CH645092A5 - MERCAPTOACYL PEPTIDES AND MEDICINAL PRODUCTS CONTAINING THEM. - Google Patents

Description

The invention relates to the subject matter characterized in the claims.

The term alkyl radical means radicals with 1 to 7 carbon atoms. The term alkanoyl radical means radicals having 2 to 7 carbon atoms. The acetyl group is preferred.

The a-amino acid and a-imino acid residues Aj and A2 can be either natural acids or synthetic acids. These residues can be derived, for example, from proline, 4-hydroxyproline, 4,4-ethylenedioxyproline, 4-methoxyproline, 4-thiazolidinecarboxylic acid, tryptophan, glycine, alanine, leucine, isoleucine and valine. The residues Aj and A2 are separated by a peptide bond -C-NH- between the a-

O

Carboxyl group of the radical A1 and the a-amino or a-imino group of the radical A2 connected.

The mercaptoacyl dipeptides of the general formula I, in which R 1 is an alkanoyl radical or a benzoyl group, can be obtained by acylation of a dipeptide of the general formula II

A, -A2 (II)

or its alkyl ester with a thioic acid of the general formula III

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R £ s (CH2) n CH — j: OH (III)

O

getting produced. The radical R'i denotes an alkanoyl radical or a benzoyl group. The acylation is carried out according to methods known per se, for example in the presence of a coupling agent such as a carbodiimide. Dicyclohexylcarbodiimide is the most used. Alternatively, the thioic acid of the general formula III can also be activated by formation of the mixed anhydride, a symmetrical anhydride, an acid chloride or an active ester or by using Woodward reagent K or N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline. A detailed discussion of the different acylation methods is e.g. B. in Methods of Organic Chemistry (Houben-Weyl), Vol. XV, Part II (1974), p. Lff. described.

An alternative synthesis for the preparation of the compounds of general formula I, in which Rx represents an alkanoyl radical or a benzoyl group, consists in the successive acylation, i.e. in the acylation of the amino acid Aj with a mercaptoalkanoyl acid of the general formula III by the process described above to form a mercaptoalkanoyl amino acid of the general formula IV

Î2

R £ S (CH2Jn CH 1 A1 (IV *

and the subsequent acylation of the amino acid A2 or its alkyl ester with the mercaptoalkanoylamino acid of the general formula IV. The acylation can be carried out by customary methods for coupling amino acids; see. Bodanszky and Ondetti, Peptide Synthesis, Intersciene Publishers (1966).

Compounds of the general formula I in which Rj is a hydrogen atom can be prepared by ammonolysis or alkaline hydrolysis of the corresponding compounds of the general formula I in which Rt represents an alkanoyl radical or a benzoyl group. Compounds of the general formula I in which Rt is a group of the general formula g

-s-ccH2) n-M-j-Al-A2

O

represents can be prepared by oxidation of the corresponding free mercaptans of the general formula I with iodine.

Alternative methods for the preparation of the compounds listed above are still available. For example, when using an ester of the dipeptide of the general formula II

Aj — A2 (II)

or an amino acid A2, the corresponding free acid can be obtained from the ester by acidic or alkaline hydrolysis.

Mercaptoalkanoyl acids of the general formula III and mercaptoalkanoyl amino acids of the general formula IV and processes for their preparation are described, for example, in US Pat. Nos. 4,046,889.4,105,776 and 4,053,651.

The compounds of general formula I according to the invention form salts with various inorganic and

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organic bases. Examples of these salts are ammonium salts, alkali metal salts, such as sodium and potassium salts, which are preferred, alkaline earth metal salts, such as calcium and magnesium salts, salts with organic bases, such as dicyclohexylamine, benzathine, N-methyl-D-glucamine and hydrodabamine , as well as salts with amino acids, arginine and lysine. Non-toxic, physiologically acceptable salts are preferred, but other salts can also be used, for example for isolating or purifying the products.

The compounds of general formula I according to the invention, their alkyl esters and salts inhibit the conversion of the angiotensin converting enzyme (ACE) and thus the conversion of the decapeptide angiotensin I to angiotensin II. They can therefore be used in the treatment of high pressure.

One of the blood pressure regulating (blood pressure increasing) systems in the human organism is the renin-angioten-sin-aldosterone system. The enzyme renin is produced in the myoepithelial cells of the juxtaglomerular apparatus of the kidney and released from there. The mechanism that regulates renin secretion appears to be primarily controlled by the salt and water content of the blood and the pressure at which blood is pumped through the kidney. If these parameters decrease, renin is released directly into the blood. There it encounters the a2-globulin angiotensinogen - an inactive protein body - as its substrate. The renin cleaves angiotensinogen in two likewise inactive components, a tetrapeptide and the decapeptide angiotensin I (A I). When circulating through the capillaries (especially the lungs), AI encounters the angiotensin converting enzyme (ACE), which is located in the endothelial cells of the capillaries. ACE cleaves the AI into a dipeptide and the octapeptide angiotensin II (A II), the strongest known blood pressure-increasing substance in the human organism. The blood pressure-increasing effect of A II takes place both locally in the vascular muscles and centrally, since the area postrema in the brain, after stimulation by A II, transmits blood pressure-increasing impulses to the organism. A II also has a second effect. It is the physiological stimulus for the secretion of the mineral corticoid aldosterone from the adrenal cortex. Aldosterone increases the reabsorption of sodium ions from the kidney tubules, which increases the effective blood volume and thus increases blood pressure via a second mechanism.

In addition to converting AI to A II, the ACE has another effect. It breaks down the vasodilating octapeptide bradykinin present in the blood into inactive components. In this role, ACE is also known as Kininase II.

In the hypertensive, the renin-angiotensin-aldosteron system is disturbed and contributes significantly to the high pressure of the patient. So far, there has been no drug that attacks the ACE to achieve the following effects, namely

1. Inhibition of the formation of A II and thereby a) direct inhibition of vasoconstriction and b) inhibition of the release of aldosterone from the adrenal cortex (inhibition of excessive water and salt absorption) and

2. by inhibiting the ACE inhibiting the cleavage of bradykinin, causing its vasodilation

(hypotensive) effect is retained.

The compounds of the general formula I are strong inhibitors of ACE. They inhibit the conversion of the decapeptide angiotensin I to angiotensin II and therefore reduce or eliminate the one caused by angiotensin II

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Hypertension. The compounds according to the invention intervene in the angiotensinogen (renin) - * angiotensin I- * angiotensin II reaction sequence by blocking the ACE and thereby reducing or eliminating the formation of the blood pressure-increasing compound angiotensin II.

The compounds of general formula I are preferably given orally, but they can also be administered parenterally, e.g. Subcutaneously, intramuscularly, intravenously or intraperitoneally in the form of conventional medicinal products.

The examples illustrate the invention.

example 1

l- [N- (acetylthioacetyl) glycyl] -L-proline

1.72 g of glycyl-L-proline are dissolved in 10 ml of sodium hydroxide solution while stirring in an ice bath. This solution is mixed with 5 ml of 2N sodium hydroxide solution and then with 1.13 g of chloroacetyl chloride. The ice bath is then removed. After standing at room temperature for 3 hours, 836 mg of mercaptoacetic acid and 960 mg of potassium carbonate in 10 ml of water are added, and the mixture is stirred at room temperature for about 16 hours. The reaction mixture is then acidified with a sulfonated polystyrene cation exchange resin, eluted with water and the eluate is evaporated to dryness under reduced pressure. The residue is taken up in acetic acid and insoluble substances {F. above 320 ° C) are filtered off. The filtrate is evaporated to dryness under reduced pressure and a 7: 3 mixture of chloroform and acetic acid is added to the residue. The mixture is centrifuged and the supernatant is added to a column filled with 60 g of silica gel and poured out with a 7: 3 mixture Chloroform and acetic acid eluted. The eluate is evaporated. 1.0 g of the title compound are obtained.

Example 2

1 - [N- (Mercaptoacetyl) Glycyl] -L-Proline

1.0 g of l- [N- (acetylthioacetyl) glycyl] -L-proline is treated for 30 minutes under argon with a solution of 6 ml of water and 6 ml of concentrated sodium hydroxide solution. The solution obtained is then evaporated under reduced pressure, the residue is acidified with a sulfonated polystyrene cation exchange resin, placed on a column filled with the same cation exchange resin and eluted with water. The eluate is evaporated. 800 mg of substance are obtained. 800 mg and 200 mg from a previous batch are chromatographed on a column filled with 30 g of silica gel with a 7: 3 mixture of chloroform and acetic acid. 740 mg of the product obtained are placed on a column filled with 85 ml of diethylaminoethyl dextran anion exchange resin and eluted with a linear gradient of 0.005 to 0.5M ammonium bicarbonate solution (750 ml each). The product in the eluate is converted into the free acid with a sulfonated polystyrene cation exchange resin and then stirred under argon as a protective gas with 7 ml of acetic acid and 350 mg of zinc dust for 6 hours. The suspension is then centrifuged and the supernatant is evaporated to dryness under reduced pressure. The residue is taken up in water and freeze-dried. The product obtained is acidified with a sulfonated polystyrene cation exchange resin, placed on a column filled with the same cation exchange resin and eluted with water. The eluate is evaporated. 365 mg of the title compound are obtained.

C9H14N204S (0.3) H20;

calc .: C 42.95 H 5.68 N 11.13 S 12.72 found: C 42.92 H 5.78 N 10.94 S 12.68 Sh calc.: 1.00; found: 0.98

Example 3

(S) -1- [3- (acetylthio) -2-methyl-1-oxopropyl] -L-prolyl-L-proIin tert-butyl ester A solution of 7.8 g of l- [S-3- (acetylthio ) -2-methyl-l-oxo-propyl] -L-proline in 75 ml of dichloromethane is mixed with 5.13 g of the tert-butyl ester of L-proline. The solution obtained is cooled to 5 ° C. and a solution of dicyclohexylcarbodiimide in 50 ml of dichloromethane is added dropwise. The reaction is carried out at room temperature for about 16 hours. The precipitate formed is then filtered off and the filtrate is washed successively with water, 5 percent sodium bicarbonate solution, water, 10 percent potassium bisulfate solution and water. The solution is then dried over sodium sulfate. The solvent is then distilled off. 12 g of crude product are obtained. After recrystallization from a mixture of ethyl acetate and pentane, the title compound of mp 104 to 106 ° C is obtained.

Example 4

l- (S-3-Mercapto-2-methyl-l-oxopropyl) -L-prolyl-

L-prolin

12 g of the tert-butyl ester of l- [S-3- (acetylthio) -2-methyl-l-oxopropyl] -L-proline are dissolved in 60 ml of distilled trifluoroacetic acid. The solution is left to stand at room temperature for 1 hour. The trifluoroacetic acid is then distilled off under reduced pressure and the residue is dried under high vacuum for 24 hours. The oil obtained is then dissolved in 30 ml of methanol and 20 ml of concentrated aqueous ammonia solution are added. The mixture is left to stand at room temperature for 1 hour. The solvent is then distilled off under reduced pressure and the oily residue is dried and mixed with 6 ml of dicyclohexylamine. The reaction mixture is then mixed with diethyl ether. An oil separates and is taken up in ethyl acetate. The ethyl acetate solution is washed with 10% aqueous potassium bisulfate solution and then dried over sodium sulfate. The solvent is then distilled off, the residue is taken up in methanol and the solution is adsorbed on silica gel. The solvent is distilled off and the product adsorbed on silica gel is placed on a column into which silica gel has been slurried with chloroform. The column is eluted with chloroform and the product-containing fractions are pooled. The solvent is distilled off and the residue is freeze-dried from water. 1.05 g of the title compound are obtained as a hygroscopic solid.

C14H22N204S • 1H20;

calc .: C 51.99 H 7.17 N 8.66 S 9.92 found: C 52.12 H 7.44 N 8.18 S 9.37 neutralization equivalent calc .: 323; found: 341.

Example 5

N- (Acetylthioacetyl) -L-valyl-L-proline A solution of 3.1 g of tert-butyloxycarbonyl-L-valyl-L-proline in 20 ml of trifluoroacetic acid is left to stand for 15 minutes at room temperature. The solvent is then distilled off under reduced pressure and the oily residue is dried under reduced pressure for several hours. The oil is then dissolved in 25 ml of dimethylformamide. 2.2 g of the N-hydroxy-succinimide ester of acetylmercaptoacetic acid are added to the solution obtained. The reaction is carried out for 17 hours at a pH of 7.5 (adjusted with triethylamine) at room temperature. Thereafter, the solvent is distilled off, the residue is taken up in ethyl acetate and the solution obtained in succession with water, 10% aqueous

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Potassium bisulfate solution, water, saturated aqueous sodium bicarbonate solution and water and finally dried over sodium sulfate. After evaporation, 4.8 g of the title compound are obtained as an oil.

Example 6 N- (Mercaptoacetyl) -L-valyl-L-proline A solution of 4.8 g of N- (acetylthioacetyl) -L-valyl-L-proline in 12 ml of methanol and 7.5 ml of concentrated aqueous ammonia solution is used for 1 hour left at room temperature. The pH is then adjusted to 7 and the methanol is distilled off under reduced pressure. The remaining aqueous phase is acidified to a pH of 2 with 2N hydrochloric acid and the solution is saturated with sodium chloride. The product is then extracted with ethyl acetate. The ethyl acetate extract is dried over sodium sulfate and the solvent is distilled off under reduced pressure. 2 g of crude product with a melting point of 158 to 162 ° C. are obtained. After repeated recrystallization from ethyl acetate with the addition of a small amount of methanol, the title compound of F. 162 to 165 ° C is obtained. After drying for 2 hours at 55 ° C, a sample melts at 190 to 192 ° C.

Example 7

1 - {S-3- (acetylthio) -2-methyl-1-oxopropyl] -L-prolyl-

glycine ethyl ester Example 3 is repeated, but 10.4 g of l- [S-3- (acetylthio) -2-methyl-l-oxopropyl] -L-proline, 100 ml of di-chloromethane, 5.6 g of glycine ethyl ester (prepared from 5.58 g of glycine ethyl ester hydrochloride and 5.6 ml of triethylamine in 50 ml of dichloromethane) and 8.24 g of dicyclohexylcarbodiimide were used. 11.2 g of the title compound are obtained as a viscous oil.

Example 8

(S) -l- (3-mercapto-2-methyl-l-oxopropyl) -L-prolyl-

glycine lithium salt A solution of 11.2 g of l- [S-3- (acetylthio) -2-methyl-l-oxopropyl] -L-prolylglycine ethyl ester in 200 ml of methanol is mixed with a solution of 9 g of potassium hydroxide in 40 ml Water added. The hydrolysis is carried out at room temperature for 2 hours. The organic solvent is then distilled off under reduced pressure and the aqueous residue is adjusted to a pH of 7.0. The product is extracted with ethyl acetate. The ethyl acetate extract is dried over sodium sulfate and evaporated under reduced pressure. The residue (3.5 g) is taken up in water and the pH is adjusted to 2.0. After the addition of sodium chloride, the free acid is extracted with ethyl acetate. The ethyl acetate extract is evaporated and freeze-dried from water. 2.16 g of the free base of the title compound are obtained. The entire product is taken up in water and the solution is adjusted to 7.0 with 0.05N lithium hydroxide solution. The solution is then freeze-dried. 1050 mg of the title compound are obtained as a hygroscopic solid.

CnH17N204S • Li;

calc .: C 47.14 H 6.12 S 11.44 Li 2.48 found: C 46.56 H 6.42 S 11.37 Li 2.43

Example 9

(S) -1 - (3-mercapto-2-methyl-1-oxopropyl) -2-L-prolyl-L-alanine Examples 3 and 4 are repeated, but 7.8 g of l- [S-3- (Acetylthio) -2-methyl-l-oxopropyl] -L-proline, 75 ml of dimethylformamide, 6.2 g of dicyclohexylcarbodiimide,

4.2 g of L-alanine methyl ester hydrochloride and 4.2 ml of triethylamine (pH 7.5 to 8.0) were used. 8 g of product are obtained. This product is placed on a column filled with silica gel, which is eluted with ethyl acetate. Two fractions are obtained with an Rf value of 0.76 and 0.4. 2 g of the less mobile fraction with an Rf value of 0.4 are dissolved in a solution of 1.5 g of potassium hydroxide in 7 ml of water and 35 ml of methanol. The solution obtained is left to stand at room temperature for 2 hours. The organic solvent is then distilled off under reduced pressure and the aqueous residue is acidified to a pH of 2.0. After adding sodium chloride, the solution is extracted 5 times with ethyl acetate. The ethyl acetate extract is dried and the solvent is distilled off under reduced pressure. The solid residue obtained is recrystallized from hot ethyl acetate. 1 g of the title compound of melting point 128 to 130 ° C. are obtained.

Example 10

Acetylthioacetyl valyl tryptophan methyl ester

A solution of 15 g of methyl ester of tert-butyloxycar-bonyl-valyl-tryptophan in 150 ml of trifluoroacetic acid and 15 ml of anisole is stirred for 1 hour under nitrogen as a protective gas. The solvent is then distilled off under reduced pressure. An oily residue remains.

A solution of 4.83 g of acetylthioacetic acid and 6.18 g of hydroxybenzotriazole monohydrate in 120 ml of tetrahydrofuran is cooled to 0 ° C. and a solution of 8.16 g of dicyclohexylcarbodiimide in 60 ml of tetrahydrofuran is added. The mixture is stirred at 0 ° C. for 30 minutes and at room temperature for a further hour. The precipitated urea compound is then filtered off and the filtrate is evaporated under reduced pressure. A white solid remains. The active ester obtained is dissolved in 90 ml of dimethylformamide and a solution of the dipeptide methyl ester trifluoroacetate obtained in 60 ml of dimethylformamide and a sufficient amount of N-methylmorpholine are added to adjust the pH to 7.5. The mixture obtained is stirred for 16 hours at room temperature. The reaction mixture is then evaporated under reduced pressure. The oily residue is shaken between ethyl acetate and water. The ethyl acetate phase is washed with 2N citric acid, water, saturated aqueous sodium bicarbonate solution and water and dried over sodium sulfate. The solution is then evaporated. The solid residue obtained is recrystallized from a mixture of ethyl acetate and pentane. 10 g of the title compound of melting point 110 to 112 ° C. are obtained.

Example 11

N- (Mercaptoacetyl) -L-valyl-L-tryptophan lithium salt

A solution of 5.0 g of acetylthioacetyl-valyl-tryptophan-methyl ester in 52 ml of methanol is mixed with a solution of 2.1 g of potassium hydroxide in 10.5 ml of water. The mixture is stirred for 90 minutes under nitrogen as a protective gas at room temperature. The methanol is then distilled off under reduced pressure. The aqueous residue is diluted with water, extracted with ethyl acetate and acidified to pH 2.0 with 6N hydrochloric acid. The solid obtained is filtered off, washed with water and dried. 3.4 g of product are obtained. This product is chromatographed on a column filled with Sephadex LH-20 and with a 7: 3 mixture of methanol and water. 1.5 g of an oil are obtained. The oil is dissolved in a 1: 1 mixture of ethanol and water and adjusted to a pH with 0.1N lithium hydroxide solution.

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Value set to 7.0. A small amount of insoluble substances is filtered off. The filtrate is evaporated under reduced pressure. The oily residue is dissolved in water and freeze-dried. 1.5 g of a hygroscopic solid are obtained.

C18H22N3O4S • Li;

calc .: C 51.54 H 6.25 S 7.65 Li 1.65 found: C 51.94 H 6.03 S 7.28 Li 1.62 neutralization equivalent calculated: 419; found: 413.

Example 12

(S) -1 - [3- (acetylthio) -2-methyl-1-oxopropyl] -L-prolyl-D-alanine methyl ester A solution of 7.8 g of I- [S-3- (acetylthio) -2 -methyl-l-oxopropyl] -L-proline in 150 ml of acetonitrile is added at 0 ° C with 4.9 g of carbonyldiimidazole. The mixture is then stirred at this temperature for 1 hour. Then the almost clear solution is mixed with 4.2 g of D-alanine methyl ester hydrochloride. The pH is adjusted to 7.5 to 8.0 with 4.2 ml of triethylamine and the mixture is stirred at room temperature for 17 hours. The precipitate formed and the solvent are then separated off and the oily residue obtained is taken up in ethyl acetate. The solution is washed successively with water, 0.1N hydrochloric acid, water, saturated aqueous sodium bicarbonate solution and water and dried. The solvent is then distilled off. The oily residue shows a spot with an Rf value of 0.5 and a trace of a spot with an Rf value of 0.75 (in ethyl acetate) on the thin layer chromatogram on silica gel with a 9: 1 mixture of ethyl acetate and chloroform as the eluent ). The yield of the title compound is 4 g.

Example 13

(S) -l- (3-mercapto-2-methyl-l-oxopropyl) -L-prolyI-

D-alanine

A solution of 4 g of (S) - [3- (acetylthio) -2-methyl-I-oxopropyl] -L-prolyl-D-alanine methyl ester in a solution of 3 g of potassium hydroxide in 14 ml of water and 70 ml of methanol is used Allow to stand for 2 hours at room temperature. The product is then isolated according to Example 9. Yield 2.1 g of the title compound of mp 152 to 155 ° C. C12H20N2O4S;

calc .: C 49.98 H 6.99 S 11.12 found: C 50.02 H 7.24 S 10.90 neutralization equivalent calculated: 288; found: 284.

Example 14 l- [N- (Mercaptoacetyl) -L-alanylI-L-proline A solution of 1.3 g of acetylthioacetic acid and 1.44 g of hydroxybenzotriazole in 39 ml of tetrahydrofuran is mixed with 2 g of dicyclohexylcarbodiimide. The mixture is stirred at 0 ° C. for 30 minutes and at room temperature for a further hour. The precipitate formed is then filtered off and L-alanyl-L-proline trifluoroacetate (prepared from 2.8 g of tert-butyloxycarbonyl-L-alanyl-L-proline and trifluoroacetic acid) is added to the filtrate. The pH is adjusted to 7.5 to 8.0 with N-methylmorpholine and the reaction is carried out for 16 hours at room temperature. The solvents are then distilled off under reduced pressure and the residue is dissolved in a mixture of 18 ml of methanol and 18 ml of concentrated aqueous ammonia solution. After 30 minutes the mixture is evaporated under reduced pressure. The residue is dissolved in water and placed on a column filled with a sulfonated polystyrene cation exchange resin and eluted with water. The eluate is evaporated. The product is then purified by chromatography on silica gel with a

7: 3 mixture of acetic acid and chloroform as eluent further cleaned. The eluate is also evaporated and the residue is recrystallized from ethyl acetate. 850 mg of the title compound of melting point 152 to 155 ° C. are obtained. [<x] d = -128 ° (c = 1.5; 50% aqueous methanol).

Example 15

1- [N- (Mercaptoacetyl) -L-alanyl] -L-thiazolidine-4-carboxylic acid Example 14 is repeated, but the L-alanyl-L-thiazolidine-4-carboxylic acid trifluoroacetate is used. The title link will be obtained.

Example 16

1- [N- (Mercaptoacetyl) -L-alanyl] -4-methoxy-L-proline

Example 14 is repeated, but the L-alanyl-4-methoxy-L-proline trifluoroacetate is used. The title link will be obtained.

Example 17

1- [N- (Mercaptoacetyl-L-alanyl] -4-hydroxy-L-proline Example 14 is repeated, but the L-alanyl-4-hydroxy-L-proline trifluoroacetate is used. The title compound is obtained.

Example 18

l- [N- (Mercaptoacetyl) -L-alanyl] -4,4-ethylenedioxy-L-proIin

Example 14 is repeated, but L-alanyl-4,4-ethylenedioxy-L-proline is used. The title link will be received.

Example 19

1 - [N- (2-MercaptopropanoyI) -L-alanyl] -L-proline Example 14 is repeated, but 2-acetylthio-propionic acid is used. The title compound is obtained as a waxy solid with a melting point of (47 ° C.) 54 to 65 ° C. (dec.).

CnH18N204S • '/ zHjO;

calc .: C 46.63 H 6.76 S 9.89 found: C 46.50 H 6.58 S 9.78 Sh titration: 100%.

Example 20

1 - [N- (3-Mercapto-l-oxopropyl-L-alanylJ-L-prolin Example 14 is repeated, but 3-acetylthio-propionic acid is used. The title compound is used as a hygroscopic solid of F. (61 ° C) Obtain 75 to 84 ° C.

C1iH18N204-0.75H20;

calc .: C 45.89 H 6.83 S 11.14 found: C 45.75 H 6.62 S 10.98 Sh titration: 99.6%.

Example 21

1- [N- (2-Mercaptopropanoyl) -L-alanyl] -4-hydroxy-L-proline

Example 14 is repeated, but 2-acetythio-propionic acid and L-alanyl-4-hydroxy-L-proIin-trifluoracetate are used. The title link will be obtained.

Example 22

l- [N- (2-mercaptopropanoyl) -L-alanyl] -L-thiazolidine

4-carboxylic acid Example 14 is repeated, but 2-acetylthio-propionic acid and L-alanyl-L-thiazolidine-4-carboxylic acid trifluoroacetate are used. The title link will be obtained.

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Example 23

1- [N- (2-Mercaptopropanoyl) -L-ananyl] -4-methoxy-L-proline

Example 14 is repeated, but 2-acetylthio-propionic acid and L-alanyl-4-methoxy-L-proline trifluoroacetate are used. The title link will be obtained.

Example 24

l- [N- (2-mercaptopropanoyl) -L-alanyl] -4,4-ethylene-

dioxy-L-prolin Example 14 is repeated, but 2-acetylthio-propionic acid and L-alanyl-4,4-ethylenedioxy-L-prolin are used. The title link will be obtained.

Example 25

l- [N- (2-Mercapto-3-phenylpropanoyl) -L-alanyl] -4-hydroxy-L-proline Example 14 is repeated, but 2-acetylthio-3-phenylpropionic acid and L-alanyl-4-hydroxy- L-proIin trifluoroacetate used. The title link will be obtained.

Example 26

1- [N- (2-Mercapto-3-phenylpropanoyl) -L-alanyl] -L-thiazolidine-4-carboxylic acid Example 14 is repeated, but using 2-acetylthio-3-phenylpropionic acid and L-alanyl-L-thiazolidine 4-carbon acid trifluoroacetate used. The title link will be obtained.

Example 27

l- [N- (2-Mercapto-3-phenylpropanoyl) -L-alanyl] -4-methoxy-L-proline Example 14 is repeated, but 2-acetylthio-3-phenylpropionic acid and L-alanyl-4-methoxy- L-prolin trifluoroacetate is used. The title link will be obtained.

Example 28

I- [N- (2-mercapto-3-propanoyl) -L-alanyl] -4,4-ethylene-

dioxy-L-prolin Example 14 is repeated, but 2-acetylthio-3-phenylpropionic acid and L-alanyl-4,4-ethylenedioxy-L-prolin are used. The title link will be obtained.

Example 29 l- [N- (Mercaptoacetyl) -L-prolil] -L-prolin Method A:

3 g of benzyloxycarbonyl-L-prolyl-L-prolin are hydrogenated in a mixture of 50 ml of anhydrous ethanol and 7.5 ml in hydrochloric acid in the presence of a 10% palladium-on-carbon catalyst. After 5 hours, the catalyst is filtered off and the solvent is distilled off under reduced pressure. The residue is dissolved in a mixture of 15 ml of dimethylformamide and 1.05 ml of triethylamine. The solution is mixed with 1.7 g of the acetyl-thioacetic acid ester of N-hydroxysuccinimide. After standing at room temperature for about 16 hours, the solvents are distilled off under reduced pressure and the residue is chromatographed on a silica gel column using a 7: 1 mixture of benzene and hexane. The eluate is evaporated. The residue is dissolved in 5 ml of trifluoroacetic acid and 2 ml of anisole and left to stand for 1 hour at room temperature. The reaction mixture is then evaporated to dryness and the residue is dissolved in 7.7 ml of sodium hydroxide solution under argon as a protective gas and with stirring. After 15 minutes the mixture is neutralized with a sulfonated polystyrene cation exchange resin, placed on a column of the same cation exchange resin and eluted with water. The eluate is evaporated and the residue is recrystallized from ethyl acetate. The title compound of F. 182 to 183 ° C is obtained.

Method B:

A solution of 1.7 g of L-proline tert-butyl ester and 1.5 g of hydroxybenzotriazole in 15 ml of dichloromethane, cooled in an ice bath, is mixed with 2.06 g of dicyclohexylcarbodiimide and 2.3 g of l- (acetylthioacetyl) -L-proline transferred. After stirring for about 16 hours at 5 ° C., the precipitate formed is filtered off and the filtrate is washed until neutral. The organic solution is evaporated to dryness and the residue is chromatographed on silica gel. The eluate is evaporated and the residue is recrystallized from a mixture of diethyl ether and hexane. A product with a melting point of 95 to 98 ° C. is obtained. This material is freed from the protective group by successive treatment with trifluoroacetic acid and sodium hydroxide solution according to method A.

Example 30

Acetylthioacetyl-L-alanyl-L-tryptophan methyl ester

A solution of 2.5 g of L-alanyl-L-tryptophan-methyl-ester acetate in 20 ml of dimethylformamide is mixed with 2.0 g of acetylthioacetic acid ester of N-hydroxysuccinimide. The pH of the solution is adjusted to 7.5 with N-methylmorpholine and the mixture is stirred at room temperature for about 16 hours. The solvent is then distilled off under reduced pressure and the oily residue is taken up in ethyl acetate, washed with water, dried and evaporated under reduced pressure. There remain 3 g of a foamy residue. This residue is taken up in ethyl acetate and placed on a column filled with 60 g of basic aluminum oxide (activity I; Woelm) and measuring 1.5 × 45 cm. It is eluted with ethyl acetate. The eluate is evaporated. 1.6 g of a foamy solid are obtained which, on thin-layer chromatography on silica gel with a 3: 1 mixture of chloroform and methanol, gives a single spot with an Rf value of 0.69.

Example 31

N- [N- (Mercaptoacetyl) -L-alanyl} -L-tryptophan lithium salt

A solution of 1.2 g of the compound obtained in Example 30 in 36 ml of methanol is stirred under nitrogen as a protective gas for 30 minutes at room temperature with 6 ml of sodium hydroxide solution. The reaction mixture is then adjusted to a pH of 6 and the solvent is distilled off under reduced pressure. The residue is shaken between a 1: 1 mixture of ethyl acetate and saturated aqueous sodium bicarbonate solution. The aqueous phase is washed with ethyl acetate, saturated with brine, acidified to pH 2.0 and extracted with ethyl acetate. The ethyl acetate extract is dried over sodium sulfate and evaporated under reduced pressure. A frothy residue is obtained. This residue is dissolved in a 50 percent aqueous ethanol solution, and the solution is adjusted to pH 7 with ln-lithium hydroxide solution. The solution is then evaporated to dryness and the residue obtained is taken up in water and freeze-dried. 390 mg of an off-white solid are obtained.

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C16Hi8N304S • Li • 2H20;

calc .: C 49.11 H 5.67 N 10.73 S 8.18

Found: C 98.78 H 5.07 N 10.41 S 7.85

Example 32

(S) -l- (3-mercapto-2-methyl-l-oxopropyl) -L-prolyl-B-alanine

A) A solution of 5.2 g l- [S-3- (acetylthio) -2-methyl-l-oxopropyl] -L-proline and 3.24 g carbonyldiimidazole is at 0 ° C with 3.06 g B- Alanine ethyl ester hydrochloride and

2.82 ml of triethylamine are added. The resulting mixture is stirred at room temperature for about 16 hours. The reaction mixture is then evaporated and the residue extracted with ethyl acetate. The ethyl acetate extract is washed successively with water, aqueous sodium bicarbonate solution saturated in hydrochloric acid, water and water and dried over sodium sulfate. The solution is then evaporated. 3.4 g of (S) -l- (3-acetylthio-

2-methyl-l-oxopropyl) -L-prolyl-B-alanine as a clear oil. This connection is used in the next stage.

B) 3.4 g of the product from stage A) are dissolved in a solution of 43 ml of methanol and 1.7 g of potassium hydroxide

8.6 ml of water dissolved. The mixture is stirred at room temperature for 1 hour, then 50 ml of water are added and the methanol is distilled off. The aqueous residue is adjusted to a pH of 2.0 with dilute hydrochloric acid. The solvent is then distilled off under reduced pressure and the residue is extracted with methanol. After drying, the methanol is distilled off. 3.2 g of an oil remain, which is chromatographed on a column filled with Sephadex LH-20 and with a 1: 9 mixture of methanol and water. 1.1 g of the title compound are obtained as a clear oil. In the thin-layer chromatogram on silica gel and with methanol as the eluent, a spot appears with an Rf value of 0.65.

C12H20N2O4S;

calc .: C 49.99 H 6.99 N 9.71 found: C 49.81 H 7.29 N 8.67

Example 33

(R) -l- [N- (2-mercapto-l-oxo-3-phenylpropyl) -L-alanyl] -L-prolin A) (R) -2-acetylthio-3-phenyl-propionic acid

A solution of 51 g of L-phenylalanine and 125 g of potassium bromide in 620 ml of 25N sulfuric acid, cooled to 0 ° C., is mixed with 32.75 g of sodium nitrite with stirring within 1 hour. The mixture is then stirred for a further hour at 0 ° C. and then for 1 hour at room temperature. The reaction mixture is then extracted with diethyl ether. The ether extract is dried over sodium sulfate and evaporated. 44.9 g of oily (S) -2-bromo-

3-phenylpropionic acid, which distill at 142 to 144 ° C / 0.25 Torr.

A mixture of 8.7 ml of thioacetic acid and 6.8 g of potassium hydroxide in 225 ml of acetonitrile is stirred at room temperature for 1 hour and 15 minutes. The mixture is then cooled in an ice bath and a solution of 25.3 g of (S) -2-bromo-3-phenylpropionic acid in 25 ml of acetonitrile is added within 10 minutes. The mixture is then stirred for 5 hours. The reaction mixture is then filtered off and the solvent is distilled off under reduced pressure. The oily residue is taken up in ethyl acetate and dried. The solvent is distilled off. 24 g of crude (R) -2-acetylthio-3-phenyl-propionic acid are obtained. The crude product is purified over the di-cyclohexylamine salt. Diethyl ether is used for »recrystallization. The free acid is regenerated from the salt with hydrochloric acid and extracted with ethyl acetate. The

Ethyl acetate extract is evaporated. [cc] d25 = + 62 ° (c = 2.8; chloroform).

B) R- (2-acetylthio-3-phenyl) propionyl-L-alanyl-L-proline The title compound is prepared from the compound obtained in step (A) by reaction with L-alanyl-L-proline according to Example 14. F. (124 ° C) 135 to 138 ° C;

[a] o2S = - 23 ° (c = 1.4; chloroform).

C) R-l- [N- (2-mercapto-l-oxo-2-phenylpropyl)] - L-

alanyl-L-prolin The product obtained in stage (B) is reacted further according to Example 14. Sodium hydroxide solution is used instead of aqueous ammonia solution. The title compound is obtained as a foamy solid;

[a] D25 = - 78.4 ° (c = 1.1; chloroform). C17H22N204S-2H20;

calc .: C 52.83 H 6.78 N 7.25 S 8.30 found: C 52.54 H 6.26 N 7.01 S 8.16

Example 34

(S) -l- [N- (2-Mercapto-l-oxo-3-phenylpropyl)] - L-alanyl-L-proline A) (S) -2-acetylthio-3-phenylpropionic acid Example 33 (A) repeated, but D-phenylalanine, mp 146 to 147 ° C, from ethyl acetate as the dicyclohexylamine salt is used. The free acid is regenerated with hydrochloric acid and extracted with ethyl acetate. [<x] d25 = -70, I ° (c = 1.91; chloroform)

B) (S) - (2-acetylthio-3-phenyl) -propionyI-L-alanyl-L-proline

The product from stage (A) is reacted with L-alanyl-L-proline according to Example 33 (B). The title compound is obtained as an oily residue, which solidifies when digested with diethyl ether and hexane. Mp 161-163 ° C; [a] D25 = -109.6 ° (c = 1.5; chloroform).

C) (S) -l- [N- (2-mercapto-l-oxo-3-phenylpropyI)] - L-alanyl-L-proline

The title compound is obtained as a foamy solid by reacting the product obtained in step (B) according to Example 33 (C);

[ab25 = -82.7 ° (c = 1.5; chloroform).

C17H22N204S • H2O;

calc .: C 55.42 H 6.56 N 7.60 S 8.70 found: C 55.25 H 6.18 N 7.36 S 8.43 Sh titration: 99.1%.

Example 35

N- [N- [2- (mercaptomethyl) -l-oxo-3-phenylpropyl] glycyl] -L-arginine

A) N- [2- [acetylthio) methyl] -1-oxo-3-phenylpropyl] glycine

13 g (0.067 mol) of benzylmalonic acid are mixed with 7.6 g (0.068 mol) of 40 percent aqueous dimethylamine and 5.4 g (0.068 mol) of 37 percent aqueous formaldehyde solution in 150 ml of water. A voluminous solid forms within 15 minutes. This solid is filtered off after 2 hours, washed with water and partially air-dried. 20.8 g of a product are obtained; theoretically 16.8 g can be expected. The solid is melted in an oil bath heated to 170 ° C. and heated for 10 minutes until the amine development and the bubble formation stop / after which the product is cooled. The mobile liquid obtained is acidified with 10% aqueous potassium bisulfate solution and extracted with hexane. The hexane extract is dried over sodium sulfate and evaporated. 6.3 g of a solid are obtained. The aqueous filtrates from the Mannich reaction are left to stand for about 16 hours and then heated on a steam bath to 100 ° C. until the bubbles stop. This takes about 2 hours. After cooling, An8

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Acidifying and extracting a further 1.2 g of solid, ie a total of 7.5 g (75% of theory) of benzyl acrylic acid, are obtained.

6.2 g (0.04 mol) of benzyl acrylic acid in 350 ml of dichloromethane are mixed with 5.4 g (0.04 mol) of glycine ethyl ester hydrochloride and 3.9 g (0.04 mol) of triethylamine. The mixture is cooled in ice and 7.9 g (0.04 mol) of dicyclohexylcarbodiimide are added. The mixture obtained is stirred at 25 ° C. for about 16 hours and then filtered. The filtrate is shaken out with aqueous sodium bicarbonate solution, 10% aqueous potassium bisulfate solution and water and then dried over sodium sulfate. The solution is then evaporated. The oily residue obtained is taken up in 200 ml of methanol and excess 10% sodium hydroxide solution is added. The mixture is heated on a steam bath for 15 minutes and then evaporated to an aqueous slurry. This slurry is filtered off and the filtrate extracted with diethyl ether. The aqueous phase is acidified with 10% hydrochloric acid and extracted with chloroform. The chloroform extract is dried over sodium sulfate and evaporated. 7.3 g (80% of theory) of an oil are obtained.

3.2 g (0.014 mol) of the glycine derivative are mixed with 10 ml of thiolacetic acid, and the solution obtained is evaporated under reduced pressure after about 18 to 20 hours. The residue is digested with diisopropyl ether. Almost white crystals are obtained in a yield of 2.9 g. After recrystallization from a mixture of ethyl acetate and hexane, 2.5 g (60% of theory) of the title compound of melting point 97 to 101 ° C. are obtained.

CI4H17N04S;

calc .: C 56.93 H 5.80 N 4.74 S 10.85 found: C 56.68 H 5.87 N 4.77 S 10.81 B) N- [N- [2- (acetylthio ) -methyl] -l-oxo-3-phenylpropyl] -glycyl-L-arginine

0.3 g (1 mmol) of the product from step (A) are dissolved in 25 ml of anhydrous tetrahydrofuran, 0.11 g (1.1 mmol) of triethylamine are added and the mixture is cooled to 0.degree.

Then 0.11 g (1 mmol) of ethyl chloroformate are added, the mixture is stirred for 40 minutes at 0 ° C. and then filtered into a solution of 0.17 g (1 mmol) of arginine in 10 ml of water. After stirring for about 16 hours at 25 ° C, the mixture is evaporated. The aqueous residue obtained is diluted to 30 ml with water and carefully extracted with ethyl acetate in order to avoid emulsion formation. The aqueous phase is then chromatographed on 50 g of Avicel in water. Fractions of 25 ml each are collected. The two Sakaguchi positive fractions are pooled, filtered through a Nucleopo-re polycarbonate filter and freeze-dried. 0.3 g (63% of theory) of a white powder with a melting point of 105 to 120 ° C. are obtained. After drying for 6 hours under reduced pressure at 95 to 100 ° C, the analysis fits the presence of 1.5 moles of water.

C20H29NsOsS • 1.5H20;

calc .: C 50.19 H 6.74 N 14.63 S 6.70 found: C 50.38 H 6.56 N 14.95 S 6.50 20 C) N- [N- [2- ( Mercaptomethyl) -l-oxo-3-phenylpropyl] -glycyl] -L-arginine 0.5 g (1.1 mmol) of the compound obtained in step (B) - are cooled in ice and concentrated under argon as a protective gas with 4 ml aqueous ammonia solution added. 25 After stirring for 1 hour, the sample is evaporated under reduced pressure. The glassy residue is digested with 25 ml of acetonitrile for 2 hours. The solvent is then decanted, replaced with fresh solvent and stirred for a further 2 lA days under argon as a protective gas. The white solid obtained is filtered off and washed with acetonitrile and diethyl ether. Finally, the product is dried at 45 ° C over potassium hydroxide, phosphorus pentoxide and paraffin at a pressure of 0.01 Torr for about 16 hours. 0.25 g (55% of theory) of the title 35 compound of melting point 120 to 137 ° C. are obtained.

C18H27N504S • 1.75H20;

calc .: C 49.02 H 6.97 N 15.88 S 7.27 found: C 48.89 H 6.67 N 16.17 S 7.28

s

Claims (36)

645 092 PATENT CLAIMS 1. Mercaptoacyl peptides of the general formula I Î2 Rj- S- (CH2) n-M-C-Ai-A2, (I) o in the Ri is a hydrogen atom, an alkanoyl radical, a benzoyl group or a group of the general formula R2 -S- (CH2) n ~ CH - ^ - Ai-A2 O R2 represents a hydrogen atom, an alkyl or phenylalkyl radical, n has the value 0 or 1 and A1 and A2 each represent an a-amino acid or a-imino acid residue which are linked to one another by a peptide bond, and their alkyl esters and Salts, where the alkyl groups contain 1 to 7 carbon atoms and the alkanoyl groups contain 2 to 7 carbon atoms. , 2nd . -S- (CH2.) N-CH.-C-Al-A2 O represents. 2. Compounds according to claim 1 of the general formula I, in which Rt represents a hydrogen atom. 3. Compounds according to claim 1 of the general formula I, in which Rj represents an alkanoyl radical. 4. Compounds according to claim 1 of the general formula I, in which Rj is a group of the general formula 5. Compounds according to claim 1 of the general formula I, in which R2 represents a hydrogen atom. 6. Compounds according to claim 1 of the general formula I, in which R2 represents an alkyl radical. 7. Compounds according to claim 1 of the general formula I, in which R2 represents a phenylalkyl radical. 8. Compounds according to claim 1 of the general formula I, in which n has the value 0. 9. Compounds according to claim 1 of the general formula I, in which n has the value 1. 10. Compounds according to claim 1 of the general formula I, in which Aj represents an a-amino acid residue and A2 represents an a-imino acid residue. 11. Compounds according to claim 10, characterized in that A2 means the rest of the L-proline. 12. Compounds according to claim I of the general formula I, in which Ai and A2, which are the same or different, the proline, 4-hydroxypropylene, 4,4-ethylenedioxyproline, 4-methoxyproline, 4-thiazolidinecarboxylic acid, tryptophan -, Glycine, alanine, leucine, isoleucine or valine residue mean. 13. l- [N- (Acetylthioacetyl) -gylcyl] -L-proline as a compound according to claim 1. 14. l- [N- (mercaptoacetyl) glycyl] -L-proline as a compound according to claim 1. 15. (S) -1 - [3- (acetylthio) -2-methyl-1-oxopropyl] -L-prolyl-L-proline tert-butyl ester as a compound according to claim 1. 16.1 - (S-3-Mercapto-2-methyl-1-oxopropyl) -L-proIyl-L-proline as a compound according to claim 1. 17. N- (acetylthioacetyl) -L-valyl-L-prolin as a compound according to claim 1. 18. N- (Mercaptoacetyl) -L-valyl-L-prolin as a compound according to claim 1. 19. l- [S-3- (acetylthio) -2-methyl-l-oxopropyl] -L-prolyl-glycine ethyl ester as a compound according to claim 1. 20. l- (S-3-Mercapto-2-methyl-l-oxopropyl) -L-prolylglycine lithium salt as a compound according to claim 1. 21. (S) -l- (3-mercapto-2-methyl-l-oxopropyl) -L-prolyl-L-alanine as a compound according to claim 1. 22. Acetylthioacetyl-valyl tryptophan methyl ester as a compound according to claim 1. 23. N- (Mercaptoacetyl) -L-valyl-L-tryptophan lithium salt as a compound according to claim 1. 24. (S) - [3- (acetylthio) -2-methyl-l-oxopropyl] -L-prolyl-D-alanine methyl ester as a compound according to claim 1. 25. (S) - (3-Mercapto-2-methyl-l-oxopropyl) -L-propyl-D-alanine as a compound according to claim 1. 26. l- [N- (mercaptoacetyl) -L-alanyl] -L-proline as a compound according to claim 1. 27.1 - [N- (3-Mercapto-1-oxopropyl) -L-alanyl] -L-proline as a compound according to claim 1. 28. Acetylthioacetyl-L-alanyl-L-tryptophan as a compound according to claim I. 29. N- [N- (mercaptoacetyl)] - L-alanyl-L-tryptophan as a compound according to claim 1. 30. (S) -l- (3-acetylthio-2-methyl-1-oxo-propyl) -L-prolyl-B-alanine as a compound according to claim 1. 31. (S) -1 - (3-mercapto-2-methyl-1-oxopropyl) -L-prolyl-B-alanine as a compound according to claim 1. 32. R- (2-acetylthio-3-phenyl) propionyl-L-alanyl-L-proline as a compound according to claim 1. 33. R-1 - [N- (2-mercapto-1-oxo-3-phenyl-propyl)] - L-alanyl-L-proline as a compound according to claim 1. 34. (S) - (2-acetylthio-3-phenyl) -propionyl-L-alanyl-L-proline as a compound according to claim 1. 35. (S) -l- [N- (2-mercapto-l-oxo-3-phenyl-propyl)] - L-alanyl-L-proline as a compound according to claim 1. 36. Medicament, characterized by a content of a compound according to claim 1.