Derived tripeptides as r - or rs-shape or their pharmaceutically acceptable non-toxic salts, pharmaceutical composition

 

(57) Abstract:

Usage: in medicine and biochemistry. The essence of the invention: derivatives of tripeptides in the form of R or RS form the General formula: A-Pro-Arg-aldehyde; A=RR, BC, R is phenyl, substituted or unsubstituted, thienyl, naphthyl, substituted or unsubstituted cyclohexyl, R1is hydrogen, lower alkyl, -lower alkyl, lower alkoxy, or N(R2)(R3), in which R2and R3independently:hydrogen or lower alkyl: or R2is hydrogen, and R3-acetyl or N-Vos (lower) alkyl; And-bicicletta group; 1, 2, 3, 4 - tetrahydroisoquinoline-1- (or 3-) - yl-carbonyl, D-perhydrosqualene-1-yl-carbonyl or their pharmaceutically acceptable non-toxic salts, which are able to inhibit thrombin. Reagent 1: AProOH. Reagent II:C - protected arginylation as trifenatate. The process is conducted in DMF at a temperature of 15 degrees. in the presence of N - methylmorpholine for 4 hours, then within about days slowly warmed to room temperature, the resulting lactam restore methylalanine hydride. 2 C. and 12 C.p. f-ly,., table 4.

The present invention relates to inhibitors of thrombin, useful as anticoagulants for humans and animals. In particular, the image is matromony activity.

Inhibition of thrombin currently achieved by the introduction of heparins and coumarins. The mechanism of action of these means were subjected to enhanced learning. Heparin can be administered exclusively parenterale, and their dosage should be carefully monitored. The effect of coumarin-based blocking or inhibiting the formation of prothrombin, you need some time to achieve maximum efficiency.

Although as heparins and coumarins are effective anticoagulants, however, there is a need to antithrombin means quick action to prevent the formation of blood clots, and not impeding the action of plasmin to dissolve existing clots.

Inhibiting thrombin compounds provided by the present invention represented by the following formula 1:

(1)

where a is 1 group of the formula:

< / BR>
in which R is phenyl of the formula:

< / BR>
where a and a' independently represents hydrogen, lower alkyl, lower alkoxygroup, halogen, trifluoromethyl, hydroxyl; or

R represents thienyl; naphthyl, unsubstituted or mono - or disubstituted by lower alkoxygroup, cyclohexyl;

RRuppo formula:

-N(R2)(R3)

in which R2and R3independently represent hydrogen or lower alkyl, or R2is hydrogen, and R3-acetyl, N-Boc (lower) alkyl, provided that when R1is methyl or ethyl, then, is different from methyl or ethyl, or a represents a bicyclic group of the formula:

< / BR>
where Q is CH2-CH2- , if Q

Y represents one radical-CH2-;

R5represents hydrogen or tert-butoxycarbonyl;

R6represents hydrogen; and the dashed circle within 6-membered cycle bicyclic group indicates perhydro-loop;

and their pharmaceutically acceptable non-toxic salt.

The peptides of formula 1 are applicable as antithrombotic means may be used as AIDS in the treatment of tissue plasminogen activator (PLA), streptokinase or urokinase.

These compounds get the usual ways. For example, BOC-D-Phg connect with the ester of L-Proline with the formation of ester BOC-D-Phg-Pro. Ester group is removed and Boc-D-Phg-Prg connect with a lactam of the formula L-arginine with getting lactam Boc-D-Phg-Pro-Arp in aminosidine form. Lactam cycle arginine reveal the Chida turn in acceptable salt form, such as acetates and sulfates.

The invention also provides a method of preventing human and animal blood clots and pharmaceutical compositions applicable in this way.

The compounds of formula 1 are tripeptides, which represent the amino acid residue, such as phenylglycine (Phg), and when And distinct from amino acid residue, i.e., when is a group other than amino or alkylamino, in this case, the compounds are N-acyl derivative of the dipeptide-Proline-arginine-aldehyde (Pro-Arg-H). As shown by formula 1, the asymmetric center of the fragment: While(C=O) has the R - or RS-configuration, while fragments of Proline and arginine-aldehyde-L-configuration.

The terms used in the definition of formula 1, have the following meanings:

"Lower alkyl" refers to C1-C4the normal alkyl or ISO-structure, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, etc.

"The lowest lexigraphy" refers to C1-C4alkoxygroup, such as methoxy, ethoxy-, n-propoxy, isopropoxy, h-butoxy-, tert-butoxypropan etc.

"Halogen" refers to fluorine, chlorine, bromine or iodine.

"Mono - or di(lower alumino-, n-butylamino, h-propylamino group, etc.

The term "C1-C6-alkyl" refers to alkido normal or ISO-structure, such as C1-C4-alkyl, as defined above, and in addition: n-pentyl, isopentyl, n-hexyl, and isomeric hexylene group, etc., "C2-C6alkenyl refers to unsaturated groups such as vinyl, allyl, butenyl, isomeric penthesilia and hextile group, "C3-C7-cycloalkyl" refers to cyclic hydrocarbon with 3-7 carbon atoms in the cycle, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl.

According to the definition of formula 1, when a is the group (R)(R1(B)C-, then R can be phenyl, which may be mono - or Disaese. Examples of such fanilow include phenyl (a and a' N), 4-were 3-ethylphenyl, 4-methoxyphenyl, 3-methoxyphenyl, 3-ethoxyphenyl, 2-methoxyphenyl, 3-isopropoxyphenyl, 4-hydroxyphenyl, 4-chlorphenyl, 3-chlorophenyl, 2-forfinal, 3-forfinal, 3-bromophenyl, 4-forfinal, 3-triptoreline, 4-hydroxymethylene, 2-hydroxymethylene, 3-AMINOPHENYL, 4-AMINOPHENYL, 3-amino-4-chlorophenyl, 3,4-dichlorophenyl, 3-hydroxy-4-forfinal, 3-hydroxy-4-were, 3-methoxy-4-hydroxyphenyl, 3-chloro-4-ethoxyphenyl and similar is disubstituted naphthyl, include 1-naphthyl, 2-naphthyl, 6-methoxy-2-naphthyl, 8-hydroxy-1-naphthyl, 8-amino-2-naphthyl, 4-methyl-1-naphthyl, 6-chloro-2-naphthyl, 4-hydroxy-6-ethoxy-2-naphthyl, 8-methylamino-4-chloro-2-naphthyl, 6,8-dimethoxy-2-naphthyl, 6-ethyl-1-naphthyl, 4-hydroxy-1-naphthyl, 3-methoxy-1-naphthyl and similar raftiline group.

Examples of the group represented in formula 1, when the amino group of the formula: -N(R2)(R3) include: amino- (R2R3H), methylamino, ethylamino, isopropylamino-, dimethylamino group and the like, an amino group; and when R2hydrogen, and R3oxycarbonyl formula: R4-O-C(O)-, then examples of such groups include C1-C6-alkoxycarbonylmethyl, such as methoxycarbonylamino, ethoxycarbonyl-, tert-butoxycarbonylamino, soliloquymonologues, etc. C2-C6-alkenylcyclopropenes, such as vinyloxycarbonyl, allyloxycarbonyl-, 2-butyloxycarbonyl, etc. C3-C7-cycloalkylcarbonyl, such as cyclopropanecarbonyl, cyclopentanecarbonyl, cyclohexyloxycarbonyloxy and so forth Externalmemory represented by the symbol, in addition, include, for example: benthological replaced phenoxycarbonylamino, where substituted phenyl fragment adopts the above values, etc.

Examples of the groups(C=O) of formula 1, when a represents a radical of the group 1 of the formula: (R)(P1)(B)C-include: phenylglycyl, 3-methoxyphenethyl, 4-methoxyphenylazo, 4-chlorophenylglycine, 3,4-dichlorophenylethyl, 3-cryptomaterial, N-(tertbutyloxycarbonyl)phenylglycyl, N-(tert-butyloxycarbonyl-11-methyl)phenylglycyl-methylphenylacetic, a-ethylvanillin, a-methoxyphenylacetyl, a-isopropoxyaniline, 1-naphthylvinyl, 2-naphthylvinyl, N-(tert-butoxycarbonyl)-2-naphthylvinyl, 2-yenilgisi, 3-yenilgisi, N-(cyclopentanecarbonyl)-2-yenilgisi, 2-fuelcell, N-ethyl-2-furyl-glycyl, Mandalay, 4-chloromandelic, 3-methoxymandelic, a-hydroxy-a-(2-naphthyl)acetyl, a-hydroxy-a-(2-thienyl)acetyl, 1,4-cyclohexadienyl, 1-cyclohexylglycine, N-(tert-butyloxycarbonyl)-1,4-cyclohexadienyl, cyclohexylglycine and is similar to A(CO) group.

The peptide derivative represented by formula 1, where a is achiral 1-aminocyclopent or 1-aminocyclohexane reflected the following structural formula:

< / BR>
where R represents a carbon-carbon bond or-CH2-; R2and R3take above what Noel)-Pro-Arg-H; N-(1-methylenecycloartanol)-Pro-Arg-H, N-(1-tert-butyloxycarbonyl)-Pro-Arg-H, etc.

Examples of the peptides is represented by formula 1, where a bicyclic group of the above formula 2 include D 1,2,3,4-tetraethylene-1-ylcarbonyl - and D-1,2,3,4-tetrahydroisoquinoline-3-ylcarbonyl-11-acyl derivatives Pro-Arg-H, which are stated below:

(A)

(B)

dihydroindol-1-ylcarbonyl derivatives, which are stated below formula:

(C)

oxo-derivative represented by the formula:

(D)

(E)

and targetirovannye derivatives. The symbols R6and R5agree to the above values. R5preferably hydrogen, preferably hydrogen, methoxy group, ethoxypropan, chlorine or methyl.

Pharmaceutically acceptable salts of the peptides of the invention include salts with acids formed by adding inorganic and carboxylic acids. Examples of forming the inorganic salts include guloudajie acid, such as chlorotoluron acid and Hydrobromic acid, phosphoric acid and sulfuric acid. Salts with carboxylic acids formed with such acids as acetic, propionic, malonic, maleic, citric, succinic who eat for example, neutralization of the compound I in the form of the free base with an acid. Featured salts with acids include sulfates and hydrochloride.

Recommended embodiments of the invention include compounds of formula I, where And represent a group of the formula:

< / BR>
in which R represent phenyl:

< / BR>
or naphthyl, or substituted naphthyl, R1is hydrogen and the amino group of the formula: -N(R2)(R3). Additional recommended compounds represented by the variant, in which R2is hydrogen and R3-oxycarbonyl formula: R4O-C(O)-

Another featured embodiment of the invention includes the compounds of formula I, where a bicyclic group of the formula (2). Recommended connection of this embodiment is represented by formula I, where A(C=O) - 1,2,3,4,-tetrahydroisoquinoline-1-ylcarbonyl (formula 2, Q-CH2-CH2-, Y -, CH - and R5R6and 1,2,3,4-tetrahydroisoquinoline-3-ylcarbonyl (formula 2,

< / BR>
The compounds of formula 1 are known ways of connecting peptides. According to one such method acid A-COOH, where a takes the values specified for formula 1, link with protected carboxyl the Proline with the formation of the dipeptide (if And amino acid), or a complex ester of N-ACELP is when the product is removed and dipeptide in the form of the free acid lactam combined with a form of arginine. The above reaction sequence is illustrated by the following scheme:

< / BR>
where R represents aminosidine group.

Obtained by joining Arg(P)-lactam product (s) to restore sociallyengaged in an inert solvent with disclosure lactam cycle and the formation of Tripeptide in arginine-aldehyde form represented by the formula:

A(C=O)-PRo-Arg(P) H,

where Arg(P)-H is aminosidine arginine-aldehyde.

Lactam form of arginine receive intramolecular reaction aminosidine arginine (Arg-OH). For example. Boc-Arg(CbZ)O represented by the formula:

< / BR>
first converted into an active ester form, such as an active mixed anhydride in the reaction with CHLOROFORMATES, for example, from ethylchloride to isobutylphthalate. The formation of ester is carried out in the presence of a tertiary amine, such as 11-methyl-formalin. Adding a stronger tertiary amine such as triethylamine leads to intramolecular acylation with the formation of the lactam form diaminohexane arginine below formula:

< / BR>
Before use in the reaction of the compound with A(C=O)-Pro-OH according to the diagram above, protect the amino group.

Connection connection asoon to complex ether Proline, when A - amino acid residue, to carry out an initial protection of the amino group of amino acids. Use regular aminosidine group, often used for temporary protection or blocking of amino groups. Examples of such protective groups include alkoxy, alkenylacyl, cycloalkyl and aryloxypropanolamine groups, such as: etoxycarbonyl, tert-butyloxycarbonyl, cyclohexyloxycarbonyl, adamantanecarbonyl, trichlorocyanuric, benzyloxycarbonyl, diphenylmethylene and similar groups. Ester group, used to protect carboxyl Proline during the reaction of accession, may be any of the commonly used easily removed hard-ester groups, such as tert-butyl, benzyl, p-nitrobenzyl, p-methoxybenzyl, diphenylmethyl, trichlorethyl, pencil or trialkylsilyl forming ester. When carrying out the reactions proceed to use Proline ester group which can be removed under conditions which aminosidine group remains intact. Aminosidine group alleluya acid (AZOOR) the result remains in place protection of the amino group during subsequent connections)(R1(B)C and the amino group-N(R2(R3), where R2the hydrogen and R3lower alkyl, is obtained on the basis of the corresponding compounds in which the amino group, known methods of alkylation. For example, N-methyl-D-phenylglycyl-L-propyl-L-arginine-aldehyde receive rehabilitation alkylation as follows. CbZ-protected D-phenylglycine combined in DMF with tert-butyl ester of L-Proline in the presence of dicyclohexylcarbodiimide (DCC) and hydroxybenzotriazole (HoBT) with the formation of the dipeptide: tert-butyl ether CbZ-D-phenylglycyl-L-Proline. The resulting peptide hydronaut in ethanol over palladium on coal with the removal of the CbZ protective group, for product recovery add the formaldehyde and the hydrogenation continued with the formation of the tert-butyl ether N-methyl-D-phenylglycyl-L-Proline. N-methyl secondary amino group phenylglycine fragment protect CbZ group in the reaction of tert-butyl ester of the dipeptide with benzylchloride in THF containing N-methylmorpholine, and the formation of N-CbZ-N-methyl-D-phenylglycyl-L-Proline tert-butyl ester. Ester tert-boutelou group is removed at room temperature in triperoxonane acid containing anisole, and get N-CbZ-N-methyl-D-phenylglycyl-L is restoring education Arg-aldehyde according to the aforementioned method. Both CbZ protective group Tripeptide removed by hydrogenation over Pd-C as catalyst to obtain N-methyl-D-phenylglycyl-L-prolyl-L-arginine-aldehyde.

The compounds of formula 1, where a group (R)(R1)(B)C-, in which R - cyclohexadienyl or cyclohexenyl and alkylamino formula: -N(R2)(R3), can be synthesized by the recovery of the amine formed in the reaction with the lowest alkilany aldehyde, lamborghini.com sodium. Similar similar N-alkylation may be carried out using alkylated and sodium hydride.

The compounds of formula 1, where a bicyclic group of the formula 2, get the same above methods of joining. For example, the peptide of formula 1, where a is 1, 2,3,4-tetrahydroisoquinoline-1-yl, obtained by acylation of ester of Proline, such as benzyl ether active derivatives of 1,2,3,4-tetrahydro-1-carboxyethylidene. Active derivatives, which can be used include galodamadruga acid such as the acid chloride or bromohydrin, acid azide, active esters and anhydrides, such as those that form in the aforementioned manner in the reaction with chloroformate. During Alliluyeva accession afalse ester of Proline apply such an active ester, as the N-Boc-1,2,3,4-tetrahydro-1-carboxyethyl formed by reaction with isobutylacetate. In the received quality of the product peptide-N-BOC-1,2,3,4-tetrahydroisoquinoline-1-ylcarbonyl-Proline-layer ether to remove the ester group, the free acid is converted into an active ester and the latter is attached to the lactam form of arginine. Obtained as a product lactam then transform the manner in aldehyde form and get the compound of formula 1, namely BOC-1,2,3,4-tetrahydroisoquinoline-1-ylcarbonyl-Pro-Arg-h

Targetirovannoy bicyclic group represented by formula 2 is obtained by hydrogenation in the usual way either partially restored, or unsaturated acids. For example, 1,2,3,4-tetrahydroisoquinoline-1-carboxylic acid hydronaut over platinum oxide in a solvent such as ethanol or acetic acid, and receive perhydro(decahydro)isoquinoline-1-carboxylic acid. Then targetability use the above method for acylation of ester of Proline. Examples of such PermitRootLogin formula 1 include: N-(D-decahydroquinoline-1 oil)-L-propyl-L-orgininally and N-(D-decahydroquinoline-3 oil)-L-prolyl-L-arginine-aldehyde.

Videopioneer approximately 15oC. Reaction of the joining is carried out in an inert organic solvent such as dimethylformamide, tetrahydrofuran, methylene chloride, chloroform and similar conventional solvents. When used in the reactions of addition of the active ester alleluya acid this reaction is usually carried out in anhydrous conditions.

Compounds of the invention are best isolated in the form of salts with acids. Salts of compounds of formula 1, formed with acids of the type mentioned above are applicable as pharmaceutically acceptable salts for the introduction of protivotrombovyh funds and for the preparation of compounds of such funds. For isolation and purification of the peptides can be used and other salts with acids. For example, there can be used salts formed with sulfonic acids, such as methanesulfonate acid, n-butanesulfonic acid, p-toluensulfonate and naphthalenesulfonate.

The recommended method for isolation and purification of the compounds of formula 1 with simultaneous achievement of the target of a stable salt form is the way in which stable inorganic salts, such as sulfate and hydrochloride, get preparative purification via chromatography the concentration of 0.01-0.05% acetonitrile, THF, methanol or other acceptable solvent is an organic component. The pH value of the acidic eluent is kept in the range of pH 4-6, the exact pH value depends on the specific peptide and used the basic resin, for example, Bio-Rad AG-H resin in the hydroxyl form. After establishing the pH of the salt solution of Tripeptide, for example, sulfate or hydrochloride lyophilized obtaining purified salt in the form of a dry powder. In the example of this method is crude sulfate D-Phg-L-Pro-L-Arg-H contaminated by impurity epimerase sulfate D-Arg-H, dissolved in about 0,01% sulfuric acid and the solution is transferred into a column for HPLC with treatment phases of Vtdc18. For elution of the column for 10 hours using a gradient of 2-10% acetonitrile in 0.01% H2O4. Selected fractions containing the desired product fraction (according to the analytical RP-HPLC) combined. The pH value of the combined fractions set in the range of 4-4 .5 by adding resin Bio-Rad AG-G in hydroxyl form. After filtrowanie solution lyophilized and get pure sulfate D-Phg-L-Pro-L-Arg-H.

Compounds of the invention of formula 1 inhibit the action of thrombin in humans and animals. Inhibition of thrombin is shown in vitro ing the I (Kkfor the interaction between the test compound (inhibitor) and thrombin. Data presented in table obtained in the test in which the thrombin hydrolyzes the chromogenic substrate N-benzoyl-D-i.e. phenylalanyl-L-poured-L-arginyl-p-nitroanilide.

The test is carried out in 50 μl buffer (0.03 M Tris, 0.15 M NaCl, pH 7.4) with 25 μl of thrombin solution of 0.21 mg/ml powder thrombostat 0.06 M Tris, 0.3 M NaCl, pH 7.4) and 150 μl of an aqueous solution of a chromogenic substrate at a concentration of 0.25 mg/ml Add solutions of the test compounds (25 μl) in different concentrations. The degree of hydrolysis of the substrate is determined by analysis of the reaction mixture at 405 nm on the allocation of p-nitroaniline. Build standard curves deposition the concentration of free thrombin on the degree of hydrolysis. The use of standard curves degree of hydrolysis observed in the presence of test compounds, and then transferred to the values of free thrombin in the respective tests. Bound thrombin (associated with the test compound) is calculated by subtracting the amount of free thrombin observed in each trial, from a known initial amount of thrombin used in the test. The amount of free inhibitor calculate each of the connection).

Values FORkare hypothetical equilibrium constants of the reaction between thrombin and the test compound (S).

Thrombin + And Thrombin-AND

< / BR>
The value of Kkcalculated for the range of concentrations of the tested compounds and average values are given in units of liter per mole.

Anticoagulant activity of the compounds of the invention are defined in the standard tests. In the following table 2 presents the data obtained for representative compounds of the invention in the tests used to determine the time, prothrombin time, thrombin time and time activated partial thromboplastin (VACHT). Numerical values in the table refer to the concentrations of the tested compounds (ng/ml) required to prolong the coagulation 2 times in three trials. Detection of thrombin time carried out in the plasma and separately identified in the buffer system at pH 7.5.

Presented in table 2 data obtained using the device Maskiner company Tecan Inc. in tests carried out by the following method.

Programmirovanie time: 50 µl of plasma. 50 μl saline, 7 µl of the test solution, 50 MK the solution, 50 μl of bovine thrombin (2 units NI H/ml).

When determining the thrombin time in the buffer with pH 7.4 instead of use plasma fibrinogen.

FACT: 50 ál plasma, 50 μl of Actin (Dade County), 7 μl of the test solution, 50 μl of CaCl2(0.01 M).

Antithrombotic activity of representative compounds of the invention was determined in the in vivo tests conducted on rats. In tests using artificially created thrombosis in the carotid artery of rats, while the measured pour the dose of the test compound necessary for the maintenance of blood flow within fifty minutes after the time of occlusion. The test is carried out as follows.

In rats artificially create thrombosis damage to the carotid artery. For damage to the vessel used topically applying solution of ferric chloride (III). Male rats Sprague-Share (375-450 g) anaesthetize with xylazine (20 mg/kg, subcutaneously) and then ketamine. HCl (100 mg/kg, SC). Animals placed on a water blanket, which circulates water with a temperature of 37oC. Access to the carotid artery get through the middle of the cervical incision. For detection of the vessel and separate it from the shell using careful peeling. Under the artery pull silk pagestart on the tape recorder chart, equipped with a writing ink timer. Small tweezers with a solution (35%) immerse diskit (dia. 3 mm) from batmanesque filter paper No. 1. Disks cut out the same size using the pointed tube (3 mm in D.) stainless steel clamped in the Chuck of a drill press. The impregnated disks are placed on each carotid artery above thermocouple. The time between the imposition of FeCl3and time of a sharp reduction in temperature is celebrated as the time of occlusion of the vessel (USS). The average time required for the blockage of both vessels, used to represent VSS for each animal.

Compound dissolved in isotonic saline solution. For infusion solutions drug use syringe pump, and the solutions begin to infuse for 15 minutes before applying FeCl3and continue to pour 60 minutes after adding the HCl3. To determine the relationship between log10pour dosages and VSS damaged arteries build curves response to the input dose. Based on the curves determine the comparative indicator of antithrombotic activity by counting pour the dose needed to maintain blood flow within 50 min (ED 50 min).

The relationship between occlusion of the vessel and is healed. Proximally to thermocouple close to the carotid artery is placed pulsation probe flow Doppler. The probe used to measure changes in flow rate, resulting in a probe set in the time of absence of thrombosis, and when the inner diameter of the vessel remains constant in the strain of the blood flow. The zero line for the temperature and flow rate (determined pulsation in the direction of the flow meter Doppler model 545, University of Iowa, Bioengineering) set before applying a 35% aqueous solution of ferric chloride (III). The results recorded in the form of percentage deviations from the initial zero line (6 min before occlusion). The time at which the temperature of the vessel decreases rapidly, arbitrarily set as zero, and the temperature and flow velocity values before and after occlusion compared to this moment.

In the following table 3 contains the results obtained for the tested compounds in the above test with chemically induced thrombosis in rats.

Compounds of the invention inhibit the formation of blood clots without noticeable effect on the body's natural ability to lyse blood clots, for example, the compounds have nizkourovnova of blood clots in humans and animals, consisting in the conduct of a person or an animal inhibitory non-toxic doses of the compounds of formula 1. Anticoagulant compound is administered orally, parenterally, for example: intravenous blivemiami (centuries), intramuscular injection (C. M.) or subcutaneously (SC ). It is recommended that introduction to exercise century. century. infusion.

Effective, inhibiting the formation of blood clots dose is in the range of 5-1000 mg Mode dose may vary, for example, in the preventive purposes can be used daily single dose or may be an acceptable purpose in several doses, for example, 3 or 5 times a day. In case of critical situations, the compound of the invention administered centuries infusion with a speed of between about 1 mg/kg/h and 50 mg/kg/h, preferably about 2.5-25 mg/kg/h

The method of the present invention is used in combination with an agent lizirovania blood clots, for example, tissue plasminogen activator (PLA), modified Apt, streptokinase or urokinase. In cases when there was the formation of clots and artery or vein was blocked partially or completely, usually use the agent lizirovania blood clot. Connection izobreteyonija formation of a blood clot.

In the process of the invention it is desirable to use recommended by the invention compounds. For example, use the preferred compounds of the above type. For your preferred peptides include: N-Boc-D-phenylglycyl-L-prolyl-L-arginyl-aldehyde and N-methyl-D-phenylglycyl-L-propyl-L-arginine-aldehyde, which are in the form of a salt, such as sulfate or hydrochloride. Especially preferred for use in this way is sulfate N-(D-1,2,3,4-tetrahydroisoquinoline-1 oil)-2-prolyl-2-arginine-aldehyde.

The invention also provides pharmaceutical preparative formula for use in the above-described therapeutic way. Pharmaceutical formulations of the invention include an effective, inhibiting the formation of blood clot amount of the compounds of formula 1 and pharmaceutically acceptable carrier. For oral administration protivotoksicheskoe compound prepared in the form of gelatin capsules or tablets which may contain various excipients, such as binders, lubricating agents, structuring agents, etc. For parenteral use of the antithrombotic agent is administered in pharmaceutically praml"ptx2">

Antithrombotic compound of the invention can be transformed into a single dosage form containing a dose between about 1 mg and 1000 mg, preferably the compound are in the form of pharmaceutically acceptable salts, such as sulfate, acetate or phosphate. For example, a unit dose preparation contains 5 mg of sulfate N-BOC-D-phenylglycyl-L-prolyl-L-arginine-aldehyde in 10 ml sterile glass vials. Another example of a single dosage form contains 10 mg of sulfate N-methyl-D-phenylglycyl-L-prolyl-L-arginine-aldehyde in 20 ml of isotonic NaCl solution enclosed in a sterile vial.

The preferred preparative form represents a single dosage form containing 5-50 mg sulfate N-(D-1,2,3,4-tetrahydroisoquinoline-L oil)-L-prolyl-L-arginine-aldehyde in sterile vials.

The compounds can be administered in various ways, including oral, rectal, transdermal, subcutaneous, intravenous, intramuscular, nasal. Compounds of the present invention preferably before assigning to receive converted to a formulation comprising an effective amount of the compounds of formula 1 or its pharmaceutically acceptable salt, /P> The active ingredient in such formulation is from 0.1% to 99.9% by weight of the form. "Pharmaceutically acceptable" means that the carrier, diluent or excipient must be compatible with other ingredients of the formulation and not to harm the recipient.

These preparative form get by known methods using known or easily available ingredients. The compositions of this invention can be formulated so as to provide quick, supported on one level or delayed release of the active ingredient after administration to the patient, using known techniques. Upon receipt of a composition or compositions of the present invention the active ingredient is usually mixed with a carrier, or diluted by a carrier or sign inside the carrier, which may be in the form of capsules, tablets, paper or other container. When the carrier serves as a diluent, it may be solid, semi-solid or liquid material which acts as a binder, excipient or medium for the active ingredient. Thus, the compositions can be in the form of tablets, pills, powders, pellets, lozenges, elixirs, suspensions, emulsions, Rastogi, sterile injectable solutions, sterile packaged powders, etc.

Below are examples of preparative forms, which illustrate but do not limit the scope of the invention. "Active ingredient" of course, means a compound of formula 1 or its pharmaceutically acceptable salt, or MES.

Preparative form 1

Hard gelatin capsules are prepared using the following ingredients quantity (mg/capsule)

The active ingredient 250

Starch, dried 200

Magnesium stearate 10

The total weight of 460 mg

Preparative form 2

Tablets are prepared using the following ingredients quantity (mg/tablet):

The active ingredient 250

Microcrystalline cellulose 400

Silicon dioxide 10

Stearic acid 5

The total weight of 665 mg

The components are mixed and pressed to form tablets each weighing 665 mg

Pre-form 3

Aerosol solution is prepared using the following components (weight):

The active ingredient of 0.25

Ethanol 25,75

Spray the substance, the propellant was 22 (Chlorodifluoromethane) 70,00

The total weight of 100.00

The active substance is mixed with ethanol, and the mixture we use is in then served in a stainless steel container and diluted with the remaining propellant. Then set the container valve.

Preparative form 4

Tablets, each containing 60 mg of active ingredient are made as follows:

The active ingredient 60 mg

Starch 45 mg

Microcrystalline cellulose 35 mg

Polivinilpirolidon (as 10% solution in water) 4 mg

Carboximetilkrahmal of 4.5 mg sodium

Magnesium stearate 0.5 mg

Talc 1.0 mg

The total number 150,00 mg

The active ingredient, starch and cellulose are passed through a sieve with holes N 45 met US and mix thoroughly. An aqueous solution containing polyvinylpyrrolidone, mixed with the resulting powder and the mixture is passed through a sieve with apertures 14 met US. The granules thus obtained, dried at 50oC and passed through sieve No. 18 in the USA. Carboximetilkrahmal sodium, magnesium stearate and talc, previously passed through a sieve with holes N 60 USA, then added to the granules which, after mixing, pressed into tablets and receive tablets, each of which weighs 150 mg.

Preparative form 5.

Capsules, each containing 80 mg of the active ingredient, are prepared as follows:

R> The total number of 200 mg

The active ingredient, cellulose, starch and magnesium stearate are blended, passed through a sieve with holes N 45 and fill with a mixture of hard gelatin capsules in 200 mg

Preparative form 6

Suppositories, each containing 225 mg of active ingredient, are prepared as follows:

The active ingredient 225 mg

Glycerides of saturated fatty acids 2000 mg

The total number 2225 mg

The active ingredient is passed through a sieve with holes N 60 U.S. and suspended in the glycerides of saturated fatty acids, previously melted using the minimum heat needed. The mixture is then poured into the mold for suppositories with a nominal capacity of 2 g and allowed to cool.

Preparative form 7

Suspensions, each containing 50 mg of active substance per 5 ml dose, are as follows:

The active ingredient 50 mg

Sodium carboxymethyl cellulose 50 mg

Syrup 1.25 ml

The solution petrol acid 0.10 ml

Odorant at the discretion of the

Dye at the discretion of the

Purified water to total amount 5 ml

The active ingredient is passed through a sieve with the hole is actor benzoic acid, the perfume and dye diluted with a portion of water and added with stirring. Then add sufficient water to obtain the desired volume.

Preparative form 8.

Preparative form for intravenous infusion is prepared as follows:

The active ingredient 100 mg

Isotonic saline 1,000 ml

The solution presented above ingredient is usually administered intravenously at a rate of 1 ml per minute.

The following examples are given to further illustrate the invention and not intended to be limiting thereof.

Values of Rfthe following examples were determined using thin-layer chromatography on Kieselgel 60F-254 (Merck, Darmstadt) in the following solvent system;

(A) chloroform-methanol-acetic acid (135:15:1.about./on);

(C) ethyl acetate-acetic acid, absolute ethanol (90:10:10./about./about. ).

In the examples used the following analytical HPLC methods.

Method 1. Chromatograph the wouterse A with column treatment phases video recorder WITH18size 0,46 cm x 10 cm Chromatogram obtained on LDC at 220 nm using a gradient And 0.01 M ammonium acetate and acetonitrile.

Method 2. the label of the gradient And either 0.01 M ammonium acetate, or In acetonitrile.

Used in the examples, the abbreviations have the following meanings:

Amino acids: Arg, arginine, Pro is Proline, the Phg phenylglycine;

Vos tert-butyloxycarbonyl;

BZ1 benzyl;

bZ benzyloxycarbonyl;

DCC dicyclohexylcarbodiimide;

DMF is dimethylformamide;

DMSO dimethyl sulfoxide;

MS-the Belarusian library Association mass spectrometry with fast atom bombardment;

MS-DP mass spectrometry with desorption in field;

THF tetrahydrofuran;

TLC thin layer chromatography.

Example 1. N-Boc-D-phenylglycyl-L-prolyl-L-arginine-aldehyde (BOC-D-Phg-Pro-Arg-H), polysulfate

1) Boc-D-Phg-Pro-OBZ1

A solution of Boc-D-phenylglycine (15 mg, 59.7 mmole) and hydrochloride benzyl ester of Proline (14, 43 g, 59.7 mmole) followed by addition of hydrate of 1-hydroxybenzotriazole (8,1 g, 59.7 mmole) and DCC (12.3 g, 59.7 mmole). The reaction mixture is stirred 3 days at room temperature, then filtered and the filter, evaporate in vacuum to obtain oil. The oil obtained is dissolved in 200 ml ethyl acetate and 150 ml of water and after shaking, the organic layer is separated, washed three times with 100 ml of 0.1 N. HCl, once with 150 ml of water, three times with 100 ml 5% sodium bicarbonate and again with 150 ml of water. Prom is 4.8 g (95% of theory (BOC-D-Phg-Pro-ObZ1, TLC Rf (A) IS 0.75; - MS-Belarusian library Association 439 (M+).

2) Boc-D-Phg-Pro-OH

Obtained in the aforementioned manner Boc-Phg-Pro-ObZ1 (24.5 g, 55.7 mmole) is dissolved in 40 ml of DMF and to the solution was added 225 ml of isopropyl alcohol and 1 g of 5% Pd on coal as a catalyst. Through distribution of the gas tube into the reaction mixture propulsive nitrogen, then 16 hours propulsive hydrogen, then 5 minutes pass nitrogen. The catalyst is filtered off through filter layer hyflo and evaporation of the filtrate to dryness in a vacuum get a solid residue. The residue is crystallized from diethyl ether containing a small amount of ethyl acetate. Received 10,35 g (yield 53% ) of the product removal of the ester group: BOC-D-Phg-Pro (2), TLC Rf (A), 0,32; MS-Belarusian library Association 349 (MN+);1H-NMR (DMSO-d6) : of 1.35 (s, N), 1,71-2,1 (m, 4H), 3,1 (m, 1H), 3,74 (m, 1H), 4,2 (m, 1H), the 5.45 (d, 1H), to 7.09 (d, 1H), 7,25 to 7.4 (m, 5H), 12.5cm (ush.with. 1H).

3) BOC-L-Arg(CbZ)-OH

In a three-neck round-bottom flask in 240 ml of 5 N. NaOH dissolve the hydrochloride of N-Mos-arginine (BOC-Arg-OH.HCl) (82,1 g, 250 mmol). The solution is cooled to -5oC and added dropwise over 55 minutes add benzylchloride (143 ml, 1 mol, 4 acsw.), maintaining a pH in the range of 13.2 to 13.5 by adding 5 N. NaOH (250 ml). Upon completion of addition, chloroformate the reaction mixture paramashiva is collected and twice extracted with diethyl ether portions 40 ml. The aqueous layer was acidified to pH 3 by addition of 3 n H2SO4(560 ml) and extracted with 550 ml of ethyl acetate. The separated aqueous layer is extracted once with ethyl acetate and the extract combined with the previously obtained extracts of ethyl acetate. The combined extracts washed with water, dried over MgSO4and evaporated to dryness in a vacuum. The residue is rinsed with ether, and the precipitated product is filtered and dried. Received of 66.1 g (65% of theory) 3) (BOC-Arg (CbZ)-OH, TLC Rf=(C) 0,43; MS-DP 408 (M+).

1H-NMR (CDCl3d: of 1.42 (s, N), 1,61 is 1.91 (m, 4H), 3,23-to 3.41 (m, 2H), 4,17 (d, 1H), total of 5.21 (s, 2H), 5,62 (d, 1H), 7.3 to 7,42 (m, 6N), of 8.37 (m, 1H).

4) BOC-Arg(Cbz)-lactam

Solution (3) BOC-Arg(CbZ)-OH (66 g, rate £ 0.162 mole), obtained as described above, in 230 ml of dry THF cooled in a bath of ice in acetone to -10oC. To the cold solution was added N-methylmorpholine (18.7 ml of 1.05 equiv.) then add isobutylparaben (22.5 ml, of 1.05 equiv.) and the mixture is stirred for 5 minutes at -10oC. then add triethylamine (23,5 ml of 1.05 equiv.) and the mixture is stirred for 1 h at -10oC and 1 h at room temperature. The reaction mixture was transferred into a one liter of a mixture of water with ice, resulting precipitated product (4). The precipitate is filtered off, washed with cold water, dried in vacuum and Krist ).

1H-NMR (CDCl3d: to 1.48 (s, N), 1,78-to 1.98 (m, 2H), 2,5 (m, 1H), 3,41 (m, 1H), 4,43 (m, 1H), 4,9 (m, 1H), 4.16 the (s, 2H), 5,27 (m, 1H), 7,28 was 7.45 (m, 6N), 9,41 (m, 1H), 9,68 (m, 1H).

5) Arg(CbZ)-lactam, triptorelin

Boc-Arg(CbZ)-lactam (4) (38 g, 0,097 mole) is mixed with 200 ml triperoxonane acid containing 20 ml of anisole, and the mixture is stirred one hour at 0oC. Then the reaction mixture is evaporated in vacuo without heating and the residue add 400 ml of diethyl ether. The solid product is filtered off, washed with diethyl ether and dried in vacuum. Obtained as a product 40.5 trifenatate (5), TLC Rf (C) 0,29; MS-DP 291 (MN+).

6) Boc-D-Pro-Arg(CbZ)-lactam

To a solution of BOC-D-Phg-Pro (14.5 g, to 41.6 mmole, obtaining see part 2 above) in 80 ml DMF, cooled to -15oC, was added 4.6 ml of N-methylmorpholine with the subsequent addition of 5.4 ml of isobutylacetate and then the reaction mixture is stirred for two minutes at -15oC.

In a separate flask in 30 ml of DMF is dissolved Arg(CbZ)-lactam. TN (15.3 g, of 37.8 mmole, obtaining see part 5) above), the solution is cooled to 0oC and to it was added 4.6 ml of N-methylmorpholine. After stirring the solution for two minutes at 0oC it is transferred into a solution of Boc-D-Phg-Pro received prior to the above method. Received Reece about the day. Then to the reaction mixture add 5% solution of NaHCO3(5 ml) and subsequent evaporation in vacuo receive oil. The oil is dissolved in 175 ml of ethyl acetate and 150 ml of water added to the solution. After shaking, the organic layer is separated, washed with 5% NaHCO3and water, dried over MgSO4and evaporation to dryness in vacuo obtain 23 g (yield 98%) of Boc-D-Phg-Pro-Arg(CbZ)-lactam (6) in the form of an amorphous substance, TLC Rf (A) 0,72; MS-Belarusian library Association 621 (MN+).

7) Boc-D-Pro-Arg(CbZ)-H

In 200 ml of dry THF was dissolved lactam (6) (23 g, 37 mmol, obtaining see part 6) above) and the solution is cooled to -15oC in nitrogen atmosphere. To the cooled solution dropwise over 10 min was added 1 M solution of sociallyengaged in THF (37 ml, 37 mmol) and upon completion of the addition the reaction mixture is heated to 0oC, and stirred for 1 h a Mixture of 12 ml of THF and 12 ml of 0.5 N. H2SO4slowly dropwise within 10 min added dropwise to the reaction mixture. Then the reaction mixture is diluted with 200 ml ethyl acetate and 200 ml of water and after shaking, the organic layer separated. The organic layer is washed three times with water portions of 150 ml, dried on MgSO4and after evaporation to dryness in vacuo gain of 19.2 g (yield 83%) of Boc-D-Phg-Pro-Aro-H, MS-the Belarusian library Association 623 (MN+); (0.5, CHCl3is I) and to the solution add 29,2 ml of 1 N. H2SO4b 2 u 10% Pd-C. suspension 5 min through the gas distribution tube propulsive nitrogen with subsequent transmission 4 h of hydrogen. At the end of the recovery again 5 min propulsive nitrogen. The reaction mixture was filtered through a layer hiflo with the removal of the catalyst and evaporation in vacuo of the filtrate concentrated to 100 ml. of water to the concentrate is added 200 ml of n-butanol and the organic layer separated from the aqueous layer. The aqueous layer was extracted three times with n-butanol portions of 100 ml, the extracts are combined and added to the organic layer. The organic layer is evaporated to dryness in vacuo, the residue is rinsed with a mixture of diethyl airdisplay ether (1:1 vol./vol.), the solid is filtered off and dried in vacuo get 10,26 g of dry product (8). The crude product is dissolved in 10% aqueous acetonitrile and the solution is transferred into a column (7.5 cm x 53 cm) resin HP-20, previously described in the balance of 10% aqueous acetonitrile. Product elute from the column stepwise elution with increasing concentrations of acetonitrile in water (10%-12%-15%). Select a number of factions that are analyzed for the presence of the product by HPLC with reversed phase. Containing the product fractions are combined and their evaporation dowiana in RP-HPLC (method 2, 10-50% In 45 min), and 32.3 minutes

Example 2. N-(tert-Butyloxycarbonyl)-D-phenylglycyl-L-prolyl-L-arginine-aldehyde (BOC-D-Phg-Pro-Arg-H), diacetate.

To a solution of Boc-D-Phg-Pro-Arg(CbZ)-N (receipt see part 7) of example 1, 38, 61 mmol) in 500 ml of isopropyl alcohol containing a 7.1 ml (2 EQ.) acetic acid added as a catalyst, 2 g of 10% Pd and coal. The mixture was washed with flowing naturally through the tube with nitrogen (5 min) and then through the mixture for 24 h miss hydrogen. At the end of recovery through the mix again 5 min flow of nitrogen. Then the reaction mixture was filtered through a layer Hiflo with the removal of the catalyst and evaporation of the filtrate to dryness gain of 33.6 g of crude product as an amorphous substance. The product was then purified portions 5 g column (5 cm x 25 cm) video recorder C18. The resulting Tripeptide elute 8 h with a gradient of 10-30% acetonitrile in 0.01 M ammonium acetate. Collect the fractions containing according to HPLC with treatment phases of the product fractions are combined and dried by freezing. Received of 11.7 g (35%) capital Tripeptide with the following indicators: MS-Belarusian library Association 489 (MN+).

Amino acid analysis: Phg 1.07, and Pro 0,94. (from 0.6, CHCl3).

Elemental analysis calculated for C28H44N6O9

Example 3. N-BOC-D-1-naphthylvinyl-Pro-Arg-H, diacetate (0.5, 50% acetic acid); MS-Belarusian library Association 539 (MN+);

HPLC (method 1, gradient 20-60% b for 60 min), retention time of 42 minutes

Example 4. N-Boc-D-2-naphthylvinyl-Pro-Arg-H, diacetate HPLC (method 2, the gradient of 30-60% In 60 min), retention time 18 min

Elemental analysis calculated for C32H46N6O9:

Theory: 58,35, N? 7.04 baby mortality, N OF 12.76%

Found: 58,59, N 6,83, N 13,03%

Example 5. N-Boc-D-(4-hydroxyphenylglycol)-Pro-Arg-H, diacetate HPLC (method 2, the gradient 10-40% b for 40 min), retention time of 26.5 min

Amino acid analysis: 4-hydroxyphenylglycol 0,99, Proline 1,01.

Example 6-23 Listed in the following table 4 compounds according to the methods of example 1 by using these amino acids or substituted acetic acids (A(C=O)) instead of used in example 1-phenylglycine. All compounds in table 1 are in the form of acetates.

Example 27 D-1,2,3,4-Tetrahydroisoquinoline-1-oil-L-prolyl-L-orgininally, sulphate

To a solution of isoquinoline-1-carboxylic acid (12.5 g, 0,072 mol) in 185 ml of glacial acetic kitlg/cm2in the apparatus for hydrogenation Parra 24 h, the Reaction mixture was filtered through a filter layer (Celite) to remove catalyst and the filtrate is evaporated to dryness in a vacuum. The solid residue is rinsed with water, filtered and after drying receive 8 g (yield 63% ) DL-1,2,3,4-tetrahydroisoquinoline-1-carboxylic acid. DP-mass spectrum 178 (MN+);1H-NMR (DMSO-d6) : 2,8-3 (m, 1H), 3.15 in (m, 1H), 3,3-3,4 (m, 2H), 7,05-7,25 (m, 4H), and 7.7 (m, 1H).

The resulting product (7,08 g of 0.04 mole) dissolved in 2 N. NaOH (40 ml, of 0.08 mole) and to the solution was added 40 ml of tert-butyl alcohol and 10.5 g (0,048 mole) di-tributylphosphate. After stirring 24 h at room temperature, the major quantity of tert-butyl alcohol is evaporated from the reaction mixture. The resulting aqueous solution extracted with diethyl ether, the aqueous layer separated and acidified with 2 N. HCl to pH 2. The acidified aqueous phase is extracted with ethyl acetate, the extract dried over MgSO4and evaporated to dryness in a vacuum. The residual oil is dissolved in diethyl ether and to the solution is added to 7.9 ml (0.04 mole) of dicyclohexylamine. After vydergivanija 4 h at 4oC sediment dicyclohexylamine salt of N-BOC-DL-1,2,3,4-tetrahydroisoquinoline-1-carboxylic acid is filtered off, washed with diethyl ether islandy for C27H42N2O4:

Theory: 70,71, H 9,23, N 6,11%

Found: 71,07, N 9,37, N 5.87 per cent

The BOC-protected derivative (73,4 g, 160 mmol) is suspended in 200 ml of ethyl acetate, washed suspension of 1.5 N. citric acid and water, dried over MgSO4and evaporated to dryness in a vacuum. The residual oil is dissolved in ethyl acetate, the solution is cooled to 0oC and to the solution was added 2,4,5-trichlorophenol (31,6 g, 160 mmol) followed by addition of DCC (33 g, 160 mmol). The reaction mixture is stirred one hour at 0oC and 1.5 h at room temperature. Then the reaction mixture was cooled to 0oC, the precipitate is filtered off and the filtrate is evaporated to dryness in a vacuum. The residual oil is dissolved in 100 ml of pyridine and to the solution was added Proline (18,42 g, 160 mmol) and triethylamine (22,3 ml, 160 mmol). After stirring 24 h at room temperature, the reaction mixture is evaporated to dryness in a vacuum. The residue is dissolved in ethyl acetate, the solution was added water and add 2 N. NaOH establish a pH of 9.5. The aqueous layer was separated, acidified with 2 N. HCl to pH 2 and extracted with ethyl acetate. The extract is dried over MgSO4, filtered and evaporated to dryness in a vacuum. The oily residue is dissolved in methylene chloride and ethyl acetate. After curing 4 hours at Kristy methylene-ethyl acetate. The obtained Boc-D-1,2,3,4-tetrahydroisoquinoline-1-oil-L-Proline (Boc-I-Tig-Pro-OH) is dried in the form of a solid product in a vacuum and get a 19.6 g (yield 33%) of pure product. TLC Rf (A) OF 0.44; MS-Belarusian library Association: 375 (MH+).

Elemental analysis calculated for C20H26N2O5:

Theory: 64,15, N 7,00, N OF 7.48%

Found: 63,26, N 6,98, N 7,52%

(0.5, methanol).

In the first flask in 100 ml of DMF is dissolved Boc-D-1-Tig-Pro (17.8 g, and 47.5 mmole), the solution is cooled to -15oC, then add to 5.3 ml (52,3 mmole) N-methylmorpholine and 6.2 ml (47,5 mmole) of isobutylacetate and the mixture is stirred for two minutes at -15oC.

In the second flask in 40 ml of DMF is dissolved With-protected arginine-lactam as trifenatate (Arg(Z)-lactam, TN) (19.2 g, and 47.5 mmole), the solution is cooled to 0oC, and then added with 5.3 ml (52,3 mmole) N-methylmorpholine. Before transferring into the first flask and the mixture is stirred for 2 min at 0oC. the Reaction mixture was stirred 4 h at -15oC, then for about days slowly warmed to room temperature, then add 5 ml of 5% NaHCO3. Evaporation of the reaction mixture in vacuo receive oil. The oil is dissolved in 175 ml of ethyl acetate and to the solution was added 150 ml of water. The organic layer is separated, the industry is t to dryness in vacuo to obtain 24.3 g (yield 79%) of Boc-D-1-Tig-Pro-Arg(Z)-lactam in the form of an amorphous substance.

TLC Rf (A) 0,71;

MS-THE BELARUSIAN LIBRARY ASSOCIATION: 647 (MN+);

(0.5, chloroform).

Obtained in the aforementioned manner Arg (Z)-lactam (23,4 g, and 36.2 mmole) is dissolved in 300 ml of dry THF and the solution is placed in an atmosphere of N2. Then the solution is cooled to -20oC and cooled the solution for 30 min added dropwise 37 ml of 1 M solution in THF of sociallyengaged. Upon completion of addition, the mixture is stirred for 30 min at -20oC, followed by dropwise within 10 min was added a mixture of 20 ml THF and 20 ml of 0.5 N. H2SO4. The reaction mixture was added 400 ml of ethyl acetate and to the solution was added 400 ml of water. The organic layer is separated, washed twice with water portions 150 ml) and dried over MgSO4. Evaporation in vacuum, washed and dried solution obtain 21 g (yield 89%) of Boc-D-1-Tig-Pro-Arg(Z)-H in the form of an amorphous substance, TLC Rf (A) of 0.28, 0.28 in. Obtained as described above is derived Arg(Z) hydronaut below method With removal of the protective group. The resulting product (18,1 g of 27.9 mmole) is dissolved in 200 ml of THF and 80 ml of water and to the solution was added 28 ml of 1 N. H2SO4and 3 g of 5% Pd on coal. Through the suspension for 5 min propulsive nitrogen flowing through the barbaterom, then 5 h miss hydrogen and again 5 min propulsive nitrogen. Calibratable layers separated. The aqueous layer was extracted three times with n-butanol portions of 100 ml and the extracts added to the organic layer. The organic layer is evaporated in vacuo, the residue is rinsed with a mixture of diethyl ether-diisopropyl ether (1:1 vol./vol.), the solid product is filtered off and dried in vacuo get 11,08 g crude product.

The product is distilled and transferred to sulfate in the following way. Obtained as described above crude product was dissolved in 20 ml of water and 20 ml of 10 N. H2SO4. The solution is heated for 25 min at 50oC, cooled to room temperature and adding resin Bio-Rad AG1-XB (hydroxide form) set pH 4. The resin is separated from the solution by filtration and lyophilisation of the solution get 8.44 grams of crude product in the form of sulfate (D-1-Tig-Pro-Arg-h H2SO4).

The resulting sulfate is dissolved in 0.01% H2SO4and the solution is transferred into two columns for HPLC with treatment phases with a size of 5 cm x 25 cm (resin video recorder WITH18) connected in series. For elution salt obtained using the gradient of increasing acetonitrile (2-10%). Selected fractions and unite on the basis of the results of the analytical RP-HPLC. Added to combined fractions resin AG1-X8 (analysis, the Ute with the removal of the resin and the filtrate lyophilized. Obtained 2.4 g (57% of theory) of the pure product.

MS-THE BELARUSIAN LIBRARY ASSOCIATION: 415 (MN+);

(0.5, 0,01 said. H2SO4);

Amino acid analysis: Pro 0,92, Tig 1,00;

Elemental analysis calculated for C21H32N6O7S:

Theory: 49,21, N 6,28, N 16,29, S OF 6.26%

Found: 51,20, N 6,17, N 16.88 In, S 5.37 Percent

1. Derived tripeptides as R - or RS-form General formula I

< / BR>
where a group of the formula

< / BR>
R phenyl group of the formula

< / BR>
where a and a1independently hydrogen, lower alkyl, lower alkoxy, halogen, trifluoromethyl or hydroxy; thienyl; neftel, unsubstituted or mono - or disubstituted by lower alkoxy group; or cyclohexyl;

R1hydrogen, methyl or ethyl;

Lower alkyl, lower alkoxy or amino group of the formula-N(R2) (R3), in which R2and R3independently hydrogen or lower alkyl, or R2hydrogen, and R3acetyl or N-Vos (lower)alkyl, provided that when R1methyl or ethyl, then b is other than methyl or ethyl,

or a bicyclic group of the formula

< / BR>
Q-CH2-CH2or

Q Y-CH2-;

R5hydrogen or tert-butoxycarbonyl;1 R6hydrogen, indicated by the dotted line circle points="ptx2">

2. Derived tripeptides on p. 1 of General formula

< / BR>
where a group of the formula

< / BR>
where R is the phenyl group of the formula

< / BR>
where a and a1independently hydrogen, lower alkyl, lower alkoxy, halogen, trifluoromethyl or hydroxy,

or R thienyl; naphthyl, unsubstituted or mono - or disubstituted by lower alkoxygroup,

or R is cyclohexyl;

R1hydrogen, methyl or ethyl;

Lower alkyl, lower alkoxy or amino group of the formula-N(R2)(R3), in which R2and R3independently hydrogen or lower alkyl, or R2hydrogen, and R3acetyl or N-Vos(lower)alkyl, provided that when R1methyl or ethyl, then b is other than methyl or ethyl.

3. Derived tripeptides on p. 2, in which the amino group of the formula-N(R2)(R3),where R2and R3independently is hydrogen or lower alkyl, or R2hydrogen, and R3tert.butoxycarbonyl.

4. Derived Tripeptide on p. 3, representing the N-BOC D-phenylglycyl-L-prolyl-L-arginal or its pharmaceutically acceptable non-toxic salt.

5. Derived Tripeptide on p. 3, representing the N-methyl-D-phenylglycyl-L-prolyl-L-arginal or its pharmaceutically acceptable non-toxic, no the genus or tert.butoxycarbonyl group;

R6hydrogen;

dashed circle indicates that the ring is bergerocactus,

or their pharmaceutically acceptable non-toxic salt.

7. Derived Tripeptide on p. 6, representing D-1,2,3,4-tetrahydroisoquinoline-1 - oil-L-prolyl-L-arginineresearch.

8. Derived Tripeptide on p. 6, representing D-perhydrosqualene-1 - yl-carbonyl-L-prolyl-L - arginine-aldehyde or its pharmaceutically acceptable salt.

9. Derived Tripeptide on p. 6, representing D-perhydrosqualene-3-yl-carbonyl-L-prolyl-L-arginine-aldehyde or its pharmaceutically acceptable salt.

10. Derived Tripeptide on p. 6, representing D-1,2,3,4-tetrahydroisoquinoline-3-oil-L-prolyl-L-arginine-aldehyde or its pharmaceutically acceptable salt.

11. Derived tripeptides according to any one of paragraphs.1 10 with antithrombotic properties.

12. The pharmaceutical composition inhibiting the clotting of blood containing the active compound, which represents the derivative Tripeptide in combination with one or more pharmaceutically acceptable carriers, excipients or diluents, wherein in cacpa p. 12, characterized in that it is intended for oral administration and comprises an active compound in an amount of about 1 to 1000 mg per unit dose, in combination with such excipients as gelatin, binder, lubricant agent, or the agent of disintegration.

14. The composition according to p. 12, characterized in that it is intended for parenteral use, and includes an active compound in an amount of about 1 to 1000 mg per unit dose in a diluent, such as isotonic saline.

Priority signs:

28.09.90 when

< / BR>
where R is a phenyl group

< / BR>
where a and a1independently hydrogen, lower alkyl, lower alkoxy, halogen, trifloromethyl or hydroxyl, or R thienyl; naphthyl, unsubstituted or mono - or di-substituted lower alkoxygroup, or R is cyclohexyl; R1hydrogen, methyl or ethyl; lower alkyl, lower alkoxy or amino group of the formula NR2, R3where R2and R3independently hydrogen or lower alkyl, or R2hydrogen, and R3acetyl or N Vos(lower)alkyl, provided that when R1methyl or ethyl, then, is different from methyl or ethyl.

06.09.91, when a group of the formula

< / BR>
where R5hydrogen or tert.butoxycarbonyl the

 

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3 cl, 16 tbl, 9 ex

FIELD: organic chemistry, medicine.

SUBSTANCE: invention represents ligands MC-4 and/or MC-3 of the formula (I): , wherein X means hydrogen atom, -OR1, -NR1R1' and -CHR1R1' wherein R1 and R1' are taken among the group: hydrogen atom, (C1-C6)-alkyl and acyl; (1) each R2 is taken independently among the group: hydrogen atom, (C1-C6)-alkyl; or (2) (a) R2 bound with carbon atom that is bound with X and Z1 and substitute R5 can be optionally bound to form carbocyclic or heterocyclic ring that is condensed with phenyl ring J; or (b) R2 bound with carbon atom that is bound with ring Ar can be bound with R7 to form ring condensed with ring Ar; each among Z1, Z2 and Z3 is taken independently from the following groups: -N(R3e)C(R3)(R3a)-, -C(R3)(R3a)N(R3e)-, -C(O)N(R3d)-, -N(R3d)C(O)-, -C(R3)(R3a)C(R3b)(R3c)-, -SO2N(R3d)- and -N(R3d)SO2- wherein each among R3, R3a, R3b and R3c, R3d, R3e when presents is taken independently among hydrogen atom and (C1-C6)-alkyl; p is a whole number from 0 to 5 wherein when p above 0 then R4 and R4' are taken among hydrogen atom, (C1-C6)-alkyl and aryl; R5 represents 5 substitutes in phenyl ring J wherein each R5 is taken among hydrogen atom, hydroxy-, halogen atom, thiol, -OR12, -N(R12)(R12'), (C1-C6)-alkyl, nitro-, aryl wherein R12 and R12' are taken among hydrogen atom and (C1-C6)-alkyl; or two substitutes R5 can be bound optionally to form carbocyclic or heterocyclic ring that is condensed with phenyl ring J; q = 0, 1, 2, 3, 4 or 5 wherein when q above 0 then R6 and R6' are taken among hydrogen atom and (C1-C6)-alkyl; Ar is taken among the group consisting of phenyl, thiophene, furan, oxazole, thiazole, pyrrole and pyridine; R7 are substitutes at ring Ar wherein each R7 is taken among hydrogen, halogen atom, -NR13R13', (C1-C6)-alkyl and nitro- wherein R13 and R13' are taken among hydrogen atom and (C1-C6)-alkyl; r is a whole number from 0 to 7 wherein when r is above 0 then R8 and R8' are taken among hydrogen atom and (C1-C6)-alkyl; B is taken among -N(R14)C(=NR15)NR16R17, -NR20R21, heteroaryl ring and heterocycloalkyl ring wherein R14-R17, R20 and R21 are taken independently among hydrogen atom and (C1-C6)-alkyl; s = 0, 1, 2, 3, 4 or 5 wherein when s is above 0 then R and R9' are taken among hydrogen atom and (C1-C6)-alkyl; R10 is taken among the group consisting of optionally substituted bicyclic aryl ring and optionally substituted bicyclic heteroaryl ring; D is taken among hydrogen atom, amino- and -C(O)R11 wherein R11 is taken among the following group: hydroxy-, alkoxy-, amino-, alkylamino-, -N(R19)CH2C(O)NH2 wherein R19 represents (C1-C6)-alkyl, -NHCH2CH2OH and -N(CH3)CH2CH2OH, or its isomers, salts, hydrates or biohydrolysable ester, amide or imide.

EFFECT: valuable medicinal properties of compounds.

18 cl, 107 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: it is suggested to apply Pro-Gly-Pro tripeptide known as anticoagulant to keep stable norglycemia and stable normoinsulinemia in circulation at no side effects because the above-mentioned tripepetide is being natural human and animal metabolite.

EFFECT: higher efficiency of application.

3 cl, 5 ex, 2 tbl

FIELD: pharmaceutics.

SUBSTANCE: the set of components is suggested containing: (a) pharmaceutical preparation including low-molecular thrombin inhibitor or its pharmaceutically acceptable derivative in the mixture with pharmaceutically acceptable adjuvant, solvent or carrier; and (b) pharmaceutical preparation including pre-medicine of low-molecular thrombin inhibitor or pharmaceutically acceptable derivative of this pre-medicine in the mixture with pharmaceutically acceptable adjuvant, solvent or carrier, where components (a) and (b), each of them, should be taken in the form suitable to be introduced together; it is, also, suggested to apply this set of components for treating the state at which it is necessary or preferably to inhibit thrombin. The innovation enables to treat thrombotic states such as thrombosis of deep veins and pulmonary embolism.

EFFECT: higher efficiency of application.

30 cl, 1 tbl

FIELD: medicine, experimental medicine.

SUBSTANCE: one should introduce tripeptide Pro-Gly-Pro for laboratory animals as injections at the quantity of 0.09-1.0 mg/kg body weight, and, also, gelatin as fodder additive. The method suggested enables to suppress appetite, decrease the quantity of fodder intake that leads to decreased body weight as a result.

EFFECT: higher efficiency.

2 cl, 5 dwg, 5 ex

FIELD: medicine, chemistry of peptides, amino acids.

SUBSTANCE: invention relates to novel biologically active substances. Invention proposes the novel composition comprising peptides of the formula: H-Arg-Gly-Asp-OH and H-Tyr-X-Y-Glu-OH wherein X means Gln and/or Glu; Y means Cys(acm) and/or Cys. The composition shows ability to inhibit proliferative activity of mononuclear cells, to induce suppressive activity and their ability for secretion of cytokines TNF-1β (tumor necrosis factor-1β) and IL-10 (interleukin-10 ).

EFFECT: simplified method for preparing composition, valuable medicinal properties of composition.

4 cl, 16 tbl, 9 ex

FIELD: biotechnology.

SUBSTANCE: invention relates to a method for preparing foodstuff containing hypotensive peptides and it using as an anti-hypertensive agent that can be used as a foodstuff. Method involves stages for fermentation of casein-containing fermenting material with lactobacillus microorganism, nanofiltration of the prepared peptide-containing fermentation product and isolation of the product. Prepared product is used as an anti-hypertensive agent and as a foodstuff also. Invention provides preparing a foodstuff with the high content of hypotensive peptides enriched by bivalent ions.

EFFECT: improved preparing method of foodstuff.

22 cl, 1 dwg, 4 ex

FIELD: medicine, pharmacy.

SUBSTANCE: invention relates to medicinal agent used for correction of metabolic vascular syndrome and diseases accompanying with vascular wall penetrability disorder and capillaries fragility and can be used as agent enhancing resistance of capillaries. Invention proposes peptide lysyl-glutamyl-aspartic acid of the general formula: H-Lys-Glu-Asp-OH corresponding to the sequence 1 [SEQ ID NO:1] possessing biological activity and eliciting enhancing effect on resistance of capillaries. Also, invention proposes pharmaceutical composition enhancing resistance of capillaries and containing effective amount of peptide lysyl-glutamyl-aspartic acid of the general formula: H-Lys-Glu-Asp-OH of the sequence 1 [SEQ ID NO:1] as an active component and a pharmaceutically acceptable carrier. Pharmaceutical composition is prepared in form suitable for parenteral administration. Also, invention proposes a method for prophylaxis and/or treatment of microcirculation disorders in organs and tissues and involves administration in patient pharmaceutical composition containing effective amount of peptide lysyl-glutamyl-aspartic acid of the general formula: H-Lys-Glu-Asp-OH of the sequence 1 [SEQ ID NO:1] as an active component in the dose 0.01-100 mcg/kg of body mass for at least once per a day for period necessary for providing the therapeutic effect. Administration is carried out by parenteral route. Proposed group of inventions provides preparing a novel peptide possessing biological activity eliciting in enhancing resistance of capillaries and using this peptide for preparing pharmaceutical composition enhancing resistance of capillaries.

EFFECT: enhanced and valuable medicinal properties of peptide and pharmaceutical composition.

7 cl, 3 tbl, 2 dwg, 6 ex

FIELD: peptides, medicine, hepatology, pharmacy.

SUBSTANCE: groups of inventions relates to medicinal agents used in treatment of liver diseases. Invention proposes a pharmaceutical composition stimulating regeneration of liver tissue and comprising the effective amount of peptide glutamyl-aspartyl-leucine as an active component of the general formula: H-Glu-Asp-Leu-OH of the sequence 1 [SEQ ID NO:1] and a pharmaceutically acceptable carrier. Invention proposes peptide glutamyl-aspartyl-leucine of the general formula: H-Glu-Asp-Leu-OH of the sequence 1 [SEQ ID NO;1] stimulating regeneration of liver tissue. Also, invention proposes a method for stimulating liver tissue involving administration in a patient a pharmaceutical composition containing peptide glutamyl-aspartyl-leucine of the general formula: H-Glu-Asp-Leu-OH of the sequence 1 [SEQ ID NO:1] as an active component used in the dose 0.01-100 mcg/kg of body mass for at least once per a day for period required for appearance of the therapeutic effect. Invention provides the development of peptide possessing the biological activity eliciting in stimulating regeneration of liver tissue, and pharmaceutical composition containing this peptide as an active component. Using this composition stimulates regeneration of liver tissue based on recovery of synthesis of tissue-specific proteins and normalization of functions of liver cells.

EFFECT: valuable properties of peptide and pharmaceutical composition.

7 cl, 5 tbl, 1 dwg, 6 ex

FIELD: medicine, pharmacy.

SUBSTANCE: invention relates to drugs used in prophylaxis and treatment of the locomotor system, in particular, degenerative-dystrophic joint and backbone diseases. Invention proposes a pharmaceutical composition normalizing metabolism in osseous and cartilage tissues and comprising the effective amount of peptide alanyl-glutamyl-aspartic acid of the general formula: H-Ala-Glu-Asp-OH of the sequence 1 [SEQ ID NO:1] as an active component, and pharmaceutically acceptable carrier. Invention proposes peptide alanyl-glutamyl-aspartic acid of the general formula: H-Ala-Glu-Asp-OH of the sequence 1 [SEQ ID NO:1] possessing the biological activity manifesting as normalization of metabolism in osseous and cartilage tissues. Invention proposes a method for prophylaxis and treatment of locomotor system by normalization of metabolism in osseous and cartilage tissues involving administration in a patient of a pharmaceutical composition containing as an active component peptide alanyl-glutamyl-aspartic acid of the general formula: H-Ala-Glu-Asp-OH of the sequence 1 [SEQ ID NO:1] in the dose 0.01-100 mcg/kg of the body mass for at least once per a day for time necessary for achievement of the therapeutic effect. Invention can be used as agent normalizing metabolism in osseous and cartilage tissues.

EFFECT: valuable medicinal properties and high effectiveness of peptide and pharmaceutical composition.

6 cl, 2 tbl, 1 dwg, 1 ex

FIELD: organic chemistry, medicine.

SUBSTANCE: invention represents ligands MC-4 and/or MC-3 of the formula (I): , wherein X means hydrogen atom, -OR1, -NR1R1' and -CHR1R1' wherein R1 and R1' are taken among the group: hydrogen atom, (C1-C6)-alkyl and acyl; (1) each R2 is taken independently among the group: hydrogen atom, (C1-C6)-alkyl; or (2) (a) R2 bound with carbon atom that is bound with X and Z1 and substitute R5 can be optionally bound to form carbocyclic or heterocyclic ring that is condensed with phenyl ring J; or (b) R2 bound with carbon atom that is bound with ring Ar can be bound with R7 to form ring condensed with ring Ar; each among Z1, Z2 and Z3 is taken independently from the following groups: -N(R3e)C(R3)(R3a)-, -C(R3)(R3a)N(R3e)-, -C(O)N(R3d)-, -N(R3d)C(O)-, -C(R3)(R3a)C(R3b)(R3c)-, -SO2N(R3d)- and -N(R3d)SO2- wherein each among R3, R3a, R3b and R3c, R3d, R3e when presents is taken independently among hydrogen atom and (C1-C6)-alkyl; p is a whole number from 0 to 5 wherein when p above 0 then R4 and R4' are taken among hydrogen atom, (C1-C6)-alkyl and aryl; R5 represents 5 substitutes in phenyl ring J wherein each R5 is taken among hydrogen atom, hydroxy-, halogen atom, thiol, -OR12, -N(R12)(R12'), (C1-C6)-alkyl, nitro-, aryl wherein R12 and R12' are taken among hydrogen atom and (C1-C6)-alkyl; or two substitutes R5 can be bound optionally to form carbocyclic or heterocyclic ring that is condensed with phenyl ring J; q = 0, 1, 2, 3, 4 or 5 wherein when q above 0 then R6 and R6' are taken among hydrogen atom and (C1-C6)-alkyl; Ar is taken among the group consisting of phenyl, thiophene, furan, oxazole, thiazole, pyrrole and pyridine; R7 are substitutes at ring Ar wherein each R7 is taken among hydrogen, halogen atom, -NR13R13', (C1-C6)-alkyl and nitro- wherein R13 and R13' are taken among hydrogen atom and (C1-C6)-alkyl; r is a whole number from 0 to 7 wherein when r is above 0 then R8 and R8' are taken among hydrogen atom and (C1-C6)-alkyl; B is taken among -N(R14)C(=NR15)NR16R17, -NR20R21, heteroaryl ring and heterocycloalkyl ring wherein R14-R17, R20 and R21 are taken independently among hydrogen atom and (C1-C6)-alkyl; s = 0, 1, 2, 3, 4 or 5 wherein when s is above 0 then R and R9' are taken among hydrogen atom and (C1-C6)-alkyl; R10 is taken among the group consisting of optionally substituted bicyclic aryl ring and optionally substituted bicyclic heteroaryl ring; D is taken among hydrogen atom, amino- and -C(O)R11 wherein R11 is taken among the following group: hydroxy-, alkoxy-, amino-, alkylamino-, -N(R19)CH2C(O)NH2 wherein R19 represents (C1-C6)-alkyl, -NHCH2CH2OH and -N(CH3)CH2CH2OH, or its isomers, salts, hydrates or biohydrolysable ester, amide or imide.

EFFECT: valuable medicinal properties of compounds.

18 cl, 107 ex

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