NO853947L - ANALOGY PROCEDURE FOR THE PREPARATION OF THERAPEUTIC ACTIVE E AMINO ACID DERIVATIVES. - Google Patents

Description

IN

This invention relates to a process for the production of new amino acid derivatives with the general formula

and salts thereof.

In formula I means:

R H, acetyl, benzoyl, pivaloyl or t-butoxycarbonyl;

R. H, acetyl, benzoyl, pivaloyl or the rest

on one of the numbers 1-4; A the rest of one of the a-amino acids or

R2hydrogen, C1-C4-alkyl, linear or branched, C5-C7-cycloalkyl on which a phenyl ring may be fused, phenyl, phenyl-C3-C4-alkyl, a hetero-C3-C4-alkyl residue where the heterocyclic group consists of a 5- or 6-ring with 1-2

of the heteroatoms 0, S or N,

R3hydrogen, -C4-alkyl, linear or branched, phenyl, phenyl-C1-C4-alkyl or a hetero-C1-C4-alkyl radical, where the heterocyclic group consists of a 5- or 6-ring with 1-2

of the heteroatoms 0, S or N, or

R2 and R3 together form with the N and C atom a 5-, 6- or 7-membered ring, which may be saturated or contain a double bond, or a 4-, 5- or 6-membered ring, which contains a one or two additional heteroatoms from 0, S

or N;

R 4 OH, C 1 -C 4 -hydroxyalkyl, C 1 -C 4 - alkoxy, phenyl -C 4 -C 4 -alkyl, C 1 -C 4 -alkylamino, (C 4 -C 4 ) 2 -dialkylamino, one of the groups

or an α-amino acid which is peptide-bonded to the C0 group of the molecule;

R,. C 1 -C 4 -alkyl, C 1 -C 4 -alkenyl, -C 4 -alkynyl, linear or branched, hydroxy, nitro, amino, -C 4 -alkyl, mercapto, C 1 -C 4 -alkylthio, hydroxy-C 1 -C 4 -alkyl , mercapto-C1-C4-alkyl, F, Cl, Br, amino-C1-C4~alkyl, sulfonamido, methylenedioxy, fluoro-C1-C4~alkyl, chloro-C1-C4-alkyl, bromo-C1-C4-alkyl , cyano

or trifluoromethyl;

R 8 is hydrogen or methyl;

Ry C 1 -C 4 -alkyl, linear or branched, where the alkyl residue may

be substituted with F, Cl, Br, CF 3 , phenyl or pyridyl; Q, Y and Z 0, S, NRCRg, CHRg,

with the proviso that only one of the residues X, Y and Z can mean 0, S,

and one or two of the residues X, Y and Z may be NRg;

R 8 is hydrogen or C 1 -C 4 alkyl, linear or branched;

Rg hydrogen, or together with a neighboring residue Rg a phenyl ring or, for m and n = 1, its dihydro form with the double bond

in conjugation to the C-terminal carboxy group, and

m and n each 0, 1 or 2, the sum of m and n being 1 or 2.

If the radicals R2 and/or R3 mean a 5- or 6-membered hetero-alkyl radical, examples of the heterocyclic ring can be mentioned thiophene, furan, pyrrole, thiazole, oxazole, isoxazole, pyrazole, imidazole, pyran, thiopyran, pyridine, morpholine, pyrazine, pyrimidine, pyridazine, oxathiine and their di- and tetrahydroforms.

The optionally substituted alkyl groups mentioned as meaning the residue R can be linear or branched. Proline, valine, leucine, isoleucine and phenylalanine come into particular consideration as α-amino acids, which are bound by a peptide bond to the C0 group of the molecule.

At one of the meanings for A, the 5- or 6-ring heterocyclic groups which are condensed on pyrrolidine or the piperidine carboxylic acid, be saturated or unsaturated. Preferred heterocyclic rings are furan, pyrrole, thiophene, benzofuran, indole, benzothiophene, oxazole, imidazole, thiazole, isoxazole, pyrazole, pyrrolidine, tetrahydrofuran, tetrahydrothiophene, pyridine, pyridazine, quinoline, isoquinoline or piperidine.

The new compounds usually exhibit several centers of asymmetry and thus exist as diastereomers or in the form of racemates or racemic mixtures. The invention encompasses both the preparation of the racemic mixtures and the individual diastereomers. The enantiomers where the asymmetric C atoms are present in S configuration are preferred.

The mentioned racemates can be enriched in their sterically uniform forms or obtained in pure form in the usual way, for example by fractional crystallization, by chemical or by biochemical cleavage.

The new compounds are available in the form of their betaines. They are thus able to form salts both with acids and with bases. Examples of inorganic bases are ammonia, alkali metal hydroxides such as sodium or potassium hydroxide, or alkaline earth metal hydroxides such as calcium or magnesium hydroxide. N,N'-dibenzylethylenediamine or triethylamine can be mentioned as organic bases. Examples of acids suitable for salt formation include halogenated hydrogen acids, sulfuric acid or sulphonic acids.

The new compounds with the general formula I can be prepared by various methods:

a) By condensation of an acid with the general formula

where

has the meaning given above, with a dipeptide of the general formula

where

A and p have the meaning given above, and X means a suitable protecting group, for example a t-butyloxy-carbonyl-, a,a-dimethyl-3,5-dimethoxy-oxycarbonyl(Ddz)-, adamantyloxycarbonyl(Adoc)-, 2-(p-biphenylyl)-isopropyloxycarbonyl(Bpoc), t-amyloxycarbonyl(Aoc), furfuryloxycarbonyl(Foc) or p-methoxybenzyloxycarbonyl-(Z(omet)) group.

b) Condensation of a compound with the general formula

where

R1rX and p have the above-mentioned meaning, with an amino acid A where A has the above-mentioned meaning.

The compound of formula IIa can optionally be obtained by a peptide connection of a compound of formula II with a compound of the general formula

c) Reaction of a compound with the general formula

where

X, p and A have the meaning given above, with a mercapto compound of the general formula

HS - R 1 (VI) where R 2 has the meaning indicated above.

The compound of formula (V) can be prepared by condensation of 3-phenyl-2-bromopropionic acid with a dipeptide of the general formula III.

d) Condensation of a compound with the general formula

where

X and p have the above meaning, with an amino acid A, where A has the above meaning, and subsequent reaction of the thus obtained condensation product of the general formula V with a mercapto compound of the general formula VI. The compound of formula IVa can be prepared by peptide coupling of 3-phenyl-2-bromo-propionic acid with a compound of the general formula IV.

e) A compound of formula I where means acetyl, benzoyl or pivaloyl, can be converted into the corresponding compound

with formula I where R^ means hydrogen,

f) a compound of formula I where R^ is acetyl, benzoyl or pivaloyl and R is H, can be converted into a compound with

formula I where R^ is hydrogen and R is acetyl, benzoyl or pivaloyl,

and any protective groups present can be cleaved at a suitable step of the methods a), b), c),

d), e) or f).

The condensations described above occur in normal conditions

methods, such as those described in Houben-Weyl, Volume XV/1, 4th edition, 1974 for the synthesis of peptides.

In general, the coupling reactions common in peptide chemistry are used using an activated form of the acids II, Ila, IV and IVa; as carboxyl-activating groups, for example, acid chlorides, azides, anhydrides, p-nitrophenyl esters, trichlorophenyl esters, thiophenyl esters, o-cyanomethyl esters, etc. can be used. As coupling agents, preferably carbonyldiimidazole, dicyclohexylcarbodiimide, ethoxyacetylene, diphenylphosphoryl azide or chloroformic acid esters are used, as the reaction takes place at temperatures between 0 and +10°C.

In the present case, the use of methylene chloride as a solvent together with cyclohexylcarbodiimide and triethylamine has proven to be advantageous.

Reaction of the bromine intermediates with a mercapto compound of the general formula VI conveniently takes place in ether as solvent. In the reaction with thiol acids (for example thiolacetic acid) the corresponding acyl-mercapto compounds are obtained, and the acetyl group can be cleaved off with strong bases.

The conversion process f) can be carried out under standard trans-acylation conditions, for example by treating the compound of formula I with a strong base followed by neutralization. The reaction can be carried out at room temperature in aqueous solution.

Dimers of the general formula I

you get, for example

a) by oxidation of a compound of the general formula I where R1 means hydrogen and the NHR group is protected with one of

the groups indicated above for X, with an oxidizing agent such as oxygen or iodine;

by reacting a compound of the general formula IVa with thiolacetic acid to a compound IIa and then saponification to a compound with a free mercapto group. The oxidation to the corresponding dimer takes place as described under a); the intermediate product obtained is condensed as described under method b), with an amino acid A.

Preference is given to such compounds of the general formula I where R and R₁ mean hydrogen, p = 4 and A means proline or thiazolidine.

According to the methods described above, the following end products can be prepared, possibly in the form of their salts: N-[Na<->(3-phenyl-2-S-acetyImercaptopropanoyl)-S-lysyl]-S-proline N-[ Na-(3-phenyl-2-S-mercaptopropanoyl)-S-lysyl]-S-proline N-[Na-(3-phenyl-2-R-mercaptopropanoyl)-S-lysyl]-S-proline

N-[N<a>.(3-phenyl-2-S-mercaptopropanoyl)-S-lysyl]-thiazolidine-4-S-carboxylic acid

N-[Na<->(3-phenyl-2-S-acetylmercaptopropanoyl)-S-lysyl]-thiazolidine-4-S-carboxylic acid

N-[Na<->(3-phenyl-2-S-acetylmercapto)-S-lysyl]S-proline pivaloyl oxymethyl ester

N-[Na<->(3-phenyl-2-S-acetylmercapto)-S-lysyl]-thiazolidine-4-S-carboxylic acid pivaloyloxymethyl ester

N,N'-[N,N'-{2,2'-S-dithiobis-(3-phenylpropanoyl)}-bis-S-lysyl]-bis-5-proline

N,N'-[N,N'-{2,2'-S-dithiobis-(3-phenylpropanoyl)}-bis-S-lysyl]-bis-S-proline pivaloyloxymethyl ester

N,N-[N,N'-{2,2'-S-dithiobis-(3-phenylpropanoyl)}-bis-S-lysyl]-bis-thiazolidine-4-S-carboxylic acid

N,N-[N,N'-{2,2'-S-dithiobis-(3-phenylpropanoyl)}-bis-S-lysyl]-bis-thiazolidine-4-S-carboxylic acid pivaloyloxy methyl ester

The starting materials are mainly known compounds, but possibly new starting materials can also be prepared by methods known per se. Compounds with the general formula III are obtained, for example, by peptide linking of a compound with the formula

with an amino acid A.

The amino acid derivatives of the general formula I have a strong, long-lasting blood pressure-lowering effect. This is due to an inhibition of the angiotensin I converting enzyme and thus a blocking of the formation of the vasoconstrictor angiotensin II from angiotensin I. In addition, the new compounds inhibit the enzyme kinase II responsible for bradykinin degradation, which can be considered identical to the above converting enzyme. As bradykinin has a vasodilating effect, the blood pressure-lowering effect is enhanced by means of this additional effect. The blood pressure lowering achieved by bradykinin in normal rats is enhanced by the new compounds. The blood pressure-lowering effect obtained in non-pretreated genetic hypertensive rats may also serve as an expression of this effect.

Thus N-[Na<->(3-phenyl-2-S-mercaptopropanoyl)-S-lysyl]-thiazolidine-4-S-carboxylic acid in anesthetized rats in a dosage of 0.3 mg/kg i.v. a long-term blood pressure reduction of 60% when pre-elevated by an i.v. administration of 0.1 mg/kg angiotensin I; for N-[Na<->(3-phenyl-2-S-mercapto-propanoyl)-S-lysyl]-S-proline the value in the same test is 57%.

N-[Na<->(3-phenyl-2-S-mercaptopropanoyl)-S-lysyl]-thiazolidine-4-S-carboxylic acid shows an inhibitory activity on the angiotensin I converting enzyme in vitro IC5Q=4.6 • 10 [M]; IC5 Qutguør for N-[Na<->(3-phenyl-2-S-mercaptopropanoyl)-S-lysyl]-S-proline 2.7 • 10~<8>[M].

The concentration of inhibitory substance which inhibits the angiotensin I converting enzyme by 50% is denoted here as ICC„ (pp. H.S. Cheung, D.W. Cushman, Biochem. Biophys. Acta 293, 451 (1973)).

For use in therapy, the new compounds are mixed with common pharmaceutical fillers or carriers, stretching, breaking, binding, sliding, thickening or diluting agents. For example, pharmaceutical preparation forms include tablets, capsules, suppositories, solutions, juices, emulsions or dispersible powders, where other known active substances such as saluretics, diuretics and/or anti-hypertonics may be added.

Suitable tablets can, for example, be prepared by mixing the active substance or substances with known excipients, for example inert diluents such as calcium carbonate, calcium phosphate or milk sugar, disintegrants such as corn starch or alginic acid, binders such as starch or gelatins, lubricants such as magnesium stearate or talc, and / or agents to achieve a depot effect, such as carboxypolymethylene, carboxymethyl cellulose, cellulose acetate phthalate or polyvinyl acetate.

The tablets can also consist of several layers.

Similarly, dragees can be produced by coating cores produced analogously to the tablets with agents that are usually used in dragee coatings, for example collidon or shellac, gum arabic, talc, titanium dioxide or sugar. To achieve a depository effect or to avoid incompatibilities, the core can also consist of several layers. Likewise, the dragée coating can also consist of several layers to achieve a depot effect, as the excipients mentioned above for the tablets can be used.

Juices of the new active substances or combinations of active substances may also contain a sweetener such as saccharin, cyclamate, glycerol or sugar, as well as a taste-enhancing agent, for example flavoring substances such as vanillin or orange extract. They may also contain suspending aids or thickeners such as sodium carboxymethyl cellulose, wetting agents such as condensation products of fatty alcohols with ethylene oxide, or protective substances such as p-hydroxybenzoates.

Injection solutions are prepared in the usual way, for example by adding preservatives such as p-hydroxybenzoates, or stabilizers such as alkali salts of ethylene-diamine-tetraacetic acid and adding suitable solvents and filled in injection bottles or ampoules.

Capsules containing one or more active substances or combinations of active substances can, for example, be prepared by mixing the active substances with inert carriers such as milk sugar or sorbitol and encapsulating them in gelatin capsules.

The daily dose for use of the new compounds of the general formula I or their salts can be 5-500, preferably 10-100 mg and can be administered in 1-4 single doses.

The following examples serve to illustrate the invention in more detail. The structure of the compounds obtained in all synthesis examples was confirmed by NMR spectra.

Example 1

N-[ N~-(3- phenyl- 2- S- acetvlmercaptopropanovl)- S- lvsvl3- S- proline

10.7 g (26.7 mmol) N-(Ne<->tert.butoxycarbonyl-S-lysyl)-S-proline-tert.butyl ester and 6.11 g (26.7 mmol) 3-phenyl-2- R-bromopropionic acid is dissolved in 270 ml of anhydrous dichloromethane and 5.5 g (26.7 mmol) of N,N'-dicyclohexylcarbodiimide are added at 0°C with stirring. Stirring continues overnight at room temperature, the precipitated urea derivative is filtered off with suction and the filtrate is evaporated in vacuo. The residue is dissolved in ethyl acetate and filtered from undissolved components. The organic phase is washed successively with 5 µg KHSO 4 solution, NaHCO 4 solution and saturated NaCl solution and dried over MgSO 4 . After distilling off the ethyl acetate in vacuum, an oily residue of 14 g (= 85.8% of the theoretical) remains, which NMR spectroscopically determined to have the structure of N-[Na<->(3-phenyl-2-R -bromopropanyl)-Ne<->tert-butoxycarbonyl-S-lysyl]-S-proline tert-butyl ester.

For the acetyl mercapto replacement, 3.48 g (45.8 mmol) of thiolacetic acid are dissolved in 120 ml of anhydrous ether and 4.62 g (45.8 mmol) of triethylamine and then 14 g (22.9 mmol) of the bromine compound prepared above are added dropwise at 0°C. Boil for a further 90 minutes under reflux and let the whole thing stand overnight at room temperature. Suction is carried out from the precipitated triethylamine hydrobromide, the filtrate is distilled in vacuum, and 15.5 g of a brown oil is obtained as a residue.

The chromatographic purification over silica gel (eluent: ethyl acetate, n-hexane = 1:1) gave 9.4 g (= 67.7% of theory) of N-[Na<->(3-phenyl-2-S- acetyl-mercaptopropanoyl)-Ne<->tert-butoxycarbonyl-S-lysyl]-S-proline-tert-butyl ester.

For saponification of the tertiary butyl ester and simultaneous cleavage of the Ne<->tert.butoxycarbonyl protecting group, 5.5 g (9.08 mmol) of the ester prepared above are dissolved in 150 ml of trifluoroacetic acid and stirred overnight at room temperature. After distilling off the trifluoroacetic acid in a vacuum, the residue is redissolved in dichloromethane and evaporated. The residue is dissolved in NaHCO 3 solution and extracted with ethyl acetate. The aqueous phase is acidified with concentrated hydrochloric acid and extracted twice with dichloromethane. The combined dichloromethane extracts are washed with water and saturated NaCl solution, dried and distilled off in vacuum. Residue 0.5 g. The aqueous phase is saturated with NaCl and extracted again with dichloromethane. After drying over MgSO 4 and evaporation, a further 3 g, thus a total of 3.5 g, are obtained as solidified foam, which, by NMR spectroscopy, is shown to have the structure of the title compound.

Rf = 0.22 (butanol, acetic acid, water = 3:1:1, silica gel).

Example 2

N-[ N~-( 3- phenvl- 2- S- mercaptopropanovl)- N~- acetvl- S- lvsvl]- S- proline

Dissolve 4.2 g (9.3 mmol) of N-[Na<->(3-phenyl-2-S-acetylmercaptopropanoyl)-S-lysyl]-S-proline in 95 ml of water, add 8.2 ml of 5 N caustic soda and stirred for 1.5 hours at room temperature and ^-atmosphere. A further 4.1 ml of 5 N caustic soda is then added and stirring is continued for a further 1.5 hours. The alkaline solution

shaken with ethyl acetate, acidified with concentrated hydrochloric acid and then extracted with dichloromethane. The organic phase is washed with saturated NaCl solution, dried and distilled off in a vacuum. 2.1 g (= 55.2% of the theoretical) of the title compound are obtained as a white, solidified foam.

Rf = 0.49 (butanol, acetic acid, water = 3:1:1, Kieselgel 60F254Merck).

Example 3

N-[ N~-( 3- phenyl- 2- S- mercaptopropanoyl)- S- lvsvl]- thiazolidine- 4- S- carboxylic acid

3.5 g (7.6 mmol) Na<->(3-phenyl-2-R-bromopropanoyl)-Ne<->tert.butoxycarbonyl-S-lysine and 1.4 g (7.6 mmol) thiazolidine- 4-S-carboxylic acid methyl ester hydrochloride is dissolved in 70 ml of anhydrous dichloromethane, and while stirring and cooling to 0°C, 769 mg (7.6 mmol) of triethylamine are added, then 1.6 g (7.6 mmol) of N,N '-dicyclohexylcarbodiimide. Stir at room temperature for approx. 17 hours, aspirate off the separated dicyclohexylurea and distill off the solvent. The oily residue is dissolved in ethyl acetate, after cooling it is filtered again, and the organic phase is washed successively with dilute KHSO^ solution, saturated NaHCO^ solution, water and saturated NaCl solution. After drying over MgSO 4 , the solvent is distilled off in vacuum.

4.1 g of the crude, oily reaction product is purified chromatographically over silica gel with ethyl acetate/n-hexane = 1:1 as eluent, thereby obtaining 2.9 g (= 68.8% of the theoretical) N-[Na<- >(3-phenyl-2-R-bromopropanoyl)-Ne<->tert-butoxycarbonyl-S-lysyl]-thiazolidine-4-S-carboxylic acid methyl ester as a solidified foam which crystallized from ether.

Rf = 0.43 (ethyl acetate, n-hexane = 2:1, silica gel).

To replace bromine with an acetylmercapto group, a solution of 792 mg (10.4 mmol) of thiolacetic acid in 35 ml of anhydrous ether is slowly added under nitrogen and ice cooling to 1.05 g (10.4 mmol) of triethylamine and then added dropwise with stirring to a solution of 2.9 g (5.2 mmol) of the above bromine compound in 20 ml of ethyl acetate. It is then boiled for a further 60 minutes under reflux, the previously cooled solution is filtered and washed successively with dilute KHSO 4 solution, NaHCO 4 solution, water and saturated NaCl solution. After drying over MgSO 4 , the solvent is distilled off in vacuo, and 3.3 g of a yellowish oil is obtained as a residue. The chromatographic purification takes place over 100 g of silica gel with ethyl acetate/n-hexane = 2:1 as eluent. One thus obtains 1.9 g (64.1% of the theoretical) of pure N-[Na<->(3-phenyl-2-S-acetylmercaptopropanoyl)-Ne<->tert.butoxycarbonyl-S-lysyl]-thiazolidine -4-S-carboxylic acid methyl ester as solidified foam.

Rf = 0.36 (ethyl acetate, n-hexane = 2:1, silica gel).

To remove the tert.butoxycarbonyl protecting group, 1.9 g (3.33 mmol) of the above-mentioned acetylmercapto compound is stirred with 40 ml of trifluoroacetic acid for 3 hours at room temperature, then evaporated in vacuo to dryness, and to remove residual amounts of trifluoroacetic acid, dissolved three times in acetone and in chloroform, and the evaporation is carried out again. 2.7 g of N-[Na<->(3-phenyl-2-S-acetylImercaptopropanoyl)-S-lysyl]-thiazolidine-4-S-carboxylic acid methyl ester are obtained in the form of a viscous oil.

R, = 0.23 (chloroform, methanol, glacial acetic acid = 90:10:5, silica gel).

For simultaneous saponification of the methyl and thiolacetic acid ester, 2.7 g (3.3 mmol) of the above reaction product are dissolved in 20 ml of methanol and 40 ml of water and 4 ml of 5 N caustic soda is added. After stirring for 1.5 hours, 2.5 ml of 5 N caustic soda is added at room temperature and further stirring is carried out for 1.5 hours. The methanol is distilled off in a vacuum, the residue is added to water and decanted with dichloromethane.

The aqueous phase is acidified with dilute hydrochloric acid, the separated viscous oil is separated in a rinsing funnel and dissolved in a little water and purified over 100 g "Dowex 50 W x 4" (elution with 5% aqueous pyridine).

700 mg (= 45.9% of the theoretical) N-[Na<->(3-phenyl-2-S-mercapto-propanoyl)-S-lysyl]-thiazolidine-4-S-carboxylic acid is obtained in NMR spectroscopic pure form.

Rf = 0.52 (ethyl acetate, butanol, water, acetic acid = 1:1:1:1, silica gel).

Preparation of the starting compounds: N a-(3-phenyl-2-R-bromopropanoyl)-N~- tert. butoxycarbonyl-5-lvsine 3.25 g (14.2 mmol) 3-phenyl-2-R-bromopropionic acid and 3.7 g (14.2 mmol) Ne<->tert.butoxycarbonyl-S-lysine methyl ester are dissolved in 70 ml of anhydrous dichloromethane, while stirring and cooling (0°C), 2.9 g (14.2 mmol) of N,N'-dicyclohexylcarbodiimide are added, and the whole is stirred further overnight at room temperature. The separated dicyclohexylurea is filtered off with suction, the solvent is removed by distillation and the residue is taken up in ethyl acetate. After cooling and re-filtration, the organic phase is washed in sequence with dilute KHSO^ solution, saturated NaHCO^ solution, water and saturated NaCl solution, dried over MgSO^ and evaporated to dryness in vacuo. The crystalline residue (6.2 g) is recrystallized from ethyl acetate and n-hexane, thereby obtaining 4.8 g (= 71.1% of theory) of the methyl ester of the title compound; m.p. 97-98°C.

R^= 0.49 (ethyl acetate, n-hexane = 2:1, silica gel),

R^ = 0.7 (chloroform, methanol, glacial acetic acid = 90:10:5, silica gel).

For saponification of the above-mentioned methyl ester, 4.8 g (10.2 mmol) Na<->(3-phenyl-2-R-bromopropanoyl)-Ne<->butoxycarbonyl-S-lysine methyl ester are dissolved in 50 ml of methanol and added 11 ml of 1 N caustic soda (11 mmol) with stirring and ice water cooling. After stirring for 3 hours at room temperature, the methanol is driven off in vacuo, the aqueous residue is diluted with water, and the cloudy solution is acidified with a 5% KHSO 4 solution and extracted with dichloromethane. The crystalline product which separates from the dichloromethane solution is suctioned off (2.32 g), and a further 2 g of crystalline mass is obtained from the filtrate. Both fractions are collected and recrystallized from chloroform and a little methanol. 3.1 g (= 66.4% of the theoretical) of the title compound is obtained. Temp. 178-179°C (decomp.).

Rf = 0.55 (chloroform, methanol, glacial acetic acid = 90:10:5, silica gel).

Example 4

N-[ Na-(3- phenyl- 2- S- acetyl mercaptopropanoyl)- S- lvsvl]- thiazolidine-4- S- carboxylic acid

The S-acyl derivative of the title compound from example 3 is obtained when, as described in example 3, instead of the thiazolidine methyl ester, the corresponding thiazolidine-4-S-carboxylic acid tert-butyl ester is used and the tert-butyl ester of the reaction product is saponified with trifluoroacetic acid in a reaction step and simultaneously cleaves off the N-protecting group.

Example 5

N-[Na-(3-phenyl-2-S-acetylmercapto)-S-lysyl]-S-proline-pivaloyl-oxymethyl ester 3 g (6.6 mmol) N-[Na<->(3-phenyl-2 -S-acetylmercapto)-S-lysyl]-S-proline is dissolved in acetone, 660 mg (6.6 mmol) KHCO 3 , 185 mg potassium iodide and 1.16 ml (8.06 mmol) chloromethyl pivalate are added, and the whole is heated in 3 hours under reflux cooling. After cooling, the precipitated KCl is filtered off, and the filtrate is evaporated. The residue is taken up in ethyl acetate, washed successively with water, dilute NaHCO 3 solution, water and saturated NaCl solution and dried over MgSO 4 . After distillation of ethyl acetate, 2.1 g of an oily residue is obtained which is purified chromatographically over silica gel (elution: initially ethyl acetate, n-hexane = 2:1, then change to ethyl acetate, methanol = 9:1). 0.8 g of an oily product is obtained which NMR spectroscopically corresponds to the title compound.

Rf = 0.36 (ethyl acetate, methanol = 9:1, silica gel).

Example 6

N-C N~-( 3- phenyl- 2- S- acetylmercapto)- S- lysvl]- thiazolidine- 4- S-carboxylic acid- pivaloyloxy methyl ester

The thiazolidine-4-S-carboxylic acid pivaloyloxymethyl ester analogous to the title compound in example 5 is obtained when, instead of the proline compound in example 5, N-[Na<->(3-phenyl-2-S-acetylmercapto)-S-lysyl]-thiazolidine- 4-S-carboxylic acid.

Example 7

N, N'-[ N, N'-{ 2, 2'- S- dithiobis-( 3- phenylpropanoyl)}- bis- S- lvsvl]- bis- S- proline

3.9 g (6.43 mmol) N-[Na<->(3-phenyl-2-S-acetylmercaptopropan-oyl)-NE<->tert.butoxycarbonyl-S-lysyl]-S-proline-tert. butyl ester is dissolved in 50 ml of methanol, 10 ml of water and 5.6 ml of 5 N caustic soda are added and stirred for 1.5 hours at room temperature. Then 2.8 ml of 5 N caustic soda are added and stirring is continued for a further 1.5 hours. The methanol is distilled off in a vacuum, the residue is dissolved in 300 ml of dichloromethane and decanted with water. The organic phase is washed successively with dilute KHSO 4 solution, water and saturated NaCl solution, dried over MgSO 4 and distilled in vacuum.

3.4 g (= 94% of theoretical) of the free mercapto compound are obtained as an oil.

For oxidation, 3.4 g (6 mmol) of the above-mentioned deacetylated compound are dissolved in 100 ml of dichloromethane, 0.8 g (8 mmol) of KHCO^ are added, and a 2% ig methanolic iodine solution is added dropwise at room temperature until permanent yellowing (consumption approx. .

25 ml).

Add approx. 50 ml of water and reduce the excess of iodine with a few drops of a Na2S203 solution. The organic phase is separated, washed several times with water and unsaturated NaCl solution, dried over MgSO 4 and evaporated to dryness in vacuo.

2.2 g (= 64.9% of the theoretical) of the dimeric bis compound is obtained as a solidified foam.

For saponification of the ester and Ne<-> protecting group, the bis compound (2.2 g = 1.97 mmol) is covered with 60 ml of trifluoroacetic acid and stirred overnight at room temperature. The trifluoroacetic acid is distilled off in a vacuum, the residue is dissolved in dilute NaHCO-j solution and decanted with ethyl acetate. The aqueous phase is then acidified with dilute hydrochloric acid and evaporated in vacuo. The white, solid residue is extracted three to four times with methanol, undissolved components are filtered off, and the solvent is evaporated in vacuo to dryness. As a yield, 1.6 g (= 100% of the theoretical) of the title compound is obtained as a solidified foam.

Rf = 0.2 (butanol, acetic acid, water = 3:1:1, silica gel).

Example 8

N. N'-[ N. N'-{ 2. 2'- S- dithiobis-( 3- phenylpropanoyl) >- bis- S- lvsvl]-bis- S- proline- pivaloyloxy methyl ester

The starting compound is the title compound from example 7. Esterification to the title compound takes place analogously to the procedure described in example 5.

Example 9

N. N-[ N, N-{ 2, 2'- S- dithiobis-( 3- phenylpropanoyl) >- bis- S- lvsvl]- bis-thiazolidine- 4- S- carboxylic acid

The starting compound is N-[Na<->(3-phenyl-2-S-acetylmercapto-propanoyl)-NE<->tert-butoxycarbonyl-S-lysyl]-thiazolidine-4-S-carboxylic acid tert-butyl ester. The dimerization and protection group removal of the title compound occurs as described in Example 7.

Example 10

N, N- CN, N'-{ 2, 2'- S- dithiobis-(3- phenylpropanoyl)}- bis- S- lysyl]- bis- thiazolidine- 4- S- carboxylic acid- pivaloyloxy methyl ester

The starting compound is the title compound from example 9. Esterification to the title compound takes place analogously to the procedure described in example 5.

Example 11

N-[N~-( 3- phenyl)- 2- S- mercaptopropanoyl)- S- lysyl)]- S- proline

A solution of 3.7 g (6.5 mmol) N-[Na< ->(3-phenyl-2-R-bromopropanoyl)-Ne<->tert-butoxycarbonyl-S-lysyl]-S-proline methyl ester in 30 ml of ethyl acetate. It is then boiled under reflux, the previously cooled solution is filtered and washed successively with dilute KHSO 3 solution, NaHCO 3 solution, water and saturated NaCl solution. After drying over MgSO4, the solvent is distilled off in water, and 4.0 g of a yellowish oil is obtained as a residue. The chromatographic purification takes place over 120 g of silica gel with ethyl acetate, n-hexane = 2:1 as eluent, and thus 3.0 g (81.9% of the theoretical) of pure N-[Na<->(3-phenyl -2-S-acetylmercaptopropanoyl)-Ne<->tert.butoxycarbonyl-S-lysyl]-S-proline methyl ester as a colorless oil.

Rf = 0.26 (ethyl acetate, n-hexane = 2:1, Kieselgel 60 60F254Merck).

By deprotection as described in example 3, 1.7 g (= 78.7 µ of the theoretical) of the title product is obtained. Rj = 0.52 (ethyl acetate, butanol, water, acetic acid = 1:1:1:1, Kieselgel 60F„C.Merck).

254

Example 12

N-[ Na-(3- phenyl- 2- R- mercaptopropanoyl)- S- lysyl]- S- proline

The R S S-diastomer analogous to the compound according to example 11 is obtained when, instead of the R-bromo compound, the corresponding S-isomer is used, namely N-[Na<->(3-phenyl-2-S-bromopropanoyl)-NE<->tert. butoxycarbonyl-S-lysyl]-S-proline methyl ester.

R^ = 0.44 (butanol, glacial acetic acid, water = 3:1:1, Kieselgel 60F254Merck).

Example 13

N-[ N~-( 3- phenyl- 2- S- acetvlmercaptopropanoyl)- S- lysvl]- S- proline-( 5- methyl- 2- oxo- 1. 3- dioxolen- 4- yl)- methyl ester

A. 2.6 g (4.7 mmol) of N-[Na<->(3-phenyl-2-R-bromopropanoyl)-Ne<->tert.butoxycarbonyl-S-lysyl]-S-proline are dissolved in 400 ml of acetone and add in order 500.5 mg (5 mmol) KHCO^, 141 mg (0.85 mmol) Kl and 965 mg (5 mmol) 4-bromomethyl-S-methyl-2-oxo-1,3-dioxolane . The solution is boiled in a ^-atmosphere with stirring for 3 hours under reflux cooling, and after cooling undissolved components are filtered off.

After distilling off the solvent, the residue is dissolved in ethyl acetate and washed with water, saturated NaHCO 3 solution, water and saturated NaCl solution. After drying over MgSO^, ethyl acetate is distilled off in vacuo. The residue of 2.8 g is purified chromatographically over 90 g of silica gel (elution with ethyl acetate, n-hexane = 2:1). You get 2.3 g (= 73.4% of the theoretical) of an oily product which NMR spectroscopically corresponds to proline-(5-methyl-2-oxo-1,3-dioxolen-4-yl)-methyl ester the derivative.

Rf = 0.68 (ethyl acetate, n-hexane = 2:1, Kieselgel 60F254Merck)-

B. To replace bromine with an acetyl mercapto group, 0.7 g (6.9 mmol) of triethylamine is slowly added to a solution of 525 mg (6.9 mmol) of thiolacetic acid in 20 ml of anhydrous ether under a N2 atmosphere and ice cooling, and then the whole is added dropwise with stirring to a solution of 2.3 g (3.5 mmol) of the above-mentioned bromine compound in 20 ml of anhydrous ether and 10 ml of ethyl acetate. It is then boiled for a further 90 minutes under reflux, the previously cooled solution is filtered and washed successively with dilute KHSO 3 solution, NaHCO 3 solution, water and saturated NaCl solution. After drying over MgSO4, the solvent is distilled off in vacuo, and 2.3 g of a yellowish oil is obtained as a residue. The chromatographic purification takes place over 7.5 g of silica gel with ethyl acetate, n-hexane = 2:1 as eluent. One thus obtains 1.7 g (= 74.4% of the theoretical) of N-[Na<->(3-phenyl-2-S-acetyImercaptopropanoyl)-Ne<->tert.-butoxycarbonyl-S-lysyl] -S-proline-(5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl ester. C. To remove the tert.butoxycarbonyl-Ne<-> protecting group on lysine, 1.7 g (2.57 mmol) of the product from step B is stirred with 40 ml of trifluoroacetic acid for 3 hours at room temperature, then the reaction mixture is evaporated to dryness, and for to remove residual amounts of trifluoroacetic acid, the residue is dissolved three times each in acetone and chloroform and evaporated again. 2.3 g of the title compound is obtained.

Example 14

The R S S diastereomers analogous to the title compounds of Examples 1 and 2, namely N-[Na<->(3-phenyl-2-R-acetylmercapto-propanoyl)-S-lysyl]-S-proline and N-[Na<- >(3-phenyl-2-R-mercapto-propanoyl)-N-acetyl-s-lysyl]-S-proline is obtained when, instead of the R-bromo compound, the corresponding S-isomer is used, namely 3-phenyl-2-S- bromopropionic acid.

1. Analogous method for the preparation of therapeutically active amino acid derivatives with the general formula

where lo^e. oJLk^-vO^

R is H, acetyl,—bonaoyl,—pivaloyl or fe bufcokaykapbonyl,—R^ is H, acetyl;—beneoyl, 'pivaloyl or the rest

p is one of the numbers 1-4;

A is the residue of one of the α-amino acids i of the heteroatoms 0, S or N, or^/ • / - .1

F*2 and R., together with the N and C atoms form a \%~ =~,—(r=—e-l-l-e-r

^^T^^åd^e^^ulg-',- - s-em--k-a-n- væ r e—m e t-t e t~e'l l"e r-- i"nn e h oid e- en

R4er 0H, ip^dC^^-h-y^ -e^-a"lkoks.y,r=,f-enyi-G-1 -C4~

the groups

R? is C1-C4-alkyl, linear or branched, jtorøj^^ik-y-l-resireTT—can

jrøEÆ=-sj±b^jptei^r1:~OTEcl---G-H-~B,rTi».C-F-3 ,-■ ^Æenyl^^el%er'-r-pyr-ridyl«;

Q» Y and Z are 0, S, NRg, d^Rg, CHRg,

fsR9 R9 R9

- CH - CH - or - C = C - ,

with the proviso that at least one of the residues X, Y and Z is 0, S,

R9<R>9<R>9R9

- CH - CH - or - C = C -

and one or two of the residue's X, Y and Z may be NRg;

R 8 is hydrogen or C 1 -C 4 alkyl, linear or branched;

Rg is hydrogen, or joins with a neighboring residue Rg a phenyl ring or, for m oc n = 1, its dihydro form with the double bond in conjugation to the C-terminal carboxy group, and

m and n are each 0, 1 or ^, the sum of m and n being 1 or

and their salts,

characterized by

a) an acid with the general formula

where

R.j has the above meaning, is condensed with a dipeptide of the general formula

where

A and p have the meaning given above, and X means a suitable protecting group, or

b) a compound of general formula IIa

where

R1;X and p have the above meaning, are condensed with an amino acid A, where A has the above meaning, or

c) a compound of the general formula (V)

where

X, p and A have the meaning indicated above, are reacted with a mercapto compound of the general formula

where

R.J has the above meaning, or

d) a compound with the general formula

where

X and p have the meanings given above, are condensed with an amino acid A, where A has the meaning given above, and the thus obtained condensation product of the general formula V is reacted with a mercapto compound of the general formula VI, and optionally: compound of formula I where means -acetyl,—b-e-uzuyl

_§JJjar—p4-valoyl is converted into the corresponding compound of formula I where R1 means hydrogen, and/or

^£4- a compound of formula I where R„ is 'acetyl,—beii<Luyl or

pi x^a.lnyi and R is H, is transformed into a compound of formula I where is hydrogen and R is ace Lyitr;—benzoyl Sllér <i

.p4-valoyl, and/or

~- q- f~ for the preparation of dimeric compounds of the general formula I where R^ denotes the group

and the other residues have the meanings indicated above, a compound of the general formula I where R^ is hydrogen and the NHR group carries a protecting group is treated with an oxidizing agent, and protecting groups present are cleaved off at a suitable step of the method, and optionally a prepared final product of the general formula I to a salt. 2. Process according to claim 1, characterized in that compounds of formula I are prepared where R is H, p = 4 and A is proline or thiazolidine, and salts thereof. 3. Process according to claim 1 or 2, characterized in that N-[Na<->(3-phenyl-2-S-mercapto-propanoyl)-S-lysyl]-thiazolidine-4-S-carboxylic acid and salts thereof are prepared. 4. Process according to claim 1 or 2, characterized in that N-[Na<->(3-phenyl-2-S-mercapto-propanoyl)-S-lysyl]-S-proline and salts thereof are prepared. 5. Method according to claims 1 to 4, characterized in that compounds are prepared in which the asymmetric carbon atoms are in S-configuration.