Derivatives of boron-containing peptides

 

(57) Abstract:

Purpose: in biochemistry and medicine, as an inhibitor of trypsin-like serine proteases, such as thrombin, kallickrein plasma and plasmin. The essence of the invention: derivatives of boron-containing peptides. See f-La. Reagent 1: the corresponding amine. Reagent 2: R - Z - OH. The condensation is carried out in the solvent in the presence of triethylamine. The resulting product is treated with alkali metal azide, followed by reduction in the presence of an organic or mineral acid, preferably benzosulfimide and interaction with cyanamide in a lower alcohol at 50 to 150°C. 18 table. Formula: where R1= H, Ac, Br, Boc, BocHu; R2=(CH2)nNHC(NH)NH2; n= 3,4; y - balance bingonova and piandiong ether; HW = C6H5SO3, HCl, HBr; Z = D - or L-Phe, DPhe - Pro, DPhe - Phe, Ala - huS, Pro - Phe, Ala - Phe, Glu - Phe, Ala - Ghe, Glu - Gly, Gly - Leu - Ala, Pyro - Glu - Phe, Val - Val, (D)Val - Leu, Lys - Pro, Leu - Thr. 18 table.

The invention relates to boron-containing peptides, a new biologically active compounds that may find application in biochemistry as inhibitors trypsinogen serine proteases, such as thrombin, kallickrein plasma and plasmin.

Known inhibitors slow the coronary thrombosis in a rabbit model [1].

Substituted argininemia consisting of secondary amines, for example (2R, 4R)-4-methyl-1 {N2-[(3-methyl-1,2,3,4-tetrahydro-8-chinoline)-sulfonyl]-L-argin - yl}- 2-piperidinecarbonitrile acid [2], are inhibitors of thrombin (Ki = 19 nm).

This inhibitor increases prothrombin time plasma analyses on blood coagulation in vitro in 2 times at 1 μm and is fibrinoliticeski amplifying means for use in combination with tissue plasminogen activator.

The most effective of the known thrombin inhibitors is N -/2-attilalongoria/-4-lidinopril-alanine-piperidine [3] (Ki = 6 nm). Its effectiveness is determined in mice and rats in vivo.

The purpose of this proposal is to create new and derivative peptide - low-emission, with a higher inhibitory activity and a broader spectrum of actions, solutions which can be prepared without the use of a solubilizer (organic solvent).

This goal is achieved by the described derivatives of boron-containing peptides of General formula

RZNHBYHW, (I) where R1= H, Ac, Bz, Boc, Boc-Leu;

R2= (CH2)nNHC(NH)NH2;

n = 3, 4;

Y = the remainder pincavage he, Ala-Glu, Glu-Gly, Gly-Leu-Ala, Pyro-Glu-Phe, Val-Val, To Dval-Leu, Lys-Pro, Leu-Thr. Get them, for example, by the interaction of the amine of the formula

H2N

(II) where n = 3, 4;

W and WI= Cl, Br;

Y is the residue bingonova or piandiong ether, enter into interaction with the amino acid or peptide of the formula

R1-Z-OH, (III) where the values of R1and Z above, and the resulting product of the formula

RZNW

(IV) where the values of R1, Z, Y, W, WIand n as mentioned above, is treated with alkali metal azide to form compounds of formula

RZNHW

(V) where the values of R1, Z, Y, W and n as mentioned above, which is then restored in the presence of an organic or mineral acid, preferably benzosulfimide, with the formation of compounds

RZNH

(VI) the values of R1, Z, Y, W and n described above, and the latter is subjected to interaction with cyanamide in a lower alcohol at a temperature of 50-150aboutC.

In the description, the following abbreviations are used for amino acid residues:

Ala = L-alanine

Arg = L-arginine

Asn = L-asparagine

Asp = L-aspartic acid

Gay = L-cysteine

Gln = L-glutamine

Glu = L-glutamic acid

Gly = glycine

His = L-histidine

Ile = L-isoleucine

Leuhr = L-threonine

Trp = L-tryptophan

Tyr = L-tyrosine

Val = L-valine

Where the extended prefix D, the above-mentioned abbreviations indicate the amino acid of D-configuration. Where the extended prefix D or L, the above-mentioned reduction indicate that the amino acid may or configuration D, or the configuration of L.

Used the term "N-terminal protective group" refers to aminocentesis protective groups used in peptide synthesis. Suitable groups include acyl protective group, acetyl (AC), benzoyl (Bz) or aliphatic urethane protective group, such as tertbutoxycarbonyl (Vos).

The following description of examples illustrate specific embodiments of the invention. All these melting points are not corrected. All parts are given in mass units, and the temperature specified in theaboutC. Chemical shifts in the spectroscopy proton nuclear magnetic resonance (NMR or1HNMR) indicated in Delta units, parts per million, based on internal tetramethylsilane standard. Numerous abbreviations used in these examples have the following meanings: FA - triperoxonane acid; DMF IS N,N-dimethylformamide; MS - mass spectromet the safety of the group of boric acid have abbreviations: -C6H12= bingonova group and-C10H16= pirandola group.

All amino acid residues are represented in the L-configuration unless otherwise stated.

TLC and RP-TLC) carried out on plates of silica gel 60 firm E. Merck and plates I18F for off-TLC firm Whatman. The neutral compound was observed in the UV region and after exposure to vapors of iodine. Compounds containing free amino groups, tinted NIN hydrinos and connection guanidinopropionic tinted dye Sakaguchi. Dye Sakaguchi reveals significant specificity to monosubstituted to guanidinum, for example contained in bargining peptides.

P R I m e R 1A. 1-Amino-4-bromo-butylboronic financial HCl.

NH2-CH[(CH2)3Br]BO2-C10H16HCl

4-Bromo-1-chlorobutyronitrile receive according to the method described Matte Son and others In the normal experiment exercise hydroporinae of allylbromide (173 ml, 2.00 mol) of borane pyrocatechin (240 ml, 2.00 mol) by the addition of borane to allylbromide followed by heating the reaction mixture for 4 h to 100aboutC in nitrogen atmosphere. The resulting product, pyrocatechin 3-bromopropionate (so Kip. 95-102aboutC, 0, what nandyala (88 g, 0.52 mol) when mixing them in 50 ml of tetrahydrofuran (THF), intensively stirring this mixture for 0.5 h at 0aboutWith and then for 0.5 h at room temperature. The solvent is evaporated and add 250 ml of hexane. Pyrocatechin removed in the form of a crystalline solid. Quantitative output obtained by serial dilution with hexane to volume of 500 and 1000 ml emitting crystals at each dilution. In the process of evaporation of the solvent receive the product (147 g) in the form of butter.

C13H22O2BrB.

Calculated, %: C 51,85; H 7,38; Br 26,54.

Found, %: C 52,85; H 7,30; Br 26,58.

4-Bromo-1-chlorocarbonate pinanediol get homologation of the corresponding propylmalonate. Methylene chloride (34.8 ml, 0,540 mol) is dissolved in 500 ml of THF and then slowly added to 1.54 M n-utility in hexane (350 ml, 0,540 mmol) at a temperature of -100aboutC. Pinanediol 3-bromopropionate (148 g, 0,490 mol) is dissolved in 500 ml of THF, cooled to the freezing point of the specified solution and injected into the reaction mixture. Zinc chloride (33,5 g, 0,246 mol) is dissolved in 250 ml of THF, cooled to 0aboutWith and added in several portions to the reaction mixture. The reaction mixture while stirring nagrevayut in hexane and washed with water. After drying over anhydrous magnesium sulfate and filtering, the solvent is removed to yield the desired product (140 g).

Pinanediol 1-amino-4-bromobutyrate obtained by dissolution of hexamethyldisilizane (28,0 g, 80,0 mmol) in 30 ml of THF, cooling the resulting solution to -78aboutWith and adding to it 1,62 N. n-utility in hexane (49,4 ml, 80,0 mol). Then the solution is slowly warmed to room temperature, re-cooled to -78aboutWith and add pinanediol 4-bromo-1- -chlorocarbonate (28,0 g, 80,0 mol) in 20 ml of THF. The reaction mixture is slowly warmed to room temperature and stirred over night. The solvent is removed by evaporation, enter dry hexane (400 ml) and obtain the residue, which is marked by filtration in a nitrogen atmosphere. The filtrate is cooled to -78aboutWith and add 4n. HCl in dioxane (60 ml, 240 mol). Then the reaction mixture is slowly warmed to room temperature and at this temperature, stirred for 2 hours the resulting product (20 g) was isolated by filtration as a solid substance. After drying in vacuum the crude product is dissolved in chloroform and nerastvorimaya substance produce by filtration. The filtrate is evaporated and the residue dissolved in ethyl acetate. Clevo is; = 1,0 in absolute methanol.

C14H26NO2BrClB.

Calculated, %: C 45,87; H 7,16; N 3,82; B 2,95.

Found, %: C 45,76; H 7,21; N 3,79; B 3.04 From.

P R I m e R 1b.

(D,L) 1-amino-4-bromobutyronitrile HCl.

(D,L) NH2-CH[(CH2)3Br]BO2-C6H12HCl.

Pinacol 4-bromo-1-chlorocarbonate receive according to the method described to obtain the corresponding pinanediol (example Ia), except that instead of pinanediol use pincon, and pinako 3-bromo-propylmalonate (so Kip. 60-64aboutC, 0.35 mm) and 4-bromo-1-chlorocarbonate pincon (so Kip. 110-112aboutWith, 0.20 mm) is distilled.

C10H19O2BrClB.

Calculated, %: C 40,38; H 6,45.

Found, %: C 40,70; H 6,37.

Hydrochloric salt of pinecone of 1-amino-4-bromobutyrate also receive according to the method of example 1A. Target product vykristallizovyvalas from a mixture of ethyl acetate:hexane his 52%.

C10H22NO2BrClB.

Calculated, %: C 38,19; H 7,05; 4,45 N; Cl 11,27; Br 25,41.

Found, %: C 38,28; H 7,39; N 4,25; Cl 11,68; Br 26,00.

P R I m e R 1C. Hydrochloric salt of pinecone 1-amino-4-chlorotoluron.

(D,L)NH2-CH[(CH2)3Cl]B 3-chloro - propyl ester Bronevoy acid (so Kip. 63aboutWith, 0.20 mm) are obtained by the method of example 1A, except that instead of allylbromide use allylchloride, and instead pinanediol take pincon.

C9H18O2ClB.

Calculated, %: C 52,85; H 8,89; Cl 17,33.

Found, %: C 53,41; H 8,15; Cl Value Of 16,81.

The homologation is also carried out according to the method of example 1A, and the resulting product are distilled (so Kip. 95aboutWith, 0.25 mm) with 65% yield.

WITH10H19ABOUT2Cl2B.

Calculated, %: C 47,47; N 7,58; Cl 28,02.

Found, %: C 47,17; H 7,45; Cl 27,75.

Hydrochloric salt of pinecone of 1-amino-4-chlorbutanol ether Bronevoy (boric) acid get by the method similar to example 1A. Target product vykristallizovyvalas from ethyl acetate to obtain 8.8 g of product (so pl. 132-135,5about(C) and 2.2 g of the product (so pl. 145-147aboutC). The product so pl. 145-147aboutTo use for analysis.

C10H22NO2ClB.

Calculated, %: C 44,47; H 8,23; N 5,19; B 4,00.

Found, %: C 44,01; H 8,23; N 4,77; B 3,80.

P R I m e R 1d (D,L) 1-amino-5-bromatylogo ether Bronevoy acid pinacol HCl.

(D,L) NH2-CH[(CH2)4Br]BO2C6H12HCl.

Pinacol 4-Brom the ester of boric acid (example 1A), except that allylbromide replace 4-bromo-1-butene, and financial - pincon. The product is isolated in the form of oil (so Kip. 77aboutC, 0.3 mm). After the homologation get 5-bromo-1-globetronica ether Bronevoy acid pinacol.

Mass spectroscopy (Cl): C11H21O2BrClB.

Computed-N: 310,47.

Found, -N: 310.

Target product hydrochloric salt of pinecone 1-amino-5-bromatylogo ether Bronevoy acid get by the method of example 1A at exit 35%.

WITH11H24NO2BrBCl.

Calculated, %: C 40,22; H Of 7.36; N, 4.26 Deaths; Cl 10,79; Br 24,32; B 3,29.

Found, %: C 39,23; H 7,18; N 4,04; Cl 15,21; Br 25,66; B 3,75.

P R I m m e R 2. Boc-(D)Phe-Pro-NH - CH[(CH2)3Br]BO2-C10H16.

The synthesis of the dipeptide Boc-(D)Phe-Pro-OH carry out first obtain a dipeptide benzyl ether, and then remove the specified complex ester by catalytic hydrogenation. Boc-(D) -Phe-OH (10.0 g, of 37.7 mmol) is dissolved in 50 ml of THF and the resulting solution was added N-methylmorpholine (4,14 ml of 37.7 mmol). The solution is cooled to -20aboutWith, and then thereto was added isobutyl ether of Harborview acid (4,90 ml of 37.7 mmol). After 5 min the reaction mixture was injected H-Pro-1 OBr-.HCl (9,11 g) and the mixture was stirred for 1 h at -20aboutC and 2 h at room temperature. The reaction mixture is filtered and the filtrate evaporated. The residue is dissolved in ethyl acetate followed by washing with 0.2 N. HCl, 5% aqueous sodium bicarbonate solution and saturated aqueous sodium chloride. The organic layer is dried over anhydrous sodium sulfate, filtered and evaporated to obtain 15.2 g of Boc-(D) Phe-Pro-OBz 1 in the form of oil. The obtained benzyl ester (15.2 g) was dissolved in 100 ml of methanol and then hydronaut at an initial pressure of 40 pounds/inch2(2,812 kg/cm2) in a Parr apparatus in the presence of 0.5 g of 10% palladium mobiles (Pd/C). The reaction solution is filtered through zeolit and evaporated to yield a solid substance. Specified solid product is isolated and washed with ethyl acetate, and then a simple ether to obtain 10.0 g of the desired product (so pl. 176,5-177aboutC).

C19H26N2O5< / BR>
Calculated, %: C 62,95; H 7,24; N 7,73.

Found, %: C 62,91; H 7,15; N 7,53.

Boc-(D)Phe-Pro-NH-CH[(CH2)3Br] BO2-C10H16get the binding specified dipeptide with an appropriate amine according to the method of the mixed anhydride. The mixed anhydride of Boc - (D)Phe-Pro-OH were obtained when dissolving the specified acid (4.94 g, to 13.6 mmol) in 30 ml THF c posleduushie isobutyl ether of Harborview acid (1.77 ml, to 13.6 mmol). After stirring for 5 minutes at -20aboutSince, in the reaction mixture is injected amine, obtained in example 1A, NH2- -CH[(CH2)3Br]BO2-C10H16Benzosulfimide.

Azide of example 3 (8,80 g of 13.8 mmol) is dissolved in 150 ml of methanol and hydronaut on installing Parra at a pressure of 40 pounds/inch2(2,812 kg/cm2in the presence of 0.50 g of 10% palladium mobiles and benzosulfimide (2,19 g of 13.8 mmol). After one hour, the catalyst was removed, and after evaporation of the solution get to 9.9 g of the desired product. of TLC in the solvent system methanol: water (85: 15) shows the spot in the UV region; Rf0.91 and stain positive for ninhydrin, Rf0,52.

P R I m e R 5. Boc-(D)Phe-Pro-NH - CH[(CH2)3-NH-C(NH)NH2]BO2-C10H16benzene - acid.

Boc-(D)Phe-Pro-broad-C10H16benzosulfimide obtained in example 4 (4.6 g, 6,11 mmol), heated under reflux at 100aboutWith in 20 ml of absolute ethanol containing cyanamide (50 mg/ml). The course of the reaction regulate of TLC in the solvent system methanol:water (85:15), where the observed disappearance of NIN - gikinoo spot of the starting material amine (Rf0,54) and the appearance of the dye Sakaguchi on polucheniya 18 h, moreover, its level gradually rises. After 7 days, when Amin is impossible to detect the reaction solution is concentrated to about 50% of the volume of passive evaporation. Then the reaction solution is filtered, concentrated and chromatographic 2.5 x 100 cm column of LH-20 in methanol. The fractions containing the desired product are pooled and, after evaporation obtain 3.7 g of the desired product. A portion of the product (2.3 g) is crystallized with a mixture of ethyl acetate:hexane, with the release of 0.89 g of the substance, and the residue (1.2 g) obtained by poroshkovaya simple ether in the form of a solid product.

Mass spectroscopy (FAB) for C34H53N6O6SB.

Calculated, % : +H 653,42.

Found, %: +H 653,38.

Elemental analysis for C40H59N6O9SB. H2O, %.

Calculated, %: C 57,95; H 7,43; N 10,14; B 1,30.

Found, %: C 57,20; H 7,14; N 10,94; B 1,01.

P R I m e R 6. H-(D)Phe-Pro-broad- -C10H162HCl.

The product of example 5, Boc-(D)Phe-Pro-broad-C10H16benzosulfimide (1,17 g, 1.54 mmol), subjected to interaction with 5 ml of 4n. hydrogen chloride in dioxane for 15 min at room temperature. The resulting product is precipitated by adding a simple ether, isolated and washed with simple EF the anion-exchange column (Cl--a form of SIV-RAD AGI X8TMRichmond, CA) c followed by washing the column with water (30 ml). Eluent evaporated in vacuo, and the residue proscout simple ether to yield the desired product (0,80 g).

Mass spectroscopy (FAB) for C29H45N6O4B:

Calculated, %: +H 553,37.

Found, %: + H 553,40 and 538,40 (not identified).

Elemental analysis H-(D)Phe-Pro-broad-C10H16I BSA.TFA:

Found, %: 553,4.

P R I m e R s 7-8. Ac-(D)Phe-Pro-broad--C10H16HCl (example 7).

Ac-(D)Phe-Pro-broad-OH HCl (example 8).

Boc-(D)Phe-Pro-broad-C10H16benzosulfimide, the product of example 5 (0,86 g, 1.13 mmol), subjected to interaction with anhydrous triperoxonane acid (TFA) (about 5 ml) for 15 min at room temperature. The excess TFA is removed by evaporation, and the resulting precipitate proscout simple ether to yield 0.76 product. The specified product (0,70 g of 0.91 mmol) dissolved in a mixture consisting of 2 ml of dioxane and 1 ml of water. Acetic anhydride (0,47 ml, 5.0 mmol) and sodium bicarbonate (0,42 g, 5.0 mmol) was added to the reaction solution. Then the resulting mixture was stirred for 20 min at room temperature. Add ethyl acetate (50 ml) and water (5 ml). The resulting f is rites by evaporation obtain 0.56 g of semi-solid substances.

The resulting sample was dissolved in 4 ml of glacial acetic acid, followed by dilution of a solution of 16 ml of water. Then immediately applied on the column with 15 ml SP-cefalexin (N+form and balance 20% solution of acetic acid. The column was washed with 300 ml of 20% acetic acid and establish a linear concentration gradient from 100 ml of 20% acetic acid to 100 ml of 20% acetic acid, hydrochloric acid. Fractions collected in a gradient concentration from 0.08 to 0.17 N. HCl contain N-acetyloxy peptide (0,29 g) in a mixture of free Bronevoy acid and complex piandiong ether.

Ester pinanediol and free Bronevoy acid separated by chromatography on a 2.5 × 100 cm column containing 1H-20 in methanol. Particle size is 8.2 ml. Financially ester (102 mg) elute in fractions 41-43, while the free boric acid (131 mg) gradually wash in fractions 45-129.

Mass spectroscopy (FAB) (example 7):

Ac-(D)Phe-Pro-broad-C10H6for C31H47N6O5B:

Calculated +H: 595,33.

Found +N :595,33.

Mass spectroscopy (FAB) (example 8):

Ac-(D)Phe-Pro-broad-OH HCl for C21H33N6O5:

Calculated +H: 449,60. The P coincide with the structure of free Bronevoy acid, as specified band emission for pinangjawa group, such as methyl singlets groups of 0.85 (3H), OF 1.30 (3H) and 1.36 (3H), were not observed. As additional evidence of this structure, the product sample free Bronevoy acid praeteritorum with the release of the material of example 7. An analytical sample (20 mg) is treated with a twofold excess pinanediol (14 mg) in 3 ml of methanol for 5 minutes, the Solvent evaporated and the excess pinanediol remove poroshkovaya simple ester of the specified sample from the output of the desired product (26 mg).

Mass spectroscopy (FAB) (found: 595,38) and NMR data coincide with the structure proposed for the esterified product and almost identical pinangjawa the product of example 7.

P R I m e R 9. Ac-Phe-broad-C10H16HCl.

According to the method described in example 2, get Ac-Phe-NH-CH[CH2)3Br]BO2- -C10H16. Then get mixed anhydride Ac-Phe-OH (0.565 g, 2,73 mmol) in 10 ml of TFA, which is associated with NH2-CH[(CH2)3Br]BO2-C10H16bansilalpet obtained by processing the compounds Ac-Phe-borough-C10H16benzosulfimide (2.0 g, 3,26 mmol) in 10 ml of a solution of cyanamide (100 mg/ml) in ethanol. RET 3 days and the reaction mixture contains a mixture of starting material and the resulting product. This mixture requires additional purification, which in all probability can not hold, extending the duration of the reaction. The reaction solution is further concentrated and chromatographic in column (2.5 x 100 cm of LH-20 in methanol. The fractions containing the desired product detected using tinted agent Sakaguchi, are combined and evaporated to yield 1.4 g of product. The resulting product (1.2 g) dissolved in 6 ml of acetic acid, diluted with 30 ml of water and get the solution milky color. This solution was applied on a column of 30 ml with SP-Seedcamp C-25 (H+-form), brought to an equilibrium state 20% aqueous solution of acetic acid. Next, the column was washed with 240 ml of 20% acetic acid and establish a linear concentration gradient from 250 ml of 20% acetic acid to 250 ml of 20% acetic acid containing 0,30 N. hydrochloric acid. Faction, erwerbende from the column in the range from 0.12 to 0.16 N. hydrochloric acid, combined with the output of 0.42 g of the desired peptide in a mixture of free boric acid and ether complex pinanediol. This mixture was dissolved in methanol (10 ml) and for the esterification of free boric the Finance sediment simple ether obtain 0.28 g of the desired product.

Elemental analysis for C26H40N5O4B HCl 2H2O, %:

Calculated: C 54,78; H 8,15; N 12,30; B 1,90.

Found: C 55,34; H 7,83; N 11,66; B 1,99.

Mass spectroscopy (FAB) for C26H40N5O4B:

Calculated +H: 498,32.

Found +N: 498,31.

P R I m e R 10. Ac-(D,L)Phe-(D,L)-broad--C6H12.

The intermediate connection of Ac - (D,L)Phe-(D,L)-NH-CH[(CH2)3Br]BO2-C6H12receive a modification of the methods described in example 1 and 2. The acid chloride Ac-Phe-OH were obtained when the interaction of Ac-Phe-OH (30 g, 0,145 mol) pentachloride phosphorous acid (30 g, 0.144 mol) in 175 ml of THF at -10aboutC. the Reaction mixture was stirred at 0aboutWith approximately 1 h, then diluted to a volume of 350 ml of chilled simple ether, the product is isolated in the form of solids, washed with cold simple ether and after drying in a vacuum get 21 g of the product. Activated acetylphenylalanine (14.8 g, 65,6 mmol) dissolved in 40 ml of THF and added to the product of the interaction of pinecone 4-bromo-1-chlorocarbonate and hexamethyldisilizane (prepared in 20 molnau concentration) at -78aboutC. Then the reaction mixture is heated to room temperature with which cetate and washed successively with water, 5% sodium bicarbonate solution and saturated aqueous sodium chloride. The organic phase of the mixture is dried over anhydrous sodium sulfate and after preconcentration get the desired product as a crystalline solid (1,37 g; so pl. 146,5-148aboutC). In the study of crystal structure obtained following chemical composition.

Elemental analysis for C21H32N2O4BrB, %:

Calculated: C 53,98; H 6,92; N 6,00; Br 17,10; B 2,31.

Found: C 54,54; H Is 6.78; N Of 5.89; Br 16,46; B 3,40.

Allylbromide converted into the corresponding azide by the method of example 3. Product vykristallizovyvalas of ethyl acetate (I. pl. 143-144aboutC).

Elemental analysis for C21H32N5O4B, %:

Calculated: C 58,74; H 7,53; N To 16.31; B 2,53.

Found: C 58,85; H Of 7.48; N 16,53; B 2,93.

The obtained azide turn in salt benzosulfimide Ac-(D,L)Phe-(D, L)-borough-C6H12according to the method of example 4, except that the hydrogenation is carried out at atmospheric pressure.

Ac-(D, L)Phe-(D, L)borough-C6H12benzosulfimide (0,243 g, 0,433 mmol) is subjected to interaction with cyanamide (0.20 g, 0,476 mmol) at 100aboutWith in 2 ml of absolute ethanol over night. characterized by positive staining of the strip when using dye Sakaguchi if borokhovich peptides Rf0-0,55, and discrete spot, Rf0,68 corresponding neproreagirovavshimi source material. The resulting product (81 mg) re-treated with 2 ml of a solution of cyanamide (10 mg/ml) during the night by the above method and after powdering simple ether obtain 71 mg of the target product.

Mass spectroscopy (FAB) for C22H37N5O4B:

Calculated +H: 446,30.

Found +N: 446,23 and 404,19 (corresponding neproreagirovavshimi branimirova the peptide) .

It should be borne in mind that the method of example 5, is the best way of obtaining borokhovich peptides and differs in the fact that they use a greater excess of cyanamide and a longer time of interaction.

P R I m e R 11. Boc-(D)Phe-broad- -C10H16-bansilalpet.

Boc-(D)Phe-Phe-OH get the procedure described to obtain the dipeptide Boc-(D)Phe-Pro-OH in example 2. After hydrogenation complex benzyl ester, the product vykristallizovyvalas from a mixture of chloroform:hexane to yield the desired peptide (I. pl. 133-133,5aboutC).

Elemental analysis for C23H28N2O5, %:

Calculated: C 66,96; H 6,86; N 6,79.

Found: C 66,75; H 6,79; -OH (6,00 g, 14.5 mmol) with NH2-CH[(CH2)3Br]BO2-C10H16HCl (example 1A, 5,33 g, 14.5 mmol) by the procedure described in example 2, except that not perform chromatography on a column of LH-20. The resulting product vykristallizovyvalas from ethyl acetate to yield in the first chase 2,47 g of the product (so pl. 132-134aboutAnd of 5.05 g of the product (so pl. 133-135about(C) in the second race. When of TLC in the system methanol:water (85:15) showed a single spot, Rf0,29.

Elemental analysis for C37H51N3O6BrB, %:

Calculated: C 61,32; H 7,11; N 5,80; Br 11,03.

Found: C 61,21; H 7,02; N 5,59; Br 10,22.

Boc-(D)Phe-Phe-NH-CH[(CH2)3N3]BO2-C10H16get in the processing of the corresponding allylbromide (7,15 g, 9,87 mmol) sodium azide according to the method described in example 3, except that purification is not necessary to perform chromatographic separation on a column of LH-20. Elute the desired product from a solvent mixture of ethyl acetate:hexane in the form of gel and after separation and washing with hexane obtain 3.0 g of the desired product in the first chase and 2.9 g of the second portion. Bansilalpet Boc-(D)Phe-Phe-borough-C10H16receive from the specified azide (lower than the 5.37 g, 7.82 mmol) by the procedure described is e ninhydrin, Rf0.42 and weak spot in the UV region, 0,92 (the formation of spots in the UV region at Rf0,92 typical amines or guanidino compounds which are salts of benzosulfimide).

Mass spectroscopy (FAB) for C37H53N4O6B:

Calculated +H: 661,76.

Found, +H: 661,14.

Boc-(D)Phe-Phe-broad-C10H16receive the procedure described in example 5. Braintoy peptide (4.83 g, 5,90 mmol) is treated with a solution of cyanamide (50 mg/ml) in 20 ml of absolute ethanol for 7 days. From the reaction mixture allocate a portion of the product corresponding to 1.0 g of starting material, and heated separately without reverse refrigerator over night with complete metamorphosis amine in guanidino connection. After chromatography on a column of LH-20 and powdering the product simple ether obtain 0.52 g of the desired Bonilla.

Elemental analysis for C44H61N6O9SB, %:

Calculated: C 61,38; H 7,16; N 9,76; B 1,25.

Found: C 59,69; H 7,41; N 9,82; B 1,26.

Mass spectroscopy (FAB) for C38H55N6O6B.

Calculated +H: 703,43.

Found +N: 703,49.

P R I m e R 12. H-(D)Phe-Phe-broad- -C10H162HCl.

Pentolair 6, except that the sample is put on ionoobmennoe column in 20% ethanol followed by washing of the specified column 20% solution of ethanol. The product is obtained (0,424 g) as a white solid.

Mass spectroscopy (FAB) for C33H47N6O4B:

Calculated +H: 603,38.

Found +N: 603,41.

P R I m e p 13. Bansilalpet Ac-Ala-Lys(Boc)-broad-C10H16.

Ac-Ala-Lys(Boc)-OH is obtained by conjugate reactions of N-oxenfree acid amidoamine Ac-Ala-OH (obtained by the method of Anderson) c H-Lys(Boc)-OH. N-oxysuccinimide Ac-Ala-OH (6.25 g, a 27.4 mmol) is dissolved in 30 ml of dioxane and added to a solution of H-Lys(Boc)-OH (7.50 g, 30.4 mmol), dissolved in a mixture consisting of 30 ml of 1.0 N. NaOH and triethylamine (2,12 ml, 15.0 mmol). The reaction mixture was stirred overnight and then acidified with hydrochloric acid. To obtain the required saturability of the solution add the desired amount of dry sodium chloride. The product is extracted in ethyl acetate and then washed with 0.2 N. HCl, obtained in a saturated aqueous solution of sodium chloride. The solvent is removed by evaporation. After crystallization from a mixture of ethyl acetate:hexane gain of 7.3 g of the desired product, (so pl. 86-89aboutC).

wow, that the resulting product was then purified fractionated by crystallization from ethyl acetate. In the product (1.13 g) obtained in the second and third fractions, there is a single spot when of TLC in a solvent mixture of methanol: water (85: 15) when the value of Rf0,51. Thin-layer plate is exposed to vapors of hydrochloric acid, which after addition of ninhydrin indicator detect target Amin.

Ac-Ala-Lys(Boc)-NH-CH[(CH2)3N3] BO2-C10H16receive from the corresponding allylbromide (1,95 g, 2,90 mmol) using the procedure described in example 3, except that the purification of the product is carried out by crystallization from ethyl acetate, but not by chromatography on LH-20. After crystallization of the crude product (1.60 g) obtain 0.55 g of the pure product (so pl. 79-84aboutC) and 0.96 g of sediment. Below is the chemical composition of the crystalline product.

Elemental analysis for C30H52N7O7B, %:

Calculated: C 56,86; H 8,29; N 15,48; B 1,71.

Found: C 56,76; H Compared To 8.26; N 15,89; B 1,65.

Bansilalpet Ac-Ala-Lys(Boc)-borough-C10H16receive from the corresponding alkylated (0,433 g, 0,683 mmol) by the procedure described in example 4. After removal of christophechristophe (FAB) for C30H54N5O7B:

Calculated +H: 608,42.

Found 608,49.

Bansilalpet Ac-Ala-Lys(Boc)-broad-C10H16get with the interaction of Broomfield peptide with cyanamide by the procedure described in example 5. Chromatographic fractions containing the desired product, proscout simple ether with a yield of 0.83 g of the product as a white solid.

Elemental analysis for C37H62N7O10BS, %:

Calculated: C 55,00; H Of 7.75; N 12,14; B 1,34.

Found: C 54,09; H 7,53; N 12,22; B 1,34.

P R I m e R 14. Ac-Ala-Lys-broad-C10H162HCl.

Bansilalpet Ac-Ala-Lys(Boc)-broad-C10H16(0,200 g, 0,248 mmol) will unlock according to the method of example 6. After ionoobmennoi chromatography, evaporation of the solvent, drying in vacuum and powdering simple ether obtain 0.14 g of the desired product.

Mass spectroscopy (FAB) for C26H48N7O5B:

Calculated +H: 550,39.

Found +N: 550,42.

P R I m e R 15. Bansilalpet Boc- -Leu-Gly-Leu-Ala-broad-C10H16.

Boc-Leu-Ala-OBzL get on the methods of synthesis of dipeptides described in example 2. Boc-Leu-Ala-OBzL (23.7 g, 57,7 mmol) dissolved in 40 the I and after processing of the resulting sludge simple ether get triptorelin H-Leu-Ala-OBzL in the form of a crystalline product (22,8 g).

Elemental analysis for C18H25N2O5F3, %:

Calculated: C 53,19; H 6,21; N 6,89.

Found: C 53,37; H Of 5.68; N 6,84.

Boc-Gly-Leu-Ala-OBzL obtained by reaction of the coupling of the BOC-Gly-OH (5,70 g, a 32.6 mmol) of H-Leu-Ala-OBzL by the method of mixed anhydride described in example 2. Product (13.8 g) obtained as amorphous solids. Boc-Gly-Leu-Ala-OBzL will unlock triperoxonane acid according to the method described for the preparation of H-Leu-Ala-OBzL, except that trifenatate salt can be dissolved in a simple ether. The product is dissolved in ethyl acetate and purified anhydrous hydrogen chloride. After deposition of the obtained product by adding a simple ester gain of 7.7 g of the chloride of H-Gly-Leu-Ala--OBzL in the first collection.

Boc-Leu-Gly-Leu-Ala-OBzL get paired reaction Boc-Leu-OH (2,62 g, 10.5 mmol) of H-Gly-Leu-Ala-OBzL according to the method of obtaining the mixed anhydride described in example 2. After bicrystalline received product from a mixture of ethyl acetate:hexane get in the first collection 2.7 g of the named product (so pl. 95-96aboutC).

Elemental analysis for C29H46N4O7, %:

Calculated: C 61,89; H Compared To 8.26; N 9,96.

Found: C 62,00; H 8,40; N 9,83.

Boc-Leu-Gly-Leu-Ala-OH get the catalytic kidogo product. After bicrystalline of the obtained product from the hot solution of ethyl acetate to obtain 1.4 g of the substance.

Elemental analysis for C22H40N4O7, %:

Calculated: 55,90: H 8,55; N Up 11,86.

Found: C To 55.42; H Of 8.47; N 11,73.

Boc-Leu-Gly-Leu-Ala-NH-CH[(CH2)3Br] BO2- C10H16obtained by reaction of the coupling of Boc--Leu-Gly-Leu-Ala-OH (1.40 g, 2,96 mmol) of the amine from example 1A according to the method described in example 2, except that not carry out chromatographic separation. After crystallization from a mixture of ethyl acetate: hexane get 1,17 g of the desired product. When TLC in the solvent system methanol:chloroform (1:9) showed a single spot when the value of Rf0,68.

Elemental analysis for C36H63N5O8BrB, %:

Calculated: C 55,10; H 8,11; N 8,93; B 1,38.

Found: C 55,96; 8,30 H; N, A Total Of 8.74; (B 1,33.

The corresponding azide receive according to the method of example 3 with 97% yield, which in turn Boc-Leu-Gly-Leu-Ala-borough-C10H16according to the method in example 4. Analiticheskoy samples prepared by deposition of the specified product simple ether and chromatographic separation on his column of LH-20, followed by elution of a mixture of chloroform with g is SS="ptx2">

Found, +H: 721,55.

Bansilalpet Boc-Leu-Gly-Leu-Ala- -broad-C10H16receive the procedure described in example 5. Appropriate braintoy peptide (0,695 g, 0,791 mmol) is subjected to interaction with 5 ml of a solution of cyanamide (50 mg/ml) in absolute ethanol. After chromatography was carried out the above mixture and powdering simple ether gain of 0.41 g of the desired product.

Mass spectroscopy (FAB) for C37H67N6O8B:

Calculated: +H: 763,53.

Found: 763,8.

P R I m e R 16. Bansilalpet H-Leu--Gly-Leu-Ala-broad-C10H16HCl.

Bansilalpet Boc-Leu-Gly-Leu-Ala- -broad-C10H16(obtained in example 15, 0,050 g, 0,0543 mmol) is subjected to interaction with 2 ml of 4N of hydrogen chloride in dioxane for 5 min at room temperature. The solvent and excess HCl is removed by evaporation. The resulting sample is dried over potassium hydroxide in vacuo over night, and then after powdering simple ether get the desired product (46 mg) in the form of a mixed salt.

Mass spectroscopy (FAB) for C32H59N8O6B:

Calculated +H: 663,47.

Found +N: 663,50.

P R I m e R 17. Bansilalpet Bz-Glu--(OBu)-Gly-boroArg-C10Oia Bz-Glu(OBu)--Gly-OH c specified amine according to the method described in example 2. The corresponding azide receive in accordance with the method described in example 3, and braintoy peptide obtained by the method of example 4.

Mass spectroscopy (FAB) for C32H49N4O7B:

Calculated +H: 613,38.

Found +N: 613,60.

The final product is obtained according to the method described in example 5.

Mass spectroscopy (FAB) for C33H51N6O7B:

Calculated +H: 655,40.

Found: 655,37.

Elemental analysis for C39H5N6O10SB, %:

Calculated: C 57,62; H 7,08; N 10,34; B 1,33.

Found: C 57,43; H 7,25; N To 9.91; B 1,23.

P R I m e R 18. Bansilalpet Bz-Glu--Gly-broad-C10H16.

Bansilalpet Bz-Glu(OBu)-Gly-broad-C10H16(of 0.13 g, 0.16 mmol) dissolved in 5 ml of dioxane, are added to benzosulfimide (0.10 g, 0.66 mmol) and the reaction solution is stirred over night at room temperature. After concentration of the solution up to a volume of about 1 ml by evaporation and subsequent poroshkovaya simple ether get a solid (0.14 g). The resulting product chromatographic 2.5 x 50 cm column of LH-20 in methanol. The fractions containing the desired product, evaporated, and theH43N6O7B:

Calculated +H: 599,34.

Found, +H: 599,35; 613,36 (not identified).

P R I m e R 18a. Bansilalpet Bz-Glu--Gly-broad-C10H16.

Bansilalpet Bz-Glu(OBu)-Gly-broad-C10H16(example 17, 0.20 g, 0,246 mmol) is treated with anhydrous hydrogen chloride by the method described in example 6, within 45 minutes After powdering the specified product, the NMR data show that about 30% of the tert-butyl protective group is still in the specified product. Then the product is subjected to interaction with anhydrous triperoxonane acid for 45 min at room temperature. TFA is removed by evaporation and after powdering the resulting sludge simple ether obtain 143 mg of the named product.

Mass spectroscopy (FAB) for C29H43N6O7B:

Calculated +H: 599,34.

Found +N: 599,35.

P R I m e R 19. Bansilalpet Bz-Pro--Phe-broad-C10H16.

Bz-Pro-Phe-OH (so pl. 200-201aboutC) receiving by the method described in example 2, used for the synthesis of dipeptides.

Elemental analysis for C21H22N2O4, %:

Calculated: C 68,82; H The 6.06; N 7,65.

Found: C 68,91; H 6,09; N 7,47 c specified amine according to General method described in example 2, except that the chromatographic separation is not carried out. At TLC in a solvent mixture of methanol: chloroform (1:9) there is a vast spot at Rf0,72 and traces when Rf0,86.

Mass spectroscopy (FAB) for C35H45N3O5BBr:

Calculated +H: 678,27.

Found +N: 677,95.

The resulting alkylhalogenide converted into azide and braintoy peptide according to the methods described in examples 3 and 4.

Mass spectroscopy (FAB) for (Bz-Pro-Phe-borough-C10H16)-C35H47N4O5B:

Calculated +H: 615,37.

Found +N: 615,42.

Target bansilalpet Bz-Pro-Phe- -broad-C10H16receive the procedure described in example 5.

Mass spectroscopy (FAB) for C36H49N6O5B:

Calculated +H: 657,39.

Found +N: 657,13.

Elemental analysis for C42H55N6O8SB, %:

Calculated: C 61,90; H 6,82; N 10,31; B 1,33.

Found: C 60,16; H 7,27; N 9,79; B 1,44.

P R I m e R 20. Chloride Bz-Pro-Phe-broad-OH.

Bansilalpet Bz-Pro-Phe-broad- -C10H16(compound of example 19, 0.64 g, of 0.79 mmol) dissolved in 4 ml of methylene chloride and cooled on the effect of 1.0 N. trichloride boron (Aldrich Chemical Co., Milwankee, WI), 50% of dry methylene chloride and placed in a bath of dry ice. The solution was stirred at -78aboutC for 5 min, and then the flask is put in a bath with ice (0about(C) where the solution is stirred for 15 minutes In the reaction solution is slowly added (5 ml) of cold water, followed by diluting it to 120 ml of 20% acetic acid. The organic phase is separated and extracted product. The aqueous phase is applied to a column volume of 20 ml for SP-Cefalexin, balanced 20% acetic acid. The column was washed with about 150 ml of 20% acetic acid, followed by establishing a linear concentration gradient from 200 ml of 20% acetic acid to 200 ml of 20% acetic acid containing 0,30 N. HCl. Product elute at a concentration of HCl in the range of 0.08 to 0.15 N. After evaporation of the solvent, drying in vacuum and powdering simple ether get the desired product (0,19 g).

Mass spectroscopy (FAB) for C26H35N6O5B:

Calculated +H: 523,29.

Found +N: 579,34 (not identified).

Elemental analysis for C26H36N6O5ClB, %:

Calculated: C 53,29; H 6,55; N 14,34; B 1,84.

Found: C 53,27; H to 6.58; N 13,25; the individual product, properties in the NMR and MS are identical to the original complex ester of example 19.

Mass spectroscopy (FAB) for C36H49N6O5B:

Calculated +H: 657,40.

Found +N: 657,39.

P R I m e R 21. Hydrochloric salt Bz-Pro-Phe-broad-F

Bz-Pro-Phe-NH-CH[(CH2)3NH-C(NH)NH2]- BF2HCl

Free Bronevoy acid (compound of example 20, 0,100 g, 0,179 mmol) dissolved in 2 ml of water. To the resulting solution was added 0,040 ml of 48% hydrofluoric acid at room temperature. Almost immediately formed solidairty the precipitate. The reaction mixture was stirred for 10 min, then frozen and vacuum to remove the excess hydrofluoric acid and water. The residue is dissolved in methanol, concentrated and proscout simple ether. The output is 0,093 g of the desired product.

Mass spectroscopy (FAB) for C26H33N6O3BF2:

Calculated +H: 527,29.

Found 527,31 and additional masses, characteristic of free Bronevoy acid.

Elemental analysis for C26H34N6O3BF2ClHCl.

Boc-Ala-Phe-NH-CH[(CH2)4NH2]BO2- -C6H12benzanilide according to the method described in example 3, except that the purification of the obtained product does not require chromatography on a column of LH-20. The obtained azide hydronaut by the method of example 4, except that take 2 EQ. benzosulfimide, and hydrogenation is carried out in the course of 2 hours Get the final product at 40% output (so pl. 154-160aboutWith decomposition).

Mass spectroscopy (FAB) for C28H46N4O6B:

Calculated +H: 547,38.

Found +N: 547,43.

P R I m e R 23. Triftoratsetilatsetonom H-Ala-Phe-(D,L)Lys-C6H12.

Bansilalpet Boc-Ala-Phe-(D, L)Lys-C6H12subjected to interaction with triperoxonane acid for 1 h at room temperature. After evaporation of the solvent and powdering sediment simple ether get the desired product in the form of a solid substance.

Mass spectroscopy (FAB) for C23H39N4O4B:

Calculated +H: 447,31.

Found +N: 447,31.

Elemental analysis for C31H46N4O9SF3B 2H2O, %:

Calculated: C 49,34; H Of 6.68; N 7,42; B 1,43.

Found: C 49,26; H 5,94; N 7,12; B 1,34.

P R I m e R 24. Bansilalpet Boc - (D)Yal-Leu-Lys-C6H12.

/P> Mass spectroscopy (FAB) for C23H36N2O5:

Calculated +H: 421,27.

Found +N: 421,38.

After hydrogenation, get free boric acid with 100% yield in the form of a crystalline solid white.

Elemental analysis for C16H29N2O5, %:

Calculated: C 59,34; H 8,87; N 8,50.

Found: C 59,34; H 8,87; N 8,50.

Boc-(D)-Yal-Leu-OH is associated with an amine, obtained in example 1d, with the release of Boc-(D)Yal-Leu-NH-CH[(CH2)4Br]BO2-C6H1297%.

Mass spectroscopy (FAB) for C27H51N3O6BBr:

Calculated +H: 604,31.

Found +N: 604,31.

The resulting allylbromide converted into the corresponding azide in 85% yield by the procedure described in example 3, with its subsequent hydrogenation. The final product is obtained as a white solid in 62% yield.

Mass spectroscopy (FAB) for C27H53N4O6B:

Calculated +H: 541,41.

Found +N: 541,46.

Elemental analysis for C33H59N4O9SB1,5H2O, %:

Calculated: C 54,62; H 8,61; N 7,73; B 1,49.

Found: C 54,58; H 8,59; N 7,92; B 1,98.

P R I m e R 25. Bansilalpet Ac-Phe--b Ac-Phe-NH-CH[(CH2)4Br]BO2-C6H12obtained in 72% yield.

Mass spectroscopy (FAB) for C22H34N2O4BBr:

Calculated +H: 481,00.

Found +N: 481,21.

Azide obtained from 57% yield. The final product obtained when 50% of its output.

Mass spectroscopy (FAB) for C22H37N3O4B:

Calculated +H: 418,29.

Found +N: 418,31.

Elemental analysis for C29H42N3O7SBH2O, %:

Calculated: C 56,66; H 7,47; N 7,08; B 1,82.

Found: C A 56.88; H 7,43; N 7,22; B 1,53.

P R I m e R 26. Bz-(D,L)borolg-C6H12HBr.

Bz-(D, L)NH-CH[(CH2)3Br] BO2-C6H12receive by reacting amine (obtained in example 16, (5.0 g, 15.9 mmol) with an equivalent amount of sodium bicarbonate in a mixture consisting of 4 ml of dioxane and 4 ml of water at 0aboutC. After the initial mixing of the reactants, the reaction mixture was diluted with 6 ml of 50% dioxane with water and warmed to room temperature. The reaction mixture was stirred for 30 min at room temperature followed by its extraction in ethyl acetate and washing with water, 0,2 N. HCl, 5% aqueous solution of bicarbonate, Natalia, after filtration and evaporation receive a crystalline product. After separation and washing with ethyl acetate receive 3,26 g connection, so pl. 176-177aboutC.

Elemental analysis for C17H25NO3BrB, %:

Calculated: C 53,44; H 6,59; N To 3.67; B 2,83.

Found: C 54,50; H 6,76; N 3,68; B 2,84.

The resulting alkylhalogenide (1,00 g, 2,62 mmol) is transformed into the corresponding salt stirone. The desired product 0.84 g will be in the form of a solid white color.

Mass spectroscopy (FAB) for C18H28N3O3SB:

Calculated +H: 378,20.

Found +N: 378,21.

Elemental analysis for C18H29N3O3SBBr, %:

Calculated: C 47,18; H 6,38; N 9,17; B 2,36.

Found: C 46,11; H Of 6.71; N 8,97; B 2,22.

P R I m e R 27. Bansilalpet Bz(D,L)broad-C6H12.

Alkylhalogenide (obtained in example 26, 2.0 g, a 5.25 mmol) is transformed into azide (0.97 g, so pl. 138-139aboutC) according to the method described in example 3. The obtained azide turn bansilalpet Bz-borough-C6H12according to the method of example 4 in almost quantitative yield.

Mass spectroscopy (FAB) for C18H27N2O3B:

Calculated +H: 319,22.

Found +what amidon according to the method described in example 5, to yield 0.65 g of crystalline product, so pl. 242-244aboutC.

Mass spectroscopy (FAB) for C18H29N4O3B:

Calculated +H: 361,24.

Found +N: 361,24.

Elemental analysis for C24H35N4O6SB, %:

Calculated: C 55,59; H 6,82; N 10,81; B 2,08.

Found: C 54,60; H 6,70; N 11,24; B 1,87.

P R I m e R 28. Bansilalpet Ac-Leu--Thr(OBu)-broad-C10H16.

Ac-Leu-Thr(OBu)-OH is obtained by binding Ac-Leu-OSu c H-Thr(OBu)-OH according to the procedure described in example 13, for dipeptide synthesis, except that the final product is obtained in the form of a white amorphous solid after chromatography on a column of LH-20. Ac-Leu-Thr(OBu)-OH (3,29 grams for 9.90 mmol) is subjected to a paired interaction with the amine (example 1A) according to the method of obtaining the mixed anhydride described in example 2, except that there is no need to chromatography on a column of LH-20.

Ac-Leu-Thr(OBu)-NH-CH[(CH2)3Br] BO2- -C10H16obtained as an amorphous solid white with a yield of 5.39, Received alkylhalogenide converted into the corresponding azide with 82% yield by the method of example 3, except that for the subsequent described in example 4. Benzene - sulfonate AC-Leu-Thr-(OBu)-borough-C10H16get 74% yield, after chromatography on a column of LH-20 and powdering simple ether.

Mass spectroscopy (FAB) for C30H55N4O6B:

Calculated +H: 579,43.

Found +N: 579,48.

Braintoy peptide turn in a final product with a yield of 86% by the method of example 5.

Mass spectroscopy (FAB) for C31H57N6O6B:

Calculated +H: 621,45.

Found +N: 621,50.

Elemental analysis for C37H63N6SO9B, %:

Calculated: C 57,05; H 8,17; N 10,79; B 1,39.

Found: C, 56.47; H 8,01; N Of 10.93; B 1,34.

P R I m e R 29. Bansilalpet Ac-Leu--Thr-broad-C10H16.

Bansilalpet Ac-Leu-Thr-(OBu)-broad-C10H16(example 28, 0,200 g, 0,257 mmol) dissolved in a mixture consisting of 2 ml of methylene chloride and 2 ml of 4 N. HCl:dioxane, and then the resulting reaction mixture is stirred for 30 minutes at room temperature. The solvent is evaporated in vacuo and the resulting residue dried in high vacuum. The desired product is obtained in the form of a white solid with a 97% yield after powdering simple ether.

Mass spectroscopy (FAB) is e R 30. Bansilalpet Ac-Lys(Boc)-Pro-broad-C10H16.

Dipeptide Ac-Lys(Boc)-Pro-OH get the procedure described in example 13. After crystallization from ethyl acetate to obtain the specified product as a white solid (temp. 160-161,5aboutC). Specified dipeptide, Ac-Lys(Boc)-Pro-OH (3,15 g, 8,18 mmol) were coupled reaction with amine (example 1A) according to the method described in example 2. The resulting product (5.8 g) used without further purification. The compound obtained is transformed into azide according to the method of example 3 with the release of his 73% after chromatography on a column of LH-20. After hydrogenation by the method of example 4, chromatography on a column of LH-20 and powdering simple ether get bansilalpet Ac-Lys(Boc)-Pro-borough-C10H16with the release of 81%.

Mass spectroscopy (FAB) for C32H55N5O7B:

Calculated +H: 634,43.

Found +N: 634,46.

Braintoy peptide (2.0 g, 2,53 mmol) is subjected to interaction with cyanamide by the procedure of example 5 to yield 1.8 g of the desired product as a solid white color.

Mass spectroscopy (FAB) for C33H57N7O7B:

Calculated +H: 676,46.

Found +N: 676,41.

Elemental analysis d; B 1,22.

P R I m e R 31. Ac-Lys-Pro-broad-C10H162HCl.

Bansilalpet Ac-Lys(Boc)-Pro-broad-C10H16(example 30, 0,30 g, 0,360 mmol) is subjected to interaction with a mixture consisting of glacial acetic acid with 4 N. HCl and dioxane at a ratio of 50:50 (%) for 15 min at room temperature. The solvent is evaporated, and the residue is dried in vacuum. The precipitate is dissolved in water and passed through a 5 ml column on AGI-X8 (Cl--form). The resulting sample is evaporated and after powdering balance of simple ether get the desired product in the form of a solid white color (230 mg).

Mass spectroscopy (FAB) for C28H49N7O3B:

Calculated +H: 576,40.

Found +N: 576,45.

P R I m e R 32. Bansilalpet Ac-Ala--Glu(OBu)-broad-C10H16.

Ac-Ala-Glu(OBu)-OH is produced by the binding of Ac-Ala-OSu c H-Glu(OBu)-OH according to the method of example 13. Product vykristallizovyvalas from a mixture of ethyl acetate:hexane (so pl. is 147.5-148aboutC).

Elemental analysis for C14H24N2O6, %:

Calculated: C 53,14; H 7,66; N Cent To 8.85.

Found: C 53,28; H 7,53; N Remaining 9.08.

Ac-Ala-Glu(OBu)-NH-CH[(CH2)3Br]BO2- -C10H16get by the method of example 2, for Estadio reaction and not spend chromatography on a column of LH-20. After evaporation of the organic layer receive 87% yield of the desired product in the form of a partially crystalline solid. Allylbromide converted into azide according to the method described in example 3. Target product (so pl. 165-166aboutC) resulted in 50% yield after crystallization of the crude product from chloroform.

Elemental analysis for C28H47N6O7B, %:

Calculated: C 53,51; H Of 7.55; N 6,69; B 1,73.

Found: C 55,51; H 7,50; N 6,50; B 1,66.

Braintoy peptide receive according to the method of example 4 with 79% yield of the desired product.

Mass spectroscopy (FAB) for C28H49N4O7B:

Calculated +H: 565,38.

Found +N: 565,51.

The final product is obtained in the form of a solid amorphous substance of white color with the release of 70% by the method of example 5.

Mass spectroscopy for C29H51N6O7B:

Calculated +H: 607,40.

Found +N: 607,41.

Elemental analysis for C35H57N6O10BS, %:

Calculated: C 54,96; H 7,53; N 10,99; B 1,41.

Found: C 54,36; H 7,71; N 11,27; B 1,21.

P R I m e R 33. Bansilalpet Ac-Ala--Glu-broad-C10H16.

Bansilalpet Ac-Ala-Glu(Bu)-broad-C120 min miss chloride. Then the solution is stirred for 1.5 h at room temperature, and after evaporation of the solvent, get the oil. The desired product is obtained in the form of a white solid (82 mg) after drying in a vacuum and powdering simple ether.

Mass spectroscopy (FAB) for C25H43N6O7B:

Calculated +H: 551,34.

Found +N: 551,41.

Using similar methods described in examples 22 and 23 received in addition, the following compounds are presented in table. 1.

The biological tests of joints produced by the described method.

It is shown that all these compounds can be classified as low-toxic substances.

The inhibition of plasma kallikrein human blood

Kallickrein human plasma obtained from protagen AG (AG) imported from Switzerland. Specific activity, as described by the supplier, is 15 units per mg. 1 unit defined as the amount of enzyme required to hydrolyze 1 µmol of substrate, H-(D)Pro-Phe-Arg-para-nitroanilide, (Kabi S2302), for 1 min at a substrate concentration of 0.50 mm at 25aboutWith in 50 mm potassium-phosphate buffer, pH 8.0.

The mother solution of the enzyme 1 poliatilenglikola 6000 (PEG). In the standard test, 10 μl of the basic solution of kallikrein was added to 990 μl of a solution containing 0.20 mm H-(D)Pro-Phe-Arg--para-nitroanilide S2302 in 0.10 mm nutrifaster buffer, pH 7.5, which includes 0.20 M sodium chloride and 0.1% PEG, 25aboutC. Effect of inhibitors appreciate when registering enzymatic activity as determined by measuring the increase in absorption at 405 nm over time in the presence and absence of inhibitors. In table. 2 shows the levels of inhibitors and activity measured in the interval from 10 to 20 min after stimulation activity. The control activity of the inhibitors is 0,0092 + 0,0095 min-1.

In table. 2 shows the biological activity of inhibitors of plasma kallikrein human blood.

The table shows that these inhibitors have values of Ki< 0,7 µm. The most effective synthetic optimizastion inhibitor is 6-amidino-2-(4-amidinophenoxy)-benzo/ /thiophene having a value of Ki= 0,7 ám.

Thus, the above table. 2 compounds are more active inhibitors of kallikrein than the well-known derivative of benzo/ /thiophene.

In addition to participating in blood coagulation, kallik the displays chemotactic effect on leukocytes and causes pain. The first two types of activity are associated with inflammation. Therefore, it is assumed that the inhibition of kallikrein has anti-inflammatory and analgesic effect.

Compounds according to the invention were tested for inhibition of thrombin.

Inhibition of thrombin (Esterna activity).

The human thrombin (specific activity 2345 N1H u/mg is obtained from R. Q. P. Laboratories, South Bend, IN /HT Lot 102/. The mother solution of thrombin is prepared in 0.01 M PIPES buffer, pH 6.0, containing 0.75 M sodium chloride. Analysis of thrombin is carried out in sodium phosphate buffer, pH 7.5, containing 0.20 M of nitriloside and 0.1% PEG-6000. The initial substrate concentration is 0.10 mm, and the concentration of thrombin is 1.0 nm per mass. In table. 3 shows the levels of inhibitors and activity, measured in the interval from 10 to 20 min after stimulation reaction. The activity of thrombin for the control group is 0,0076-0,0005 min-1.

The results are shown in table. 3.

Biological data presented in the table. 3, confirm the superiority of the compounds in accordance with the described invention over the known compound N -(2-naphthyl-sulfanilyl)-4-amidinotransferase. Although the data in the table. 3 paulenich in table. 3. The value of Kiis defined as the concentration of inhibitor required for 50% inhibition of the enzyme in the absence of substrate. The presence of the substrate, for example thrombin, has a protective effect on the enzyme. From the data presented in table. 3, it follows that all tested compounds have a value of Kibelow 6 nm.

For the following connection values of Kiwere determined by the method of Leinweber and Brooke after establishment of the equilibrium of the enzyme and inhibitor at pH 7.5 (see table. 4).

These compounds, including Ac-Ala-Glu-boroArg-C10H16which has a value of Ki(final concentration) of 10 nm, i.e., a value which is approximately equal to Ki6, superior to comparative connection is still for the reason that they can be entered without the use of organic solvent - solubilizer.

In the case of peptides derived Bronevoy acid, the solubility of which is 1-50 mg/ml, the organic solvent solubilizer, is not required.

Inhibition of blood clotting, are shown at the definition of ART and RT.

Effect of protease inhibitors on blood coagulation in vitro is determined by measuring their actions in two different clinical settings is specified analyses supplies GeneraI Piagnostics, Jessup MD. Royal solutions of inhibitors are prepared in 25 mm HEPES-buffer, pH 7.5, containing 0.10 M sodium chloride. For ART analysis, the inhibitor solution (0.100 ml) incubated normal human blood plasma (0.100 ml) and spontaneous reagent ART (0.100 ml). After incubation for 5,0 min at a temperature of 37aboutWith the specified solution was added calcium chloride (0.100 ml) and the coagulation time measured in seconds, determine the fiberscope. In table. 5 shows the effect of different concentrations of inhibitor at the time of blood coagulation compared to the clotting time of a control group, established in the absence of inhibitor.

For RT analysis, solutions of inhibitor (0.100 ml) incubated usually plasma of human blood (0.100 ml) for 2 min at a temperature of 37aboutC. Then incubated solution was added simplistically reagent (0,200 ml) and determine the coagulation time of blood, are given in table. 6.

In table. 8 shows the summary results presented in table. 5 and 6, showing the estimated concentration of inhibitor necessary to increase the time partially activated thromboplastin (hartt) and prothrombin time (HRT) twice.

Ingebor protease Ac-(D)- Phe-Pro-broad-OH on the coagulation of blood in vitro is determined by measuring its effect on thrombin time (TT). A mixture consisting of 0.2 ml normal rabbit plasma and 0.05 ml of the buffer containing the inhibitor 6-fold amount of the desired final concentration, heated to 37aboutC. the Clotting of blood is stimulated by injection of thrombin (0.05 ml 6 times the number of final concentration). Used thrombin purchased from Sigma Chemical Comp. (NT-6634, activity 1190:11H IU/mg protein) and is prepared in the buffer. Used for both inhibitor and thrombin, the buffer is 0.1 M Tris-buffer (12,10 g/l) containing 0,154 M NaCl (8,84 g/l) and 2.5 mg/ml bovine serum albumin, pH 7.4. Thrombin time, measured in seconds, determine the fiberscope. In table. 9 presents the data of the influence of inhibitor on blood clotting time compared with the coagulation time of the control group without the use of inhibitors. Impact measures represent an average value of at least three measurements. If the reaction of the coagulation does not occur within 300 C, then it ceased.

The stability of the inhibitors in human plasma determined when ART

The stability of inhibitors in plasma assessed on their ability to inhibit blood clotting. First of all, the basic solutions of (1.0 μm) inhibitors, podliasie plasma human blood. The mixture is prepared at 0aboutWith, then aliquots (0,200 ml) is taken and incubated for 2 min at 37aboutC. Add the same amount of spontaneously prepared reagent ART and measuring the clotting time of blood, as described. The final concentration of the inhibitor during the analysis of blood coagulation is 250 nm. Incubation time (given in hours) and clotting time (measured in seconds) for each inhibitor are given in table. 10. Stability parameters for compounds E and F are determined simultaneously with the control group. Indicators for connections And received the next day.

The stability of the inhibitors in buffer solution

Inhibitors, each at a concentration of 1.0 μm, incubated at room temperature in a 0.20 M nutrifaster buffer, pH 7.5, containing 0.20 M sodium chloride and 0; 10% PEG. Then select aliquots (4,0 mm) and analyzed by thrombin sample, as described in the examples 72-110. Table 8 presents data thrombin activity (%) remaining after incubation and duration of stay in nutrifaster buffer of the tested inhibitors. In the case of the inhibitor And there is little loss of inhibitory activity. The inhibitor loses its biologization in vivo.

Female rats (Sprague Dawley CD Rats, weighing 130-140 g, provided by Charles River Labs, Inc., Wilmington, MA) anaesthetize pentobarbital sodium (50 mg/kg, intraperitoneally). On the ventral surface of the neck perform median incision and into one of the carotid arteries injected polyethylene catheter with removing it from the back of the neck. After removing the rats from narcosis, control blood samples taken of the carotid artery catheter, produce anticoagulation with sodium citrate and centrifuged (2000 x rpm, 10 min). The plasma is transferred into plastic tubes and kept on ice until analysis. Thrombin time is determined in the fiberscope.

Rats get proteiny inhibitor Ac-(D)Phe-Pro-broad-OH in the media, or the media, via a stomach tube in a volume of less than 4 ml of media using 5% dimethylsulfoxide in saline. Blood samples taken at different times after oral administration, which analyzed as described above. In the following table. 12 shows the data obtained clotting time of blood in seconds. When the clotting time exceeds 300, the table is specified as >300. Other data show the average time required for coagulation, measured ErrorLog receive inhibitor.

To further demonstrate the ability of this compound to inhibit blood clotting in vivo, rats anaesthetize pentobarbital sodium (50 mg/kg, intraperitoneally), and then in the jugular vein insert the catheter and the hole closed. After excretion in rats of orally administered anesthesia or 5 mg/kg of protease inhibitor, Ac-(D)Phe-Pro-broad-OH dissolved in water or an equal volume of water. After 30-60 min, all rats injected with 500 IU/kg of thrombin for 1 min All 14 rats receiving only water died within 10 min after infusion of thrombin. In contrast, only 8 out of 17 rats that received inhibitorsdisease water died within 10 min, and the remaining lived 1 hour, and at this time they were euthanized.

Inhibition of in vivo coagulation of blood after oral, topical and rectal administration.

General method:

Male Lewis rats weighing 300-350 g anaesthetize pentobarbital sodium (50 mg/kg, intraperitoneally) and jugular vein insert slastikov cannula attached to a polyethylene tube. The specified receiver output at the back of the neck and attach to the syringe through a shut-off valve. Blood samples (0.5 ml) fill the syringe, washed with citrate buffer prior to each blood sampling before it is placed in a sealed container containing citrate buffer. In addition, after each blood sampling, the cannula is washed with saline solution. Then blood samples are centrifuged (2500 rpm for 15 min) and for the measurement of coagulation time of use 0.2 ml plasma samples. Time measurement of blood coagulation is performed on the fiberscope, as described below. First of all, plasma (0.2 ml) is placed in fibroscan and add Tris-buffer (50 μl), pH 7.4. The buffer solution containing the plasma, incubated at 37aboutC for 1 min, then add 50 ál of 24 µg/ml of thrombin solution in Tris buffer, and measuring the coagulation time in seconds. If the coagulation time exceeds 300, then indicate how > 300.

Oral administration:

Rats, introduced into the jugular vein catheter, remove from narcosis to oral administration of the inhibitor. An aqueous solution of the protease inhibitor Ac-(D)Phe - Pro-broad-OH containing 3 mg of inhibitor per kg of weight in rats (approximately 1 mg per rat) in 0.75 ml of water per 1 kg of weight of rats injected through the probe. In the following table. 13 shows the data obtained.

Local application:

In the abdominal cavity of rats with the introduction of the jugular vein catheter, perform incision 3 cm, while they are still in narcotization protease Ac-(D)Phe-Pro-broad-OH, containing 3 mg of inhibitor per 1 kg in rats (approximately 1 mg per rat) in 1 ml of water per 1 kg of body weight rats, injected into the ascending part of the cavity of the large intestine. The incision is closed using parentheses to suture. The following table summarizes the data obtained.

Rectal use:

As a technique for rectal administration of the inhibitor to rats with inserted into the jugular vein catheter using the methodology described KAMIJO in J. Pharm. Sci., 71: 621 (1982).

Made device consisting of 0.89 cm and 0.71 cm silicone gasket, United in length by 2 cm of the wire. The specified device is inserted into the rectal orifice of the rat, and the first strip of larger size and then glued to the anal passage of an appropriate adhesive. The introduction of the inhibitor is carried out by injection through located outside of the gasket. Dose for rectal administration is 3 mg of protease inhibitor Ac-(D)Phe-Pro-broad-OH 1 kg in rats (approximately 1 mg per rat) in 0.6 ml of water per 1 kg of body weight of the rat. In the following table. 15 shows the data obtained.

Inhibition of in vivo inflammatory process caused by Croton oil

Get two solutions: in the first moderator), and the second solution contains 5% Croton oil in acetone medium to which was added 10 mg/ml of the proposed connection (test solution). CROTONALDEHYDE solution (10 μl), or alternatively, the test solution was applied to the right ear of each animal (rat Sprague Dawley CD, weighing 130-140 g, provided by Charles River Labs, Inc., Wilmington, MA). The acetone carrier pure (acetone solution) (10 ml) is applied to the left ear of each animal. One hour after treatment of animals euthanized, their ears cut off, punched on drives larger 1/4 inch in diameter and weighed. Swelling is measured as the difference in mass between Croton solution, which treated the right ear and the acetone solution is left ear. The obtained data are compared with indomethacin, which is known as non-steroidal anti-inflammatory drugs, prepared and applied in the same way as the test compound. In table. 16 shows the averaged values for compounds F, Ac-Phe-broad- -C10H16. As used below, the term "dose" refers to the number of active anti-inflammatory ingredient in mcg (connection A, C, D, E, F or G, or indomethacin) in solution, applied to the right ear, and n indicates the number of rats, kaliterna activity of compounds a and C, taken under the same conditions (dose of 100 mcg).

Inhibition of plasmin morrisonville peptides.

The plasmin human plasma get from American Diagnostics, Greenwich. The concentration of active centers in the product is determined by a method similar to the method described by Coleman and other analysis of the enzymes that perform the spectrophotometer Perkin Elmer Lambda 4C using computational machine type Perkin-Elmer 7300. As buffer using 50 mmol Tris-buffer, pH 7,40 containing 110 mmol of sodium chloride, 0.1% PEG 6000 and 0.30 mmol H-(D)Val-Leu-Lys-p-nitroanilide. To this mixture was added the appropriate amount of inhibitor and then plasmin. At 25aboutTo measure the increase in absorption at 405 nm over time. Because the inhibitors bind slowly, the reaction rate is determined by the tangent held to the curve of the reaction after 10 min from the beginning of the reaction. In each case carry out a control test without the use of an inhibitor. In all cases the concentration of plasmin is less than 10% concentration of inhibitor.

The results are shown in table. 18.

The tests showed that the compounds according to the invention have low toxicity, have high ingibiruet is imeneniya solvent-solubilizer.

DERIVATIVES of BORON-containing PEPTIDES of General formula I

R1-Z-NH--Wu-HW

where R1- H, Ac, Bz, Boc, Boc - Leu;

R2- (CH2)n, NHC(NH)NH2;

n = 3,4;

y is the residue bingonova or piandiong ether;

HW = C6H5SO3H, HCl, HBr;

Z = D or L - Phe, DPhe - Pro, DPhe - Phe, Ala - Lys, Pro - Phe, Ala - Phe, Glu - Phe, Ala - Glu, Glu - Gly, Gly - Leu - Ala, pyro - Glu - Phe, Val - Val, (D)Val - Leu, Lys-Pro, Leu - Thr.

 

Same patents:

The invention relates to the field of Bioorganic chemistry, namely to new connections dibromide 6-(D-leucyl-L-prolyl-L-arginyl), aminonaphthalene-1-pentanitroaniline (1) and its benzyloxycarbonyl (Z) is the derivative (2)

The invention relates to medicine, namely to dermatology, and can be used in the treatment of psoriasis
The invention relates to medicine, in particular to rheumatology

The invention relates to medicine and can be used in the treatment of ischemic disorders of cerebral circulation, resulting in complete or partial blockage of an artery by a thrombus or embolus, or ischemia of the brain due to cerebral vascular insufficiency

The invention relates to the field of treatment for erectile dysfunction, in particular to rodstvennym gene calcitonin peptides, analogs or partial sequences and pharmacologically tolerable salts, intended for the treatment for erectile dysfunction

The invention relates to medicine, namely to pharmacology

The invention relates to veterinary medicine, in particular to the means for interrupting estrous females cats

The invention relates to the field of preparative biochemistry, namely the method of production of calmoduline - Ca2+- dependent low-molecular-weight protein that is involved in regulating the activity of several enzyme systems and processes in the cell, mainly phosphodiesterase of cyclic nucleotides
The invention relates to medicine, in particular to the treatment of infectious diseases
The invention relates to obstetrics and can be used in hospitals when you stop hypotonic uterine bleeding in childbirth, at the time of caesarean section and postpartum period

FIELD: pharmaceutical chemistry.

SUBSTANCE: invention relates to (i) essentially crystalline melagatran in the form of hydrate, which is characterized by x-ray diffraction pattern on powder having crystalline peaks with following d values: 21.1, 10.5, 7.6, 7,0, 6.7, 6.4, 6.2, 5.7, 5.4, 5.3, 5.22, 5,19, 5.07, 4.90, 4.75, 4,68, 4.35, 4.19, 4.00, 3.94, 3.85, 3.81, 3.73, 3.70, 3.63, 3.52, 3.39, 3.27, 3,23, 3.12, 3.09, 3.06, 2.75, 2.38, and 2.35 Å and/or water content 4.3%; and (ii) essentially crystalline melagatran in the form of anhydrate, which is characterized by x-ray diffraction pattern on powder having crystalline peaks with following d values: 17.8, 8.9, 8.1, 7.5, 6.9, 6.3, 5.9, 5.6, 5.5, 5.4, 5.3, 5.2, 5.0, 4.71, 4.43, 4.38, 4.33, 4.14, 4.12, 4.05, 3.91, 3.73, 3.61, 3.58, 3.56, 3.47, 3.40, 3.36, 3,28, 3.24, 3.17, 3.09, 3.01, 2.96, 2.83, 2.54, 2.49, 2.41, 2.38, and 2.35 Å. Invention also relates to a method for preparation of indicated form, a method for interconversion of anhydrite form, to use of indicated compounds as pharmaceutical agent, and to preparation of drugs. Pharmaceutical preparation is suitable for treatment of condition, in case of which inhibition of thrombin is needed or desirable. Invention provides a method for treatment of such condition.

EFFECT: increased chemical stability and solid state stability as compared to amorphous forms of melagatran.

14 cl, 4 dwg, 3 tbl, 9 ex

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention relates to compounds of the formula (I):

wherein r = 1, 2 or 3; s = 0; t = 0; R1 is taken among group including R11-CO and R12-SO2- wherein R11 is taken among group including (C6-C14)-aryl, (C1-C8)-alkyloxy-group wherein all given group are unsubstituted or substituted with a single or some similar or different substitutes R40; R12 means (C6-C14)-aryl wherein indicated group is unsubstituted or substituted with a single or some similar or different substituted R40; R2 means R21(R22)CH-, R23-Het-(CH2)k-, R23(R24)N-(CH2)m-D-(CH2)n- or R25(R26)N-CO-(CH2)p-D-(CH2)q- wherein D means bivalent residue -C(R31)(R32)-, bivalent (C6-C14)-arylene residue or bivalent residue obtained from aromatic group Het comprising 5 or 6 atoms in cycle among them 1 or 2 are similar or different cyclic heteroatoms taken among group including nitrogen and sulfur atoms; numbers k, m, n, p and q = 0, 1, 2; R21 and R22 that are independent of one another can be similar or different and taken among group including hydrogen atom, (C1-C12)-alkyl, (C6-C14)-aryl and so on; R23 means hydrogen atom, R27-SO2- or R28-CO-; R24, R25 and R26 mean hydrogen atom; R27 is taken among group including (C1-C8)-alkyl, (C6-C14)-aryl and so on; R28 is taken among group including R27, (C1-C8)-alkyloxy-group; R31 and R32 mean hydrogen atom; R40 is taken among group including halogen atom, hydroxy-, (C1-C8)-alkyloxy-group, (C1-C8)-alkyl, (C6-C14)-aryl and so on; R91, R92, R93 and R96 means hydrogen atom; R95 means amidino-group; R97 means R99-(C1-C8)-alkyl; R99 is taken among group including hydroxycarbonyl- and (C1-C8)-alkyloxycarbonyl-; Het means saturated, partially unsaturated or aromatic monocyclic structure comprising from 3 to 6 atoms in cycle among them 1 or 2 are similar or different heteroatoms taken among group comprising nitrogen and sulfur atoms; in all its stereoisomeric forms and also their mixtures in any ratios, and its physiologically acceptable salts. Invention proposes a method for preparing compound of the formula (I). Also, invention proposes a pharmaceutical preparation eliciting inhibitory activity with respect to factor VIIA and containing at least one compound of the formula (I) and/or its physiologically acceptable salts and pharmaceutically acceptable carrier. Invention provides preparing compounds of the formula (I) eliciting power anti-thrombosis effect and useful for treatment and prophylaxis of thrombosis-embolic diseases.

EFFECT: valuable medicinal properties of compounds and composition.

10 cl, 70 ex

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention relates to compounds of the formula (I):

wherein r = 1, 2 or 3; s = 0; t = 0; R1 is taken among group including R11-CO and R12-SO2- wherein R11 is taken among group including (C6-C14)-aryl, (C1-C8)-alkyloxy-group wherein all given group are unsubstituted or substituted with a single or some similar or different substitutes R40; R12 means (C6-C14)-aryl wherein indicated group is unsubstituted or substituted with a single or some similar or different substituted R40; R2 means R21(R22)CH-, R23-Het-(CH2)k-, R23(R24)N-(CH2)m-D-(CH2)n- or R25(R26)N-CO-(CH2)p-D-(CH2)q- wherein D means bivalent residue -C(R31)(R32)-, bivalent (C6-C14)-arylene residue or bivalent residue obtained from aromatic group Het comprising 5 or 6 atoms in cycle among them 1 or 2 are similar or different cyclic heteroatoms taken among group including nitrogen and sulfur atoms; numbers k, m, n, p and q = 0, 1, 2; R21 and R22 that are independent of one another can be similar or different and taken among group including hydrogen atom, (C1-C12)-alkyl, (C6-C14)-aryl and so on; R23 means hydrogen atom, R27-SO2- or R28-CO-; R24, R25 and R26 mean hydrogen atom; R27 is taken among group including (C1-C8)-alkyl, (C6-C14)-aryl and so on; R28 is taken among group including R27, (C1-C8)-alkyloxy-group; R31 and R32 mean hydrogen atom; R40 is taken among group including halogen atom, hydroxy-, (C1-C8)-alkyloxy-group, (C1-C8)-alkyl, (C6-C14)-aryl and so on; R91, R92, R93 and R96 means hydrogen atom; R95 means amidino-group; R97 means R99-(C1-C8)-alkyl; R99 is taken among group including hydroxycarbonyl- and (C1-C8)-alkyloxycarbonyl-; Het means saturated, partially unsaturated or aromatic monocyclic structure comprising from 3 to 6 atoms in cycle among them 1 or 2 are similar or different heteroatoms taken among group comprising nitrogen and sulfur atoms; in all its stereoisomeric forms and also their mixtures in any ratios, and its physiologically acceptable salts. Invention proposes a method for preparing compound of the formula (I). Also, invention proposes a pharmaceutical preparation eliciting inhibitory activity with respect to factor VIIA and containing at least one compound of the formula (I) and/or its physiologically acceptable salts and pharmaceutically acceptable carrier. Invention provides preparing compounds of the formula (I) eliciting power anti-thrombosis effect and useful for treatment and prophylaxis of thrombosis-embolic diseases.

EFFECT: valuable medicinal properties of compounds and composition.

10 cl, 70 ex

FIELD: pharmaceutical chemistry.

SUBSTANCE: invention relates to method for production of acetylamidiniophenylalanylcyclohexylglycilpypidinioalanin amides of formula I , wherein X anions are physiologically acceptable anions, and analogous thereof. Said compounds are effective inhibitors of fibrillation factor Xa and are useful, for example, in prevention of thrombosis. Claimed method includes coupling of 2-[2-acetylamino-3-(4-amidinophenyl)-propionylamino]-2-cyclohexylacetic acid, obtained from 2-[2-acetylamino-3-(4-cyanophenyl)acryloylamino]-2-cyclohexylacetic acid by assimetric hydration and converting of cyano group to amidine, or salt thereof with 3-(2-amino-2-carbamoylethyl)-1-methylpyridinic acid or salt thereof. Also are disclosed starting materials and intermediated used in this method, process for production the same and acetyl-(S)-4-amidiniophenylalanyl-(S)- cyclohexylglycil-(S)-(1-methyl-3-pypidinio)alanin amide in form of ditosylate.

EFFECT: simplified method; increased commercial availability of compounds with applicable anion.

14 cl, 16 ex

FIELD: organic chemistry, medicine, pharmacology.

SUBSTANCE: invention relates to new inhibitors of thrombin of the formula (I)

,

method for their preparing, intermediate compounds used for their preparing of the formula (II)

and a pharmaceutical composition comprising compounds of the formula (I). Invention provides enhancing effectiveness in inhibition of thrombin.

EFFECT: improved preparing method, valuable medicinal properties of compounds.

23 cl, 61 ex

FIELD: chemistry of peptides, microbiology, biotechnology.

SUBSTANCE: L-alanyl-L-glutamine is prepared by incubation of a mixture containing a microorganism able to produce L-alanyl-L-glutamine from L-alanine ester and L-glutamine, L-alanine ester and L-glutamine, and isolation of the end product. Using the invention allows simplifying the process for preparing L-alanyl-L-glutamine. Invention can be used in pharmacy and food processing industry.

EFFECT: improved preparing method of dipeptide.

3 cl, 3 tbl, 1 ex

Peptide compounds // 2281955

FIELD: chemistry of peptides, medicine, pharmacy.

SUBSTANCE: invention relates to compound of the formula (I): wherein R1 represents benzofuranyl substituted with halogen atom or styryl substituted with halogen atom; R2 represents substituted hydroxyl substituted with mercapto-group or substituted sulfonyl, or its pharmaceutically acceptable salts. Compound of the formula (I) and its pharmaceutically acceptable salts possess the strong inhibitory effect on production of nitrogen oxide (NO) and can be useful in prophylaxis and/or treatment of NO-mediated diseases in humans and animals.

EFFECT: valuable medicinal and biochemical properties of compounds.

FIELD: pharmaceutical dosage forms.

SUBSTANCE: invention relates to compositions and methods for delivery of drugs. Composition according to invention represents powdered composite containing polymer, therapeutical agent, and complexing agent, said polymer containing one or more cyclodextrin fragments. Polymer interacts with complexing agent in a host/guest or guest/host manner resulting in formation of inclusion complex. Both polymer of composite and complexing agent may be used in a way to introduce functionality into therapeutical composition. Invention also discloses a method for preparing composition and to a method for delivering therapeutical agent.

EFFECT: widened choice of drug delivery methods.

26 cl, 31 dwg, 1 tbl, 65 ex

FIELD: organic chemistry, medicine, endocrinology, pharmacy.

SUBSTANCE: invention relates to novel compounds of the formula (I): or their pharmaceutically acceptable salts and composition containing thereof that possess activity with respect to release of growth hormone. In the formula (I) R1 represents hydrogen atom or (C1-C6)-alkyl; R2 represents hydrogen atom or (C1-C6)-alkyl; L represents compound of the formula: wherein R4 represents hydrogen atom or (C1-C6)-alkyl; p = 0 or 1; r = 1; q, s, t and u = 0, 1, 2, 3 or 4 being independently of one another; the sum q + r + s + t + u = 0, 1, 2, 3 or 4; R9, R10, R11 and R12 represent independently of one another hydrogen atom or (C1-C6)-alkyl; Q represents >N-R12 wherein R13 represents hydrogen atom or (C1-C6); or L represents compound of the formula: , wherein r = 0 or 1; q, s, t and u independently of one another = 0, 1, 2, 3 or 4; the sum q + r + s + t + u = 0, 1, 2, 3 or 4; R9, R10, R11 and R12 represent independently of one another hydrogen atom or (C1-C6)-alkyl; Q represents >N-R13 or compound of the formula: , wherein o = 0, 1 or 2; T represents -N(R15)(R16) or hydroxyl; R13, R15 and R16 represent independently of one another hydrogen atom or (C1-C6); R14 represents hydrogen atom; G represents compounds of the formula or wherein R17, R18, R19, R20 and R21 represent independently of one another hydrogen atom or aryl; J represents compounds of the formula or wherein R22, R23, R24, R25 and R26 represent independently of one another hydrogen or halogen atom; a = 0, 1 or 2; b = 0, 1 or 2; c = 0, 1 or 2; d = 0 or 1; e = 0, 1, 2 or 3; f = 0 or 1; R5 represents hydrogen atom; R6 and R7 represent independently of one another hydrogen atom or (C1-C6)-alkyl; or R6 and R7 can form -(CH2)i-U-(CH2)j- optionally wherein i and j = 1, 2 or 3 independently one another, and U represent a valence bond; R8 represents hydrogen atom or (C1-C6)-alkyl; M represents arylene or -CR27=CR28- wherein R27 and R28 represent independently one another hydrogen atom or (C1-C6)-alkyl. Also, invention relates to a pharmaceutical composition comprising compounds of the formula (I) or its pharmaceutically acceptable salt as an active component in common with a pharmaceutically acceptable carrier or excipient. Also, invention relates to a method for stimulation of growth hormone secretion from mammalian hypophysis that involves administration to mammal the effective dose of compound or its pharmaceutically acceptable salt. Invention provides synthesis of novel compounds and their composition containing thereof that can be used in treatment of diseases associated with the growth hormone deficiency.

EFFECT: improve stimulating method, valuable medicinal properties of compounds and pharmaceutical composition.

13 cl, 29 ex

Up!