S. et al. Res. Chem. Intermed., Springer, 2017, 43 (4), 2471]. The introduction of a 2-oxoimidazolidinyl group into a heterocyclic scaffold based on substituted indoles provides its additional interaction with hydrophobic pockets of the target protein. At the same time, 3- (2-oxoimidazolidinyl-5) indoles I, as potential drugs, also contain other pharmacophore elements, in particular acceptors and hydrogen bond donors, in positions 1 and / or 2.

Derivatives of imidazolininols of general formula A are described, which can be considered as structural analogues of the claimed compounds [WO 00/78750 A1, 2000, C07D 403/04], however, they contain an aromatic fragment of imidazolone instead of imidazolidinone, as in the case of compounds of formula I, and are obtained by intramolecular condensation of the corresponding compounds of formula B. Obtaining compounds of formula I in this way is not possible. Compounds A exhibit physiological activity and are recommended for use in the treatment of inflammatory, immune and several other diseases.

where R ₁ = H, C ₁ -C ₆ alkyl, fluoro-substituted C ₁ -C ₆ alkyl, phenyl, carboC ₁ -C ₆ alkoxy;

R ₂ = H, C ₁ -C ₆ alkyl, aminoC ₁ -C ₆ alkyl, hydroxy-C ₁ -C ₆ alkyl;

R ₃ = H, C ₁ -C ₆ alkoxy; R ₄ = H, hydroxy-C ₁ -C ₃ alkyl or amidinothio-C ₁ -C ₃ alkyl.

A known method of producing derivatives of indoles by amidoalkylation, which consists in the reaction of α-hydroxyalkylamides (structurally similar to the claimed) with indole [Sviridova LA, Afanasyeva SV, Golubeva GA, Terentyev PB, Bundel Yu. G. CGS, 1990 (9), 1207-1213]. However, to extend this method to obtain 3- (2-oxoimidazolidinyl-5) indoles failed. In addition, under the conditions described by the authors, a 2,3-Planchet rearrangement characteristic of the indole derivatives [Ciamician G., Plancher G. Chem. Ber., 29, 2475 (1896)], and a difficultly separable mixture of 2- and 3-derivative indoles is formed.

Closest to the claimed method is a known method for producing bis-heterocyclic compounds based on indole, specifically a method for producing pyrrolidinylindoles, analogues of bisheterocycles of formula I [Sadovoy A.V., Kovrov A.E., Golubeva G.A., Sviridova L.A. Chem. Heterocycl. Comp., 2011, 46, (10), 1215]. However, this method has been tested only for derivatives of cyclic acid amides and has limitations from the point of view of the design of physiologically active bisheterocyclic derivatives of indoles, which is due to the low content of aliphatic heteronuclei capable of entering into such interactions.

The present invention is the creation of previously unknown 3- (2-oxoimidazolidinyl-5) indoles, including a combination of pharmacophore elements of two different classes of heterocycles - indoles and 2-imidazolidinones.

The combination of pharmacophore elements such as indole and imidazolidine nuclei in the structure of the claimed 3- (2-oxoimidazolidinyl-5) indoles of formula I can provide a wide range of their physiological activity.

EFFECT: new compounds with anti-inflammatory activity are obtained, which can be used in medicine, and a technologically advanced method for their preparation is developed.

The problem is solved by new compounds 3- (2-oxoimidazolidinyl-5) indoles of the general formula I,

where R ¹ = H, C ₁ -C ₆ alkyl; R ² = H, C ₁ -C ₆ alkyl, C ₆ H ₄ X, where X = H, Me; R ³ = aryl with anti-inflammatory activity, as well as the method of their preparation.

The inventive method includes the interaction of 5-hydroxyimidazolidin-2-ones of General formula II

где R³ = арил,

where R ³ = aryl,

with an equimolar amount of indole of formula III

where R ¹ = H, C ₁ -C ₆ alkyl; R ² = H, C ₁ -C ₆ alkyl, C ₆ H ₄ X, X = H, Me,

in the presence of 1-10 mol. % boron trifluoride etherate in tetrahydrofuran at room temperature.

The inventive method allows to obtain compounds of formula I in one stage, it is based on the electrophilic substitution of indoles of general formula III at the most active position 3 in the interaction with 5-hydroxyimidazolidin-2-one II (Scheme 1) in the presence of catalytic amounts of boron trifluoride etherate.

The reaction is carried out under mild conditions: at room temperature in THF for a short time (0.5-3 h), and the claimed 3- (2-oxoimidazolidinyl-5) indoles I are obtained in 19-32% yields.

The undoubted advantage of the proposed method is the ability to obtain compounds of formula I in one step from the available starting compounds - 1- and / or 2-substituted indoles III and easily synthesized from glycine hydroxyimidazolidin-2-ones II [Cortes S., Kohn N., J. Org. Chem., 1983, 48 (13), 2247] (Scheme 2), using a standard catalyst, boron trifluoride etherate.

A technological method is proposed for producing indole derivatives of general formula I, which does not require heating of the reaction medium, the use of special equipment, involves the regeneration of spent solvents and the ability to scale the process without reducing the yield of the target product.

In FIG. Figure 1 presents the results of a study of the anti-inflammatory effect of Ia-If compounds on a BV-2 cell line treated with 1 μg / ml bacterial lipopolysaccharide, 18 hours of incubation, Gris reaction, mean ± standard deviation (based on 3 experiments).

In FIG. 2 shows data on the cytotoxicity of compounds Ia-If in relation to cells of the human neuroblastoma SH-SY5Y (24 hr incubation), MTT test, the average value ± standard deviation (based on 3 experiments).

In FIG. Figure 3 presents the results of a study of the cytotoxicity of Ia-If compounds with respect to BV-2 mouse microglia cells (24 hr incubation), MTT test, mean ± standard deviation (based on 3 experiments).

Anti-inflammatory effect

Under the conditions of the induction of an inflammatory response by bacterial lipopolysaccharide on the mouse microvial BV-2 cell line, anti-inflammatory activity was recorded for all compounds (Fig. 1). Compounds Ie and Id were most active, providing suppression of up to 50% of the response even in the low concentration range from 0.1 to 10 μM.

Cytotoxicity

For SH-SY5Y neuroblastoma cells, the compounds were low toxic (Fig. 2). Compounds Ia, Ib and Ic did not show toxic effects up to a concentration of 100 μM. For compounds Ie, Id, If, cytotoxicity was recorded in the concentration range from 10 to 100 μM with an EC _{50 of} 147 ± 10, 95 ± 5, 57 ± 5 μM, respectively (average value ± standard deviation, based on 3 experiments).

EC ₅₀ values were calculated using GraphPad Prism 6.01 software by approximating the experimental data of a curve described by the Hill formula:

where E is the observed effect, n _H is the Hill coefficient characterizing the slope of the curve, and A is the concentration of the substance.

For line BV-2 also low cytotoxicity was recorded (Fig. 3). Substances Ia and Ic were again non-toxic, compounds Ie, Id and Ib caused the death of 20 to 40% of cells at a concentration of 100 μM. If compound showed significant toxicity in the concentration range of 10-100 μM with EC ₅₀ = 19 ± 3 μM (mean ± standard deviation, based on 3 experiments).

The invention is illustrated by the specific embodiments below.

¹ H NMR spectra of the obtained compounds were recorded on Bruker Avance 400 (operating frequency 400 MHz) and Agilent 400-MR (operating frequency 400 MHz) in DMSO-d ₆ , ¹³ C NMR spectra on a Bruker Avance ™ 600 instrument (operating frequency 150.93 MHz). Elemental analysis was performed in the laboratory of elemental analysis of INEOS RAS. Melting points were determined on an Electrothermal IA 9100 instrument. The reaction progress and product purity were monitored by TLC on Silufol-UV ₂₅₄ wafers with a fixed layer of silica gel. The eluent CHCl ₃ - Meon (10: 1), the manifestation of iodine vapor and Kovac's reagent. Solvents were purified by standard methods. The starting compound 1-phenyl-5-hydroxyimidazolidin-2-one (II) was obtained by reduction of LiAlH _{4 of the} corresponding hydantoin derivative synthesized from glycine and phenylisocyanate [Cortes S., H. Kohn, J. Org. Chem., 1983, 48 (13), 2247]. The yield is 56%. T. pl. 112-114 ° C. IR spectrum, υ, cm ^-1 : 1700 (С = O). ¹ H NMR spectrum, δ, ppm (DMSO _d6 ) 3.08 (1Н, d, J = 3 Hz, Н-4), 3.58-3.62 (1Н, m, Н-4 '), 5.58-5.62 ( 1H, dd, H-5), 6.42 (1H, d, OH), 6.95-7.05 (2H, m, Ph), 7.24-7.30 (2H, m, Ph), 7.35 (1H, s, NH), 7.60 (2H, d, Ph).

General Procedure for the Preparation of Ia-If Compounds

1 mmol of 1-phenyl-5-hydroxyimidazolidin-2-one (II) and 1 mmol of indole III are dissolved in 10 ml of dry tetrahydrofuran. With stirring, a few drops (1-10 mol.%) Of boron trifluoride etherate are added. The reaction mixture is stirred for half an hour to three hours, the precipitate formed is filtered off and washed with a small amount of diethyl ether. The filtrate was evaporated, the resulting oil was crystallized from ethyl alcohol.

Example 1

3- (1-Phenyl-2-oxoimidazolidinyl-5) -1H-indole (Ia)

To a solution of 0.188 g (1 mmol) of 1-phenyl-5-hydroxyimidazolidin-2-one II and 0.117 g (1 mmol) of indole III in 10 ml of tetrahydrofuran, 5 mol are added with stirring. % boron trifluoride etherate. The reaction mixture was stirred for 1 hour, then the precipitate formed was filtered off and washed with a small amount of diethyl ether. The combined filtrate was evaporated, the resulting oil was crystallized from ethyl alcohol. Obtain 0.083 g of 3- (1-phenyl-2-oxoimidazolidinyl-5) -1H-indole (Ia). Yield 30%. T. pl. 244 ° C. IR spectrum, υ, cm ^-1 : 1680 (C = O); 3350 (NH). Found,%: C 71.41; H 5.46; N 14.23. 2C ₁₇ H ₁₅ N ₃ O × H ₂ O. Calculated,%: C 71.31; H 5.63; N, 14.68. Mass spectrum, m / z: 277 [M] ⁺ .

¹ H NMR spectrum, δ, ppm: 3.35-3.37 (1Н, m, Н-4), 3.84-3.88 (1Н, m, Н-4 '), 5.72 (1Н, dd, J = 6.23 Hz , J = 9.24 Hz, H-5), 6.87 (1H, t, J = 7.36 Hz, p-Ph), 6.95 (1H, s, NH _imid ), 6.97-7.00 (1H, m, Ind), 7.06- 7.08 (1H, m, Ind), 7.13-7.15 (2H, m, m-Ph), 7.31 (1H, s, 2-Ind), 7.33-7.34 (1H, d, J = 7.66 Hz, Ind), 7.46 -7.47 (2H, d, J = 8.01 Hz, o-Ph), 7.62-7.63 (1H, d, J = 7.66 Hz, Ind), 10.87 (1H, s, NH _ind ). ¹³ C NMR spectrum, δ, ppm: 45.80, 53.86, 112.25, 114.58, 119.15, 119.34,120.88 (2С), 121.70, 122.67, 124.57, 125.46, 128.42 (2С), 137.28, 140.22, 159.62.

Example 2

1-Methyl-3- (1-phenyl-2-oxoimidazolidinyl-5) indole (Ib)

Analogously to example 1, from 0.178 g (1 mmol) of 1-phenyl-5-hydroxyimidazolidin-2-one II and 0.131 g (1 mmol) of 1-methylindole III, after 0.5 hours of stirring, 0.073 g of 1-methyl-3- (1-phenyl -2-oxoimidazolidinyl-5) indole Ib. Yield 25%. T. pl. 233 ° C. IR spectrum, υ, cm ^-1 : 1695 (C = O). Found,%: C 71.77; H 5.72; N 13.63. 2C ₁₈ H ₁₇ N ₃ O × H ₂ O. Calculated,%: C 71.98; H 6.04; N 13.99. Mass spectrum, m / z: 291 [M] ⁺ .

¹ H NMR spectrum, δ, ppm: 3.30 (1Н, m, Н-4), 3.68 (3Н, s, N-CH ₃ ), 3.82 (1Н, t, J = 9 Hz, Н-4 '), 5.71 (1H, dd, J = 6.06 Hz, J = 9.20 Hz, H-5), 6.87 (1H, t, J = 7.36 Hz, p-Ph), 7.01-7.03 (1H, m, Ind) , 7.11-7.16 (1H, m, Ind, 2H, m, m-Ph), 7.33 (1H, s, 2-Ind), 7.35-7.37 (1H, d, J = 7.7 Hz, Ind), 7.46-7.47 (2H, d, J = 8.03 Hz, o-Ph), 7.61-7.62 (1H, d, J = 7.7 Hz, Ind).

¹³ C NMR spectrum, δ, ppm: 32.79, 45.63, 53.17, 53.21, 110.51, 113.47, 119.34, 119.51, 120.60 (2С), 121.86, 122.68, 125.59, 128.52 (2С), 128.83, 137.50, 139.86 159.50.

Example 3

2-Methyl-3- (1-phenyl-2-oxoimidazolidinyl-5) -1H-indole (Ic)

Analogously to example 1, from 0.178 g (1 mmol) of 1-phenyl-5-hydroxyimidazolidin-2-one II and 0.131 g (1 mmol) of 2-methylindole III, after 2 hours of stirring, 0.081 g of 2-methyl-3- (1-phenyl -2-oxoimidazolidinyl-5) -1H-indole Ic. Yield 28%. T. pl. 275 ° C. IR spectrum, υ, cm ^-1 : 1681 (C = O); 3240 (NH). Found,%: C 74.51; H 6.27; N 14.32. C ₁₈ H ₁₇ N ₃ O. Calculated,%: C 74.20; H 5.88; N 14.32. Mass spectrum, m / z: 291 [M] ⁺ .

¹ H NMR spectrum, δ, ppm: 2.41 (3Н, s, СН ₃ ), 3.34 (1Н, m, Н-4), 3.76 (1Н, m, Н-4 '), 5.72 (1Н, dd, H-5), 6.84-6.87 (1H, m, p-Ph), 6.90-6.92 (1H, m, Ind), 6.95 (1H, m, Ind), 7.11-7.16 (2H, t, J = 7.8 Hz, m-Ph), 7.19-7.20 (1Н, d, J = 7.9 Hz, Ind), 7.33-7.34 (2Н, d, J = 8.0 Hz, o-Ph), 7.48-7.50 (1Н, d, J = 7.9 Hz, Ind), 10.88 (1H, s, NH _ind ).

¹³ С NMR spectrum, δ, ppm: 11.71, 44.86, 52.53, 109.21, 111.13, 118.28, 119.12, 120.81, 120.86, 122.81, 128.39 (4С), 133.79, 135.62, 139.74, 159.52.

Example 4

1,2-Dimethyl-3- (1-phenyl-2-oxoimidazolidinyl-5) indole (Id)

Analogously to example 1, from 0.178 g (1 mmol) of 1-phenyl-5-hydroxyimidazolidin-2-one II and 0.145 g (1 mmol) of 1,2-dimethylindole III, after 2 hours of stirring, 0.058 g of 1,2-dimethyl-3- is obtained (1-phenyl-2-oxoimidazolidinyl-5) indole Id. Yield 19%. T. pl. 278 ° C. IR spectrum, υ, cm ^-1 : 1699 (С = O). Found,%: C 73.18; H 6.07; N 13.32. 2C ₁₉ H ₁₉ N ₃ O × H ₂ O. Calculated,%: C 72.59; H 4.41; N 13.37. Mass spectrum, m / z: 305 [M] ⁺ .

¹ H NMR spectrum, δ, ppm: 2.45 (3Н, s, 2-СН ₃ ), 3.30-3.34, (1Н, m, Н-4), 3.58 (3Н, s, N-CH ₃ ) , 3.78-3.82 (1Н, m, Н-4 '), 5.76-5.80 (1Н, m, Н-5), 6.84-6.86 (1Н, m, p-Ph), 6.94-6.96 (1Н, m, Ind ), 7.02-7.04 (1Н m, Ind), 7.11-7.13 (2Н, m, m-Ph), 7.30-7.33 (1Н, m, Ind; 2Н, o-Ph), 7.51 (1Н, d, J = 7.9 Hz, Ind).

¹³ С NMR spectrum, δ, ppm: 10.33, 29.67, 44.97, 52.80, 109.30, 109.84, 118.42, 119.41 (2С), 120.91 (2С), 122.85,128.83 (3С), 135.54, 137.01, 139.74 159.89 .

Example 5

2-p-Tolyl-3- (1-phenyl-2-oxoimidazolidinyl-5) -1H-indole (Ie)

Analogously to example 1, from 0.178 g (1 mmol) of 1-phenyl-5-hydroxyimidazolidin-2-one II and 0.207 g (1 mmol) of 2-p-tolylindole III, after 2 hours of stirring, 0.118 g of 2-p-tolyl-3- is obtained (1-phenyl-2-oxoimidazolidinyl-5) -1H-indole Ie. Yield 32%. T. pl. 317 ° C. IR spectrum, υ, cm ^-1 : 1684 (C = O); 3282 (NH). Found,%: C 74.27%; H 5.69; N, 10.70. C ₂₄ H ₂₃ N ₃ O. Calculated,%: C 74.78; H 6.01; N, 10.90. Mass spectrum, m / z: 367 [M] ⁺ .

¹ H NMR spectrum, δ, 2.42 (3Н, s, Tol - С Н ₃ ), 3.54-3.58 (1Н, m, Н-4), 4.01-4.06 (1Н, m, Н-4 '), 5.59- 5.63 (1Н, m, Н-5), 6.77-6.79 (1Н, m, p-Ph), 6.94-7.08 (4Н Ph, m, 1Н, m, Ind), 7.02-7.04 (1Н, m, Ind) 7.21 (1H, s, NH _imid ), 7.28-7.31 (1H, m, Ind), 7.38-7.46 (4H, m, Tol), 7.57-7.59 (1H, m, Ind), 11.32 (1H, s, NH _ind ).

¹³ С NMR spectrum, δ, ppm: 21.34, 44.93, 52.85, 109.79, 111.96, 119.47, 119.65, 120.32 (2С), 121.95, 122.60, 128.26 (3С), 129.03 (2С), 129.42 129.93 (2С ), 136.59, 137.59, 138.13, 139.63, 159.40.

Example 6

1-Methyl-2-p-tolyl-3- (1-phenyl-2-oxoimidazolidinyl-5) indole (If)

Analogously to example 1, from 0.178 g (1 mmol) of 1-phenyl-5-hydroxyimidazolidin-2-one II and 0.221 g (1 mmol) of 1-methyl-2-p-tolyl-indole III, after 3 hours of stirring, 0.107 g of 1- methyl -2-p-tolyl-3- (1-phenyl-2-oxoimidazolidinyl-5) indole If. Yield 28%. T. pl. 183 ° C. IR spectrum, υ, cm ^-1 : 1701 (С = O). Found,%: C 78.44; H, 6.02; N, 10.65. C ₂₅ H ₂₃ N ₃ O. Calculated,%: C 78.71; H 6.08; N 11.02. Mass spectrum, m / z: 381 [M] ⁺ .

¹ H NMR spectrum, δ, ppm: 2.45 (3Н, s, Tol - CH ₃ ), 3.43-3.45, (1Н, m, Н-4), 3.49 (3Н, s, N-СН ₃ ) , 3.88-3.91 (1Н, m, Н-4 '), 5.19 (1Н, m, Н-5), 6.80-6.82 (1Н, m, p-Ph), 7.00-7.06 (2Н, m, Ph; 2Н , m, Tol; 1Н, m, Ind), 7.13-7.16 (1Н, m, Ind), 7.31-7.33 (2Н, m, Ph), 7.41-7.45 (1Н, m, Ind; 2Н, m, Tol) 7.60-7.62 (1H, m, Ind).

¹³ С NMR spectrum, δ, ppm: 21.40, 30.57, 45.30, 54.45, 109.57, 110.16, 119.75, 119.99, 122.01, 122.39, 123.83, 125.12, 127.70, 128.20 (4С), 129.34 (2С), 130.39 , 137.36, 138.38, 138.91, 140.09, 160.17.

Methods for determining the cytotoxicity and anti-inflammatory effects of compounds Ia-If

For biological experiments, DMEM, L-glutamine, penicillin, streptomycin, amphotericin B, MTT, trypsin solution in EDTA, Earl solution, D-glucose, Versen solution from Paneko, Russia were used; DMSO, lipopolysaccharide, sulfonamide, naphthylethylenediamine and Triton X-100 from Sigma-Aldrich, USA; isopropanol and HCl company Himmed, Russia; fetal cow serum from BioSera, France.

The experiments were performed on cells of mouse microglia of the BV-2 line (CVCL_0182) and human neuroblastoma of the SH-SY5Y line (ATCCCRL-2266). Cells were cultured in DMEM, to which the following components were added to the following final concentrations: L-glutamine - up to 4 mM; fetal cow serum - up to 10%; penicillin - up to 100 units / ml; streptomycin - up to 100 μg / ml and amphotericin B - up to 2.5 μg / ml. All cell lines were cultured at 37 ° C in a humidified atmosphere with 5% CO ₂ content.

To evaluate the anti-inflammatory effect, on the eve of the experiment, cells in a culture medium were seeded in a 96-well plate with a density of 15 thousand per well (100 μl per well). Inflammation was induced by bacterial lipopolysaccharide (LPS). For this, 100 μl of fresh culture medium containing LPS (1 μg / ml) and the test substance at a concentration of 100, 10, 1, and 0.1 μM (obtained by serial dilution from stock solution in DMSO) were added to 100 μl of the old medium in wells. Cells were incubated for 18 hours, after which the inflammatory response was assessed by the accumulation of nitric oxide metabolism product NO ₂ ^- in the medium. The concentration of nitrite anion was measured according to the Gris method. For this, 12.5 μl of a 0.04% aqueous solution of sulfanilamide was added to 75 μl of the analyzed mixture, kept at room temperature for 10 min with protection from light, then 12.5 μl of a 2% solution of naphthylethylenediamine in 3M HCl in water was added, kept for another 10 min at room temperature with protection from light, after which the optical absorption at a wavelength of 540 nm was determined. As a control, cell samples were used only with culture medium and with culture medium supplemented with LPS.

To assess cytotoxicity, cells (15 thousand cells in 100 μl of culture medium) were scattered into 96-well plates with a density of 15 thousand per well. 100 μl of the solutions of the test substances in the culture medium prepared by serial dilution of their stock solutions in DMSO to final concentrations of 100, 10, 1, and 0.1 μM (final DMSO content not more than 0.5% by volume) were added to the wells. The control contained only DMSO, the positive control - 0.5% Triton X-100. Cells were incubated at 37 ° C in a humidified atmosphere with 5% CO ₂ for 24 hours. Cell viability was assessed using the MTT test. For this, the medium was removed and replaced with a MTT solution in Earl liquid (0.5 mg / ml) with the addition of 1 g / L D-glucose and incubated for 1.5 h in a CO ₂ incubator. After that, an equal volume of 0.04 M HCl in isopropanol was added to the wells and incubated with stirring at 37 ° C for 30 min. The optical density of the resulting solution was evaluated at wavelengths of 594 and 620 nm using an Efos 9304 apparatus (MZ Sapphire, Russia). The percentage of cells that survived under the action of a specific dose of the test compound was calculated as the quotient of dividing the average optical density in the wells after incubation with this dose by the average optical density in the control wells. The action of each compound in all concentrations was studied three times and average data were obtained from the results of three experiments.

Claims (8)

1. The method of obtaining 3- (2-oxoimidazolidinyl-5) indole of General formula I

where R ¹ = H, C ₁ -C ₆ alkyl; R ² = H, C ₁ -C ₆ alkyl, C ₆ H ₄ X, where X = Me; R ³ = phenyl, including the interaction of 5-hydroxyimidazolidin-2-one of General formula II

where R ³ = phenyl, with an equimolar amount of indole of the general formula III

where R ¹ = H, C ₁ -C ₆ alkyl; R ² = H, C ₁ -C ₆ alkyl, C _b H ₄ X, where X = Me, in the presence of 1-10 mol. % boron trifluoride etherate in tetrahydrofuran at room temperature.

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