NL8104046A - Hypotensive mercaptoacyl-proline derivs. - prepd. by acylating proline or its analogues with mercapto-carboxylic acid derivs. (BE 11.8.77) - Google Patents

Abstract

Cpds. of formula (I) and theis basic salts are new. In the formula, R is OH, amino or lower alkoxy; R1 and R4 are H, lower alkyl, phenly or phenyl-(lower alkyl); R2 is H, lower alkyl, phenyl (opt. substd. by halogen, lower alkyl or lower alkoxy), phenyl-(lower alkyl), diphenyl-(lower alkyl), triphenyl-(lower alkyl), lower alkylthiomethyl, phenyl-(lower alkylthiomethyl), lower alkanoylaminomethyl, -COR5, -C(=M)-M-R5, -C(=M)-NHR5, -SR6 or R7; R3 is H, OH or lower alkyl; R5 is lower alkyl, phenyl or phenyl-(lower alkyl); R6 is lower alkyl, hydroxy-(lower alkyl), amino (carboxy)-substd. lower alkyl, phenyl, or phenyl substd. by halogen, lower alkyl or lower alkoxy; R7 is a residue of formula (Ia) M is O or S; m is 1, 2 or 3; and n and p are 0, 1 or 2. (I) may be in the form of optical isomers or racemates (asterisks in formula (I) indicate asymmetric centres). (I) are blockers of angiotensin converting enzyme. They inhibit conversion of angiotensin I into the hypertensive angiotensin II, and thus reduce or eliminate angiotensin-induced hypertension. (I) can be administered orally or parenterally. Dosage for reducing blood pressure is pref. ca. 1-50 mg/kg/day. Specific (I) include 1-(2-benzoylthioacetyl)-L-proline, prepd. e.g. by acylating L-proline with chloroacetyl chloride in the presence of NaOH and reacting the product with thiobenzoic acid in the presence of K2CO3.

Description

- 1 - i a *>

A method of preparing a therapeutic preparation, as well as a method of preparing suitable compounds,

The invention relates to a method for preparing a therapeutic composition with an antihypertensive effect, in which a proline is brought into a form suitable for therapeutic use. Such a method can be considered to be known from the German "Offenlegungsschriften" 2,108,043 and 2,124,451. According to these "Offenlegungsschriften" one can use certain peptides with terminal proline groups to combat blood pressure increase. Namely, it has been found that these peptides are capable of inhibiting the enzyme, that angiotensin I, which substance in the. body is converted into angiotensin II, which has an antihypertensive effect.

According to the invention, a compound of the formula (I) of the formula sheet in which R represents a hydroxy or amino group or a lower alkoxy group, R 1 and R 4, is separately introduced a hydrogen atom, an optionally substituted lower alkyl group or a phenyl group, R 1 a hydrogen atom or a hydroxy or lower alkyl group and R2 represents a lower alkyl or phenyl group or a substituted phenyl group, the substituent of which is a halogen atom, a lower alkyl or alkoxy group, a phenyl-lower alkyl, diphenyl-lower alkyl or triphenyl -lower alkyl, a lower alkylthiomethyl, phenyl-lower alkylmethyl, or lower alkanoylamidomethyl group or the group Μ M or a

II II

R5-M-C, R5-NH-C, Rg-S

25 represents the group of formula 29, wherein Rj. a lower alkyl, phenyl or phenyl-lower alkyl group and Rg is a lower alkyl or phenyl group or a substituted phenyl group with a halogen atom, a lower alkyl or alkoxy group, a hydroxy-lower alkyl or amino (carboxy) substituent -lower alkyl group and M represents an acid P, and 30 substance or sulfur atom, / n = 0, 1 or 2 and m = 2, or a basic salt thereof in a form suitable for therapeutic application, 8-1 0 4 0 4 6 - 2 -

Namely, it has been found that these much simpler prolines, which are not peptides, are also suitable for effectively combating high blood pressure by the same mechanism. It is further important that they also exert this action when administered orally.

The stereoisomers of the L-form prolines of formula 2 are preferred.

The lower alkyl groups represented by the above symbols include straight and branched chain hydrocarbon radicals from methyl to heptyl, for example methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, isopentyl and the like. The lower alkoxy groups are of the same type with 1-7 carbon atoms, attached to the oxygen atom, for example, methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, t-butoxy and the like. The groups with 1-4 carbon atoms and in particular with 1 or 2 carbon atoms are preferred. Phenylmethyl is the preferred phenyl-lower alkyl group.

The lower alkanoyl groups are those with the acyl radicals of the lower C 1 -C 3 fatty acids, for example, acetyl, propionyl, butyryl, isobutyryl and the like. The lower alkanoyl groups with up to 4 carbon atoms, especially acetyl, are preferred.

The term "halogen" includes the four usual halogens, but chlorine and bromine are preferred. The substituted phenyl groups preferably contain the substituent in the 4-position of the ring. The hydroxy lower alkyl groups contain a hydroxy group on an alkyl chain in the same manner as those described above, preferably on the terminal carbon atom, for example, hydroxymethyl, 2-hydroxyethyl, etc. The amino (carboxy) lower alkyl groups contain one amino group and one carboxy group on a lower alkyl group, such as those described above, preferably both on one carbon atom, for example on the terminal carbon atom, as in the preferred 2-amino-2-carboxyethyl group.

The new compounds of the general formula I can be prepared in a manner known per se.

8104046 Λ ~ - 3 -

Generally, the compounds of the invention are prepared by acylating a compound of Formula 3 with an acid of Formula 4 or a chemical equivalent thereof.

The final product can be obtained not only by direct acylation with an acid of the formula IV but also via intermediates, such as (a) hal-haloalkanoic acids of the formula 5, wherein X is bromine, chlorine or iodine, or (b) a tosyloxyalkane -acid, that is to say an acid of the formula 5, wherein X is tosyloxy 10 (ch3 - <§> - “SC ^ O-) or (c) a substituted acrylic acid of the formula 6. The product of this acylation is then subjected to rearrangement or addition with the anion of a thiol or thioic acid of the formula 7.

When the acid of the formula IV is used as an acylating agent, acylation can be accomplished in the presence of a coupling agent, such as dicyclohexylcarbo-diimide or the like, or the acid can be activated by forming its mixed anhydride, symmetrical anhydride, acidic chloride, acid ester or use of the Woodward reagent K, N-20 ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline, or the like. In connection with the acylation methods, reference is made to Methods of Organic Chemistry (Houben-Weyl), Vol. XV, Part II, p. 1 et seq. (1974).

Compounds of formula 3 include, for example, proline, hydroxyproline, 4-methylproline and its lower alkyl esters and the like. The acylation of such compounds is described in more detail below.

According to a preferred method of preparing a compound of the formula 1, an acid or ester of the formula 3 is coupled with a haloalkanoic acid of the formula 5, wherein X represents halogen, preferably chlorine or bromine. This can be accomplished by using any of the known procedures, wherein the acid 4 is activated for the reaction with the acid 3 by forming a mixed anhydride, symmetrical anhydride, acid chloride or active ester or by using 8104046 * * - 4 - *

Woodward reagent K, EEDQ (N-ethoxycarbonyl-2-ethoxy-1,2-dihydroxy-quinoline) or the like.

The product of this reaction is a compound of the formula 10. The product of the formula 10 is subjected to a displacement reaction with the anion of a thioic acid of the formula 7 to form a product of the formula 11. When R is an ester group (ie R represents a lower alkoxy group), the ester group can be removed from an ester of the formula 11 by treatment with trifluoroacetic acid and anisole to yield the corresponding free acid. The corresponding acid can also be obtained by alkaline hydrolysis.

A variation of this procedure involves the use of an acrylic acid of the formula VI as starting material. This acrylic acid is first converted to the acid halide form and then reacted with a compound of the formula 3 to form a compound of the formula 13 and this intermediate is subjected to the addition reaction with the thiol or thioic acid 7 as described above.

A tosyloxyalkanoic acid of the formula 14 can also be used as the acylation agent of the acid of the formula 3, after which the acylation product is subjected to the displacement reaction, etc., as described above.

On the other hand, the acrylic acid of the formula 6 can first be reacted with the thioic acid of the formula 7 to form a product of the formula 15, which is converted into its acid halide, for example with thionyl chloride, and then coupled with the compound with formula 3, after which the procedure described above is further followed.

The acid or ester of the formula III can also be acylated with a "protected" form of a merc-mercaptoalkanoic acid of the formula 16, wherein Rg is the "protecting" group. Such "protecting" groups can be in the form described above.

After acylation, the protecting group can be removed from the product by any of the above-described 8104046 * 5 methods.

Further details regarding the preferred methods of preparing the compounds of the invention are set forth in the specific examples below.

According to a particularly preferred modification of the process, the acid or ester of the formula 3 is acylated with a haloalkanoyl halide of the formula 17, wherein each X independently represents a halogen, preferably chlorine or bromine, R represents hydrogen, lower alkyl or phenyl-lower alkyl and n is 0, 1 or 2. This reaction is effected in an alkaline medium, for example, a dilute alkali metal hydroxide solution, alkali metal hydrogen carbonate or alkali metal carbonate solution, at a reduced temperature, for example about 0-15 ° C. The reaction product is subjected to a displacement reaction with the anion of the thiol or thioic acid of the formula 7, also in an alkaline medium, preferably an alkali metal carbonate solution, and / or subsequently worked up in the usual manner. When an acid of the formula III is used as the starting material, the end product obtained as the free carboxylic acid can then be converted into its ester, for example by esterification with a diazoalkane, such as diazomethane, a 1-alkyl-3-p-tolyltriazene, such as ln -butyl-3-p-tolyltriazene or the like. The treatment of an ester, preferably the methyl ester, with an alcoholic ammonia solution converts the free acid 25 into the amide, that is to say in a product in which R represents nh 2.

According to another variation of the process, an ester, preferably the t-butyl ester, of the formula 3 in an anhydrous medium, such as dichloromethane, tetrahydrofuran, dioxane or the like, is treated with a thioalkanoic acid of the formula 18 in the presence of dicyclohexylcarbodiimide, N. N'-carbonyl bis-imidazole, ethoxyacetylene, diphenylphosphoryl azide or analogous scalping agents at a temperature of about 0-10 ° C. The ester group (R) can then be removed, for example, by treatment with trifluoroacetic acid and emisole at room temperature.

8104046 «δ - 6 -

Products of the formula 1, in which the group M

1R, -Mc- is prepared by reacting a compound of the formula 12 (see parent patent application 77.01457) with a halogenated compound of the formula 19 or by reacting a compound of the formula 10 with an alkali metal salt or alkaline earth metal salt of a compound of formula 20, wherein Me represents the alkali metal or alkaline earth metal.

A product of formula 1, where 1 * 2 represents the group

10 M

II

R 1 -NH-C- is prepared by reacting a compound of the formula 12 with a suitably substituted isocyanate or isothiocyanate of the formula 21. On the other hand, the same products can be obtained by coupling an acid of the formula 22 with a 15 amino acid of the formula 3.

Compounds of formula 1, wherein R 1 represents lower alkyl, phenyl, substituted phenyl, phenyl-lower alkyl, triphenyl-lower alkyl, lower alkylthiomethyl or phenyl-lower alkyl-thiomethyl, are obtained by reacting a compound of the formula 12 with the corresponding halide R2X or by reacting a compound of the formula 10 with the corresponding thiol R2SH in the same manner as described above.

A compound of formula 1, wherein R 2 represents a lower alkanoylaminomethyl group, can be prepared by condensing a compound of formula 12 with a corresponding hydroxymethyl-lower alkanoylamide of formula 23 in the presence of an acidic catalyst, such as trifluoroacetic acid.

A compound of formula 1, wherein R2 represents the group R1, -S, can be prepared using known methods for the synthesis of mixed disulfides, for example, by reacting a compound of formula 12 with a thiosulfinate of the formula 24, thiosulfonate of the formula 25, sulfenyl halide of the formula 26, thiosulfate of the formula 27, or sulfenyl thiocyanate of the formula 28.

In the special case where a group with the 81 0 4 0 4 ο ί * - 7 - represents formula 29 and R, Rj, R ^ and R ^ have the same meanings as the corresponding substituents in formula 1 and p is 0 the symmetrical disulfides are obtained by direct oxidation of a compound of formula 12 with iodine.

When p is 1 or 2, such products can be obtained by stepwise oxidation of the corresponding compound, wherein p is 0. Mixed disulfides are obtained using the modification described in the examples.

Products of the formula I have one or more asymmetric carbon atoms. When R ^, R ^ or R ^ have a meaning other than hydrogen, the carbon atom to which this substituent is attached is asymmetric. These carbon atoms are indicated by an asterisk in formula 1. The compounds therefore exist in stereoisomeric forms or in racemic mixtures thereof. All these forms and mixtures are within the scope of the invention. In the syntheses described above, the starting materials may be the racemate or one of the enantiomers. When the racemic starting material is used in the synthesis, the stereoisomers obtained in the product can be separated using conventional chromatographic or fractional crystallization methods. Generally, with respect to the carbon atom of the amino acid, the L isomer is the isomeric form, which is preferred. Also preferred is the D isomer with respect to the α-carbon atom in the acyl side chain (ie, the carbon atom that bears it).

The compounds of the invention form basic salts with different inorganic and organic bases, which salts are also within the scope of the invention. Such salts include ammonium salts, alkali metal salts such as sodium and potassium salts (preferred), alkaline earth metal salts such as the calcium and magnesium salts, salts with organic bases, for example the dicyclohexylamine salt, benzathine, N-methyl-D-glucamine hydrabamine salts, salts with amino acids such as arginine, lysine and the like. The non-toxic, physiologically acceptable salts are preferred, although others 8104043 ♦? Salts are also useful, for example, in the isolation or purification of the product, as illustrated in the examples for the case of the dicyclohexylamine salt.

The salts are conventionally formed by reacting the free acid form of the product with one or more equivalents of the appropriate base providing the desired cation in a solvent or medium in which the salt is insoluble or in water and removal of the water by freeze drying. By neutralizing the salt with an insoluble acid such as a hydrogenation cation exchange resin (eg polystyrene sulfonic acid resin such as Dowex 50) or with an aqueous acid and extraction with an organic solvent eg ethyl acetate, dichloromethane or the like, the free acid form and, if desired, another salt are formed.

Further experimental details are indicated in the examples, which describe preferred embodiments and also serve as a model for the preparation of other members of the group.

The compounds of the invention inhibit the conversion of the decapeptide angiotensin I to angiotensin II and are therefore useful for lowering angiotensin-related hypertension. Angiotensin I is formed by the action of the enzyme renin on angiotensinogen, a pseudoglobulin in blood plasma. Angiotensin I is converted by angiotensin converting enzyme (ACE) into angiotensin II. The latter is an active blood pressure-increasing agent, which is considered to be the cause of different forms of hypertension in different mammalian species, eg rats and dogs. The compounds according to the invention block the conversion angiotensin (renin) - * angio-tensin I - angiotensin II by inhibiting the angiotensin converting enzyme and reducing or eliminating the formation of blood pressure-increasing angiotensin II, By administration of a composition containing one or more more compounds of the formula 1 and / or physiologically acceptable salts thereof, the angiotensin-induced hypertension in affected mammals, may 8104046; «- 9 - are reduced. A single dose or preferably 2-4 daily divided doses based on about 0.1-100 mg per kg per day and preferably about 1-50 mg per kg per day is suitable for lowering blood pressure, as shown by the animal experiments, described by SL Engel, T.R. Schaeffer, M.H. Waugh and B. Rubin, Proc. Soc.

Exp. Biol. Med. 143, 483 (1973). The substance is preferably administered orally, but parenteral administration, such as subcutaneous, intramuscular, intravenous or intraperitoneal administration, can also be used.

The compounds of the invention can be used for lowering blood pressure in the form of preparations such as tablets, capsules or elixirs for oral administration or in sterile solutions or suspensions for parenteral administration. About 10-500 mg of a compound or mixture of compounds of the formula 1 or physiologically acceptable salts thereof are mixed with physiologically acceptable vehicles, carriers, excipients, binders, preservatives, stabilizers, flavors, etc. to form a unit dosage form as required in pharmaceutical practice. The amount of active substance in these compositions is such that an appropriate dose is obtained within the above range.

Examples of additives that can be included in tablets, capsules and the like are the following! a binder such as gum tragacanth, acacia gum, corn starch or gelatin, an excipient such as dicalcium phosphate, a disintegrant such as corn starch, potato starch, alginic acid and the like, a lubricant such as magnesium stearate, a sweetener such as sucrose, lactose or saccharin, and such as peppermint, wintergreen or cherry oil. When the unit dosage form is a capsule, it may additionally contain, in addition to the materials of the above type, a liquid carrier, such as a fatty oil. Various other materials may be present as coatings or to further modify the physical form of the unit dose . For example, tablets can be coated with shellac, sugar or both. A syrup or elixir may contain the active compound, 8104046 * C-10 sucrose as a sweetener, methyl and propyl parabenz as preservatives, a colorant and a flavoring agent such as cherry or orange flavor.

Sterile preparations for injection can be prepared according to conventional pharmaceutical procedures by dissolving or suspending the active substance in a vehicle, such as water for injection, a naturally occurring vegetable oil, such as sesame oil, coconut oil, peanut oil, cottonseed oil, etc. , or a synthetic fat vehicle, such as ethyl oleate or the like. If necessary, buffers, preservatives, anti-oxidants and the like can be included.

The invention is further illustrated but not limited by the following examples.

Example_I

IV 3- (Ethoxy) carbonyl / thio / propanoyl-L-proline.

Ethyl chloroformate (1.2 g) is added to a solution of 3-mercaptopropanoyl-L-proline (2.03 g) in normal sodium hydrogen carbonate (30 ml) and the mixture is stirred vigorously at 5 ° C for 1 hour and at room temperature for 2 hours. After acidification with concentrated hydrochloric acid, the mixture is extracted with ethyl acetate. The organic phase is washed with water, dried over magnesium sulfate and evaporated to dryness to yield the title compound.

25 example_II

1-3 / 3 / (Ethoxy) thiocarbonyl / thio / porpanoyl / L-proline.

Aqueous 2 N. sodium hydroxide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added to a solution of 30 L-proline (5.75 g) in N. sodium hydroxide (50 ml), cooled and stirred in an ice bath. After 5 min. The ice bath is removed and stirring is continued at room temperature. After 3 hours, ethyl cantoic acid potassium salt (9.6 g) is added and the mixture is stirred at room temperature overnight. The solution 35 is acidified with concentrated hydrochloric acid and extracted with ethyl acetate, the organic layer is evaporated to dryness and the residue is chromatographed on a column of silica gel with a mixture of benzene-acetic acid (7µl) as solvent, the title being connection is obtained? m.p. 94-95 ° C.

Example III

1- / 3 - / - / (Benzylthio) carbonyl / thio / propanoyl / - L-proline.

A solution of benzylthiocarbonyl chloride (11 ml) in dioxane (20 ml) is added in 5 portions to a solution of 1- (3-mercaptopropanoyl) -L-proline (1.6 g) in normal sodium hydrogen carbonate (24 ral) cooled in an ice bath over a 30 minute period. The ice bath is removed and stirring is continued at room temperature for 2.5 hours. After acidification with concentrated hydrochloric acid, the aqueous phase is extracted with ethyl acetate. The organic phase is dried over magnesium sulfate and evaporated to dryness to yield the title compound.

Example IV_ 1V 3 * V / (Ethylthio) thiocarbonyl_ / thio_ / propanoyl _ / - L-proline, 20

Aqueous 2N. sodium hydroxide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added to a solution of L-proline (5.75 g) in N. sodium hydroxide (50 ml), cooled and stirred in an ice bath. After 5 min. The ice bath is removed and stirring is continued at room temperature. After 3 hours, ethyl trithiocarbonate potassium salt (10.5 g) is added and the mixture is stirred at room temperature overnight. After acidification with concentrated hydrochloric acid, the mixture is extracted with ethyl acetate. The organic layer is dried over magnesium sulfate and evaporated to yield the title compound.

Example V

3- / / (Methylamino) thiocarbonyl / thio / propionic acid.

35 Methyl isothiocyanate (4 g) is added to 8104046%

·· X

- 12 - s a solution of 3-mercaptopropionic acid (5.3 g) in a mixture of pyridine (250 ml) and 0.5 N sodium hydroxide (100 ml). The solution is kept at 40 ° C for 2 hours and evaporated to dryness in vacuo. The residue is dissolved in water (100 ml), acidified with concentrated hydrochloric acid and extracted with ether. The organic phase is evaporated to dryness to yield the title compound; m.p. = 86-87 ° C.

Example VI

1- / 3-LL (Me thylamine) - thiocarbony 1 / thio / propanoy 1 / -L-pro 1 in -10 t-butyl ester.

To a solution of L-proline tert-butyl ester (1.71 g) and hydroxybenzotriazole (1.35 g) in dichloromethane (10 ml), cooled and stirred in an ice bath, dicyclohexylcarbo-15 diimide (2.06 g) and 3-methylaminothiocarbonylthiopropionic acid (1.79 g). After 15 min, the bath is removed and stirring is continued overnight. The precipitate is filtered off and the filtrate diluted with ethyl acetate and washed until neutral reaction. The organic phase is evaporated to dryness to yield the title compound; m.p. = 129-130 ° C.

Example_VlI

1 - / _ 3 / _ / (Methylamino) thiocarbony 1 __ / thio __ / propanoyl _ / - L-proline.

(A) 1- (Methylaminothiocarbonylthiopropanoyl) - 25 L-proline tert.-butyl ester (0.98 g) is dissolved in a mixture of anisole (3.6 ml) and trifluoroacetic acid (7.5 ml). After 1 hour at room temperature, the mixture is evaporated to dryness in vacuo and the residue precipitated 3 times from ether-hexane. This material is chromatographed on a column of silica gel with a solvent mixture of benzeneacetic acid (75:25) to yield the title compound; = 0.4 silica gel-benzene: acetic acid (75:25) /. The dicyclohexylamonium salt has a m.p. from 127-129 ° C.

(B) Methyl isothiocyanate (4g) is added to a solution of 3-mercaptopropanoyl-L-proline (10.1g) in a mixture of pyridine (250ml) and 0.5N sodium hydroxide (100ml). The solution is kept at 40 ° C for 2 hours and then evaporated to dryness in vacuo. 8104046 - 1 - 13. The residue is dissolved in water (100 ml), acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic phase is evaporated to dryness to obtain the title compound.

Example VIII

1- / 3- / (Ethylamino) carbonyl / thio / propanoyl / L-proline.

Ethyl isocyanate (0.45 ml) is added to a solution of 1- (3-mercaptopropanoyl) -L-proline (1 g) in a mixture of N. sodium hydroxide (5 ml) and pyridine (5 ml). The solution is heated at 40 ° C for 4 hours and concentrated in vacuo. The residue is partitioned between 0.1 N, hydrochloric acid and ethyl acetate. The organic layer is washed with water, dried over magnesium sulfate and evaporated to dryness to yield the title-15 compound. The dicyclohexylammonium salt is prepared by adding dicyclohexylamine to a solution of the free acid in ethyl acetate; m.p. = 150-152 ° C.

Example IX

1- / 3- / / (Ethoxy) carbony l __ / thio_y-2-thy thypropanoyl __ / - L-proline.

20

By using 1- (3-mercapto-2-methylpropanoyl) -L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example I, the title compound is obtained,

Example_X

25 l - / _ 3- / L (Ethoxy) carbonyl / / thio / / butanoyl / / - L-proline.

By using 1- / 3-mercaptobutanoyl / L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example I, 1V 3- / / (ethoxy) carbonyl / thio / butanoyl / - L- 30 proline obtained.

v22E5eelÉ- $ I

1- / 4- / (Benzylthio) carbonyl / thio / butanoyl / L-proline.

By using 4-mercaptobutanoyl-L-proline instead of 35 3-mercaptopropanoyl-L-proline in the procedure of 8104046 t-14 - Example III, 1 - / - 4 - / / (benzylthio) carbonyl_ / thio_ / propanoyl / L-proline.

Example_XII

1 - / "2-ΓΓ (Benzyl thio) carbonyl / thio / propanoyl / L-proline.

5

Using 2-mercaptopropanoyl-L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example III gives 1- / 2- / / (benzylthio) carbonyl / thio / propanoyl / -L-proline.

10 Y22? * S? ISL2iii 1 - / - 3 - / f (Ethylthio) thiocarbonyl / thio / propanoyl / L-proline methyl ester.

A solution of 1 - / - 3 - / / (ethyl thio) thio-15 carbonyl / thio / propanoyl / l-proline in ethyl acetate is treated with an ethereal solution of diazomethane until a permanent yellow color is obtained. After removing the yellow color with a few drops of acetic acid, the solvents are removed in vacuo to yield the title compound.

20 Y22E ®2IIL5IY_ 1 - / 3 - / _ / _ (Methylamino) thiocsu: bonyl / thio> _ / - 2-methylpropanoyl / / - L-proline amide.

By using 1- (3-mercapto-2-methylpropanoyl) -L-proline-25 amide instead of 3-mercaptopropinoyl-L-proline in procedure B of Example VII, 1 - / 3V / (methylamino) thio-carbonyl_ / thio-2-methylpropanoyl-1-L-proline amide. Example XV

1- / 3- / / (Phenoxy) carbonyl / thio / propanoyl / -L-proline.

30

By using phenyl chloroformate instead of ethyl chloroformate in the procedure of Example 1, 1 / 3- / (phenoxy) carbonyl / thio / propanoyl / L-proline is obtained.

35 8104046 * - 15 -

Example_XVI

1 - / 3 - // ^ (Penoxy) carbonyl / thio / butanoyl / L-proline.

By using phenyl chloroformate instead of ethyl 5 chloroformate and 4-mercaptobutanoyl-L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example I, IV 3V / (phenoxy) carbonyl / thio-Butanoyl-L-proline obtained.

Example Xyl 10 IV 3V / (Phenylamino) carbonyl / thio / propanoyl / L-proline.

By using phenyl isocyanate instead of ethyl isocyanate in the procedure of Example VIII, IV 3 - / - / (phenylamino) carbonyl / 1hio / propanoy 1 / - L-proline is obtained.

Example XVIII

IV 3- / / (Phenethylamino) carbonyl / thio / propanoyl / L-proline.

By using phenethyl isocyanate instead of ethyl isocyanate in the procedure of Example VIII, 20 IV of 3V / (phenethylamino) carbonyl / thio / propanoyl / -L-proline is obtained.

Y ° ° £ image JCIX

IV 3- / / (Ethylamino) carbonyl / thio / 2-benzylpropanoyl / L-proline.

By using 1- (3-mercapto-2-benzylpropanoyl) -L-proline instead of 1- (3-rnercaptopropanoyl) -L-proline in the procedure of Example VIII, 1V 3- / A (ethylamino) carbon- nyl / thio-A-2-benzylpropanoyl / L-proline.

Example XX

1- (3-Methylthiopropanoyl) -L-proline.

(A) Methyl 3-methylthiopropionate (51 g) is saponified with a 10% sodium hydroxide solution (150 ml, 30 min at 100 ° C). The cooled solution is extracted with ether 35 and then acidified. The crude acid thus obtained is distilled and converted into the acid chloride with thionyl chloride.

A solution of L-proline (11.5 g) in N. sodium hydroxide (100 ml) is cooled in an ice bath and 3-raethylthiopropanoic acid chloride (6.9 g) is added dropwise with vigorous stirring over a period of 10 min. After 5 hours, the reaction mixture is acidified and extracted with ethyl ether to yield the title compound. The dicyclohexylammonium salt is prepared by adding dicyclohexylamine to a solution of the free acid in ethyl acetate; m.p. 169-171 ° C.

(B) Methyl iodide (71 g) is added to a solution of 1- (3-mercaptopropanoyl) -L-proline ethyl ester (115 g) and sodium (11.5 g) in ethanol (400 ml). The reaction is allowed to proceed overnight, then the ethanol is removed in vacuo and the residue is dissolved in a mixture of ethyl acetate and water. The organic layer is dried and evaporated to dryness in vacuo.

The resulting 1- (3-methylthiopropanoyl) -L-proline ethyl ester (98g) is suspended in a mixture of methanol (200ml) and 5N sodium hydroxide (200ml) and stirred at room temperature for hours. The methanol is removed in vacuo and the aqueous phase is extracted with ethyl acetate, acidified and re-extracted with

ethyl acetate. The latter organic phase is washed with water, dried and evaporated to dryness to yield the title compound. Example ^ XXI

1- / 3- (4-Chlorophenylthio) propanoyl / L-proline.

25

Aqueous 2 N, sodium hydroxide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added to a solution of L-proline (5.75 g) in N. sodium hydroxide (50 ml), cooled and stirred in a ice bath. After 5 min. The ice bath is removed and stirring is continued for 3 hours at room temperature. The reaction mixture is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is washed with water, dried and evaporated to dryness in vacuo. The residue is dissolved in a mixture of 4-chlorobenzenethiol (8 g), sodium hydroxide (4.2 g) and ethanol (300 ml). The solution becomes 8104046 V r - 17 -

heated under reflux for 6 hours. The solvent is removed in vacuo and the residue dissolved in water, acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is washed with water, dried and evaporated to dryness in vacuo to give the title compound. Example XXII

1- / / (3-Benzylthiomethyl) thio / propanoyl / L-proline.

1- (3-Mercaptopropanoyl) -L-proline (8.1 g) is dissolved in boiling liquid ammonia (100 ml) and small pieces of sodium are added until a permanent blue color is obtained, which is then removed with a small piece ammonium chloride. Benzylthiomethyl chloride (6.9 g) is added and the ammonia is allowed to evaporate. The last traces of ammonia are removed in vacuo, the residue is dissolved in water and extracted with ethyl acetate. The aqueous phase is acidified with concentrated hydrochloric acid and extracted with ethyl acetate.

The organic layer is washed with water, dried and evaporated to dryness in vacuo to give the title compound.

1- / / (3-Acetamidomethyl) thio / propanoyl / L-proline.

1- (3-Mercaptopropanoyl) -L-proline (2 g) and N-hydroxymethylacetamide (0.89 g) are dissolved in trifluoroacetic acid (10 ml) and the solution is stored at room temperature for 1 hour. The excess trifluoroacetic acid is removed in vacuo and the residue is precipitated several times from ether-hexane. Finally, the residue is partitioned between dilute hydrochloric acid and ethyl acetate. The organic layer is washed with water, dried and evaporated to dryness to yield the title compound.

Example XXIV 1- (Methylthioacetyl) -L-proline.

Using methylmethylthioacetate instead of 8104046-18-methyl-3-methylthiopropionate in procedure A of Example XX gives 1- (methylthioacetyl) -L-proline, (mp = 123-124 ° C).

Example XXV

5 l- (Benzylthioacetyl) -L-proline.

Using benzylthioacetyl chloride instead of 3-methylthiopropanoyl chloride in procedure A of Example XX gives 1- (benzylthioacetyl) -L-proline (mp = 86-88 ° C).

Example 10CVI - 1/3 - / (2-Phenylethyl) thio / propanoyl / L-proline.

Using phenethyl bromide instead of methyl iodide in procedure B of Example XX gives 1 - / - 3 - / - (2-phenylethyl) thio / propanoyl / - L-proline.

Example_XXVII

1- / 3- / (Triphenylmethyl) thio / propanoyl / L-proline.

Using triphenylmethyl chloride instead of methyl iodide in procedure B of Example XX gives 1 - / - 3-f - (triphenylmethyl) thio / propanoyl / - L-proline.

1- (3-Methylthio-2-methylpropanoyl) -L-proline amide.

25

By using 1- (3-mercapto-2-methylpropanoyl) -L-proline amide instead of 1- (3-mercaptopropanoyl) -L-proline ethyl ester in procedure B of Example XX and omitting the saponification step 1 - (3-Methylthio-2-methylpropanoyl) -L-proline-30 amide.

Example_XXIX

1- / 3- (4-Methoxyphenylthio) propanoyl_yL-proline.

Using 4-methoxybenzenethiol instead of 4-35 chlorobenzenethiol in the procedure of Example XXI gives 1 - / - 3 - (4-methoxyphenylthio) propanoyl / - L-proline.

8104046 - 19 -

Vqorbegld XXX

1- / 2- (4-Chlorophenylthio) propanoyl / L-proline.

Using 2-bromopropionyl chloride instead of 5 3-bromopropionyl chloride in the procedure of Example XXI gives 1- / 2- (4-chlorophenylthio) propanoyl / L-proline.

Example XXXI

1- / 3- / (Diphenylmethyl) thio-2-benzylpropanoyl-L-proline.

10

Diphenylmethanol (0.92 g) and 1- (3-mercapto-2-benzylpropanoyl) -L-proline (1.5 g) are dissolved in trifluoroacetic acid (10 ml) and the solution is left in room temperature for 30 min. temperature. The excess trifluoroacetic acid is removed in vacuo to give 1- / 3- / (diphenylmethyl) thio-2-benzylpropano-yl / L-proline.

Example XXXII

1- / 4- (4-Chlorophenylthio) butanoyl / L-proline.

Using 4-bromopropionyl chloride instead of 3-bromopropionyl chloride in the procedure of Example XXI gives 1- / 4- (4-chlorophenylthio) butanoyl / L-proline.

1- / 3- / (Benzylthiomethyl) thio / butanoyl / / L-proline.

By using 3-mercaptobutanoyl-L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XXII, 1 - / - 3 - / - (benzylthiomethyl) thio - / butanoyl - / - L-30 proline is obtained.

Example XXXIV

1- / / 4 - / "(Acetamidomethyl) thio / - 2-methylbutanoyl_y-L-proline.

By using 1- (4-mercapto-2-methylbutanoyl) -L-proline 35 instead of 1- (3-mercaptopropanoyl) -L-proline in the 8104046-20 procedure of Example XXIII, 1 / £ 4- (accetamidomethyl) -thio-7-2-methylbutanoyl / L-proline.

Example XXXV

1 -f ~ 3- (Ethyldithio) propanoyW-L-proline.

5 (A) 3-Mercaptopropinoyl-L-proline (10 g) is added to a solution of ethyl thiosulfinate (8.4 g) in methanol (100 ml) and the reaction mixture is stirred vigorously at room temperature for 4 hours. The methanol is removed in vacuo to yield 1- (3-ethyldithiopropanoyl) -L-proline.

(B) A solution of ethyl thiosulfinate (8.4 g) in ethanol (50 ml) is added to an aqueous solution of 3-mercaptopropanoyl-L-proline (10 g), which is kept at a pH of 6-7 by gently add sodium hydroxide.

The mixture is stirred vigorously at room temperature until the reaction to thiol is negative. The mixture is diluted with water, adjusted to pH 8 and extracted with ethyl acetate, the aqueous phase acidified to pH 3 and extracted again with ethyl acetate.

The latter extract is washed with water, dried and evaporated to dryness to yield 3- (ethyldithio) propanoyl-L-proline.

1- / 3- / (4-Methylphenyl) dithio / propanoyl / L-proline.

25

A solution of 4-methylphenylsulfenyl chloride (1.76 g) in ether (20 ml) is added to a solution of 3-mercaptopropanoyl-L-proline (2 g) in 0.5 N. sodium hydroxide (20 ml) cooled in an ice bath . The mixture is stirred vigorously for 1 hour and the aqueous phase separated, acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic phase is washed with water, dried and evaporated to dryness, leaving IV / .3- (4-methylphenyl) dithiol. / propanoyl / L-proline is obtained, 810104046

J

- 21 -

Example xxxv IV 3- (Phenyldithio) propanoyl / L-proline.

Phenyl thiosulfinate / prepared from phenyl disul-5-fide according to U. Weber and P. Hartter, Z. Physiol. Chem., 351, 1384 (1970) in instead of using ethyl thiosulfinate in the procedure of Example XXXV, IV 3- {phenyldithio) propanoyl / L-proline is obtained.

Example XXXVIII

10 1V. 3V (2-Penylethyl) dithio / propanoyl / L-proline.

Using 2-phenylethylthiosulfinate (prepared from phenyldisulfide) in place of ethylthiosulfinate in the procedure of Example XXXV gives 1- / 3- (2-phenylethyl) dithio-propanoyl-L-proline.

Example XXXIX IV 3V (2-Hydroxyethyl) dithio / ptopanoyl / L-proline.

Mercaptoethanol (4.2 g) is added to a solution of 1,1'V (sulfinylthio) bis-20 (3-propanoyl) _J-bis-L-proline (21 g) in methanol (100 ml) and the reaction mixture is stirred vigorously at room temperature for 4 hours. The methanol is removed in vacuo and the residue is purified by silica gel column chromatography to give 1- / 3V (2-hydroxyethyl) dithio [propanoyl] L-proline.

Example XL

1- / 2- (E thyldi thio) propanoyl / L-proline.

Using 2-mercaptopropanoyl-L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XXXV gives 1- / 2- (ethyl dithio) propanoyl / -L-proline.

Example_XLI

IV 3V (4-Methylphenyl) dithio / / butanoyl / / -L-proline.

35 8104046

V

- 22 -

Using 3-mercaptobutanoyl-L-proline in place of 3-mercaptopropanoyl-L-proline in the procedure of Example XXXVI gives 1 - 3 - (4-methylphenyl) dithio / butanoyl / L-proline.

5 Y22E522i! Lïïi 1 - / - 3- (Ethyldithio) - 2 -me thy lpr opanoy 1 __ / - L-pro line -me thy le s ter.

By using 1- (3-mercapto-2-methylpropanoyl) -L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XXXV and then treating it with diazomethane according to the procedure of Example V, 1 - / 3- (ethyldithio) -2-methylpropanoyl / - L-proline-methyl ester. Example_XLIII

1 3 - / (4-Methylphenyl) dithio-2-methylpropanoyl / 2L-hydroxyproline.

15

By using 1- (3-mercapto-2-methylpropanoyl) -L-hydroxy-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XXXVI, 1 / 3- / (4-methylphenyl) -dithio_ / -2-methyl propanoyl / / -L-hydroxyproline.

20 Y ° 2 £ image_XLIV

IV 3V (2-Amino-2-carboxyethyl) dithio / propanoyl / L-proline.

A 0.5 M solution of thiocyanogen in ice vinegar is prepared by shaking for 10 min in a sealed flask 600 mg dry lead thiocyanate with a solution of 75 µl bromine in 3 ral acetic acid. After the removal of lead bromide and the excess lead thiocyanate by centrifugation, 2.5 ml of this solution are mixed with 2.5 µl of a 0.41 M solution of cysteine hydrochloride, which has been previously neutralized with dilute sodium hydroxide. This mixture is immediately added to 0.75 ml of a 1.9 M solution of 3-mercaptopropanoyl-L-proline, which has been previously neutralized with dilute sodium hydroxide.

After 20 min. The mixture is titrated with alcoholic iodine to an incipient brown color and adjusted to a pH of 3. The precipitate is removed by filtration and the filtrate is applied to a column of cation exchange resin (Dowex 50) 8104046-23. The column is washed with water until no more acidic material is removed and then eluted with pyridine acetate buffer of pH 6.0. The fractions containing the disulfide of cysteine and 3-mercaptopropanoyl-5 L-proline are collected and evaporated to dryness.

Example XLV

1,1 '/ Dithiobis (3-propanoyl) / bis-L-proline.

3-Mercaptopropanoyl-L-proline (0.95 g) is dissolved in water (20 ml) and the pH is adjusted to 6.5 with N. sodium hydroxide. An ethanolic solution of iodine is added dropwise while maintaining the pH at 6.5 by cautious addition of N, sodium hydroxide. When a permanent yellow color is obtained, the addition of iodine is stopped and the color is removed with a small amount of sodium thiosulfate.

The reaction mixture is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic phase is washed with water, dried and evaporated to dryness to give 1.1 * dithiobis (3-propanoyl) / bis-L-proline. The dicyclohexyl ammonium salt is prepared by adding dicyclohexylamine to a solution of the free acid in acetonitrile; m.p. = 179-180 ° C. Example XLVI

1,1 '- / Dithiobis (2-D-methyl-3-propanoyl) - bis-L-proline.

By using 3-mercapto-2D-methylpropanoyl-L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XLV, 1,1 '/ dithiobis (2-D-methyl-3-propanoyl) - bis-L-proline obtained, m.p. = »236-237 ° C.

Example_XLVII

1.11 - / Dithiobis (2-propanoyl) / bis-L-proline.

By using 2-mercaptopropanoyl-L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XLV, 1.1'-dithiobis (2-propanoyl) / bis-L-proline is obtained.

8104046 - 24 -

Example_KLVIII

1.1 * - (Dithiobisacetyl) bis-L-hydroxyproline.

Using 1- (2-mercaptoacetyl) -L-hydroxyproline instead of 3-mercaptopropionyl-L-proline in the procedure of Example XLV gives 1.1 '- (dithiobisacetyl) bis-L-hydroxy-proline.

Y2Ieldi-XL1x 1.1'-Dithiobis (3-propanoyl) _-bis-4-methyl-L-proline.

10

By using 1- (3-mercaptopropanoyl) -4-methyl-L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XLV, 1.1 '/ dithiobis (3-propanoyl) __ / - bis- 4-methyl-L-proline obtained.

Example L-1.1 '/ Dithiobis (2-benzyl-3-propanoyl) / bis-L-proline.

By using 1- (3-mercapto-2-benzylpropanoyl) -L-proline instead of vein 3-mercaptopropanoyl-L-proline in the procedure of Example XLV, 1.1 · / dithiobis (2-benzyl-3-propanoy1) / bis-L-proline obtained.

1.1'Dithiobis (4-butanoyl) / bis-L-proline.

Using 4-mercaptobutanoyl-L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XLV gives 1.1'-dithiobis (4-butanoyl) bis-L-proline.

Lil 30 1.1 '- / Dithiobis (2-benzyl-4-butanoyl) _ / - bis-L-proline.

Using 1- (4-mercapto-2-benzylbutanoyl) -L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XLV gives 1.1 '/ dithiobis (2-benzyl-4-buta-35) noyl) bis-L-proline.

8104046

X

- 25 -

Example sentence 1.11 - / Dithiobis (3-butanoyl) / bis-L-proline.

Using 3-mercaptobutanoyl-L-proline instead of 5-3-mercaptopropanoyl-L-proline in the procedure of Example XLV gives 1.1 dithiobis (3-butanoyl) bis-L-proline.

Example LIV

1.1 '- / Dithiobis (3-propanoyl) V-bis-L-proline-me thyles ter10

A solution of 1,1'-dithiobis (3-propanoyl) -1-bis-L-proline in methanol is treated with ethereal diazomethane until a permanent yellow color is obtained. After 15 minutes, a few drops of acetic acid are added and the solvents are removed in vacuo to yield the title compound. Example LV

1.1 '- / Dithiobis (3-propanoyl) / bis-L-proline amide.

A solution of 1.1'-dithiobis (3-propanoyl) -120 bis-L-proline methyl ester in methanol is saturated with ammonia while cooling in an ice-water bath. The reaction mixture is stored in a pressure bottle at room temperature for 16 hours and then the solvents are removed in vacuo to yield the title compound.

25 Example LVI

1.1 '- / Dithiobis (2-phenyl-3-propanoyl) - bis-L-proline.

By using 1- (3-mercapto-2-phenylpropanoyl) -L-proline instead of 3-mercaptopropanoyl-L-proline in the procedure of Example XLV, 1.1 * / dithiobis (2-pheny1-3-propanoyl) / bis-L-proline obtained.

Example LVII

1.1 (sulfinylthio) bis (3-propanoyl) -1 bis bis-L-proline.

While cooling in an ice bath, 0.12 ml of 8104046-26-peracetic acid are added to a stirred solution of 1.1'V dithiobis (3-propanoyl) V-bis-L-proline (40 g) in glacial acetic acid (500 ml ). The reaction mixture is left overnight at room temperature and then the solvent is removed in vacuo to give the title compound.

Example LVIII

1.1 (Sulfonylthio) bis- (3-propanoyl) -1 bis-L-proline.

A 30% solution of hydrogen peroxide (2.0-10 ml) is added to a solution of 1.1'dithiobis (3-pro-panoyl) -1 bis bis-L-proline (4 g) in glacial acetic acid (80 ml) and the solution is stored at room temperature for 30 hours. The solvent is removed in vacuo to yield the title compound.

15 Example_LIX

1.1 (Sulfinylthio) bis (2-propanoyl)> bis bis L-proline.

By using 1,1 '- / dithiobis (2-propanoyl) __ / - bis-L-proline instead of 1,1dithiobis (3-propanoyl) _ / - bis-L-proline 20 in the procedure of Example LVIX, 1.1 - / (sulfinylthio) bis- (2-propanoyl) / bis-L-proline.

Example LX

By using 1.1 '- (dithiobisacetyl) bis-L-azetidine carboxylic acid instead of 1.1' - / dithiobis (3-propanoyl) _ / - bis-L-25 proline in the procedure of Example LVI1, 1.1 '- / (sulfinylthio) bis-acetyl / bis-L-azetidine-2-carboxylic acid.

Example LXX

1.1 * - / - (Sulfinylthio) -bis- (3-propanoyl) _ / - bis-4-methyl-L-proline.

30

By using 1.1 '- / dithiobis (3-propanoyl) __ / - bis-4-methyl-L-proline instead of 1.1' - / dithiobis (3-propanoyl) _ / - bis-L-proline in the procedure of Example LVII gives 1.1 '- / (sulfinylthio) -bis- (3-propanoyl) -1 / - bis-4-methyl-L-proline.

8104046 - 27 -

Example LXII

1.i '- / (Sulfinylthio) -bis- (2-benzyl-3-propanoyl) -1- bis-L-proline.

By using 1.1 '- / dithiobis (2-benzyl-3-propanoyl) 5 bis-L-proline instead of 1.1' - / dithiobis (3-propanoyl) _ / - bis-L-proline in the example procedure LVII gives 1,1 '- / (sulfinylthio) -bis- (2-benzyl-3-propanoyl) -1 / bis-L-proline.

1.1 '- / ~ (Sulfylylthio) -bis- (4-butanoyl) 1 / bis-L-proline.

By using 1.1 * - / dithiobis (4-butanoyl) _ / - bis-L-pro-line instead of 1.1'V dithiobis (3-propanoyl) ^ / bis-L-proline in the procedure of Example LVII, 1.1 '- / (sul-15-finylthio) -bis- (4-butanoyl) -7-bis-L-proline.

Example LXIV

1.1 '- / (Sulfinylthio) -bis- (3-butanoyl) _ / - bis-L-proline,

By using 1.1 '- dithiobis (3-butanoyl) _ / - bis-L-pro-20 line instead of 1.1' V dithiobis (3-propanoyl) __ / - bis-L-proline in the procedure of Example LVII 1.1 '- / (sulfinylthio) -bis- (3-butanoyl) -1 / - bis-L-proline is obtained.

Example_LXV

1.1 '- /' * (Sulfinylthio) -bis- (2-methyl-3-propanoyl) -1- bis-L-proline.

25

By using 1.1 '- dithiobis (2-methyl-3-propanoyl) _ / - bis-L-proline instead of 1.1'V dithiobis (3-propanoyl) _ / - bis-L-proline in the procedure of example LVII gives 1.1 * - / (sulfinylthio) -bis- (2-methyl-3-propanoyl) -1- bis-L-proline.

Example LXVI

1.1 * - / Sulfinylthio) bis- (2-phenyl-3-propanoyl) / bis-L-proline.

By 1.1 '- / "dithiobis (2-pheny 1-3 -propanoyl) __ / -bis-35 L-proline instead of 1.1' - / dithiobis (3-propanoyl) _ / - bis-L-proli- When used in the procedure of Example LVII, 1.1 '- ((sulfinylthio) bis- (2-phenyl-3-propanoyl) -1 / bis-L-proline is obtained.

1- / 3- / 3- (2-Carboxy-1-pyrrolidinyl) -3-oxopropyl / dithio-2-methyl-5-propanoyl / L-proline.

By using 1.1 '- / {sulfinylthio) bis (2-methyl-3-propanoyl) _ / - bis-L-proline instead of ethylthiosulfinate in the procedure of Example XXXV, \ - / _ 3 - / _ / 3- (2-carboxy-1-10 pyrrolidinyl) -3-oxopropyl / dithio-2-methylpropanoyl / L-proline.

Preliminary LXVIII

1.1 '- / (Sulfonylthio) -bis-acetyl_ / -bis-L-hydroxyproline.

By using 1.1 '- (dithiobisacetyl) -bis-L-hydroxyproline instead of 1.1 * - / _ dithiobis {3-propanoyl) -bis-L-proline in the procedure of Example LVIII, 1.1' - / _ (sulfonylthio bis-acetyl / bis-L-hydroxyproline.

Y22 ^ 2ë ^ _ ^ H

1.1 '- / (Sulfonylthio) -bis- (2-benzyl-4-butanoyl) -1- bis-L-proline.

By using 1.1 '- / dithiobis (2-benzyl-4-butanoyl) _ / - bis-L-proline instead of 1.1' - / _ dithiobis (3-propanoyl) __ / - bis-L-pro-line for the procedure of example LVIII is obtained 1.1'-25 / (sulfonylthio) -bis- (2-benzyl-4-butanoyl) -1- bis-L-proline.

Example LXX

a) 3 - / - (4-Methoxyphenyl) methylthio / - 2-methylpropanoic acid.

30 p-Methoxy-α-toluenethiol (15.4 g; 0.1 mol) is added to a solution of methacrylic acid (8.6 g; 0.1 mol) in 50 ml of 2N sodium hydroxide. The mixture is heated on a steam bath for 3 hours, then refluxed for 2 hours and cooled. The mixture is extracted with ether and the aqueous layer is acidified with concentrated HCl 8104046-29 and extracted with dichloromethane. The acidic extracts are washed with table salt solution, dried (magnesium sulfate) and evaporated in vacuo. The resulting semi-solid is taken up in 50 ml of dichloromethane, diluted with 50 ml of hexane and cooled. The title compound is obtained in the form of a white crystalline solid; m.p. 74-82 ° C (5.5 g).

b) 3- (4-Methoxyphenyl) methylthio / 2-methylpropanoyl-L-proline t-butyl ester.

10 3- / (4-Methoxyphenyl) methyl thio-2-methylpropanoic acid (3.6 g, 0.015 mol) L-proline tert-butyl ester (2.6 g, 0.015 mol) and dicyclohexylcarbodiimide (3.1 g 0.015 mol) are dissolved in 50 ml of dichloromethane and stirred at 0 ° C for 30 min. The cooling bath is removed and the mixture is stirred overnight (16 hours). The resulting suspension is filtered and the filtrate washed with 5% potassium hydrogen sulfate, saturated sodium hydrogen carbonate and brine, then dried (magnesium sulfate) and evaporated in vacuo. The resulting clear oil is applied to a 250 ml silica gel column and chromatographed using 20% ethyl acetate / hexane as the eluent. The main fraction (R ^ = 0.70, silica gel, ethyl acetate) is evaporated to 5.5 g (93%) 1 - / - 3 - (4-methoxyphenyl) methyl-thio / -2-methylpropanoyl-L-proline. tert-butyl ester as a clear oil R ^ = 0.70 (silica gel, ethyl acetate), Rf = 0.60 (silica gel, ether).

Example LXXI

1 - / - Dithiobis (2-methyl-3-propanoyl) / bis-L-proline.

By using 3,3'-dithiobis-2-methylpropanoic acid instead of 3-acetylthio-2-methylpropanoic acid in the procedure of example, 1- / dithiobis- (2-methyl-3-propanoyl) __ / - bis-L-proline obtained.

Example LXXII

3-Benzylthio-2-methylpropanoic acid 8104046-30 - a) By using cx-toluenethiol instead of p-methoxy-a-toluenethiol in the procedure of Example LXXa, 3-benzylthio-2-methylpropanoic acid is obtained.

b) 1 3- (Benzylthio) -2-methylpropanoyl-1-L-proline tert-butyl ester.

5 ”"

By using 3-benzylthio-2-methylpropanoic acid instead of 3V (4-methoxyphenyl) methylthio / - 2-methylpropanoic acid in the procedure of Example LXXb, 1 - / - 3 - (benzylthio) - 2-methylpropanoyl / - L-proline tert-butyl ester obtained.

1 ° Example LXXIII

1- / 3- (Benzylthio) -2-methylpropanoyl / L-proline.

1- / 3- (Benzylthio) -2-methylpropanoyl / L-proline ter t-butyl ester (7.8 g) is dissolved in a mixture of anisole 15 (55 ml) and trifluoroacetic acid (11-0 ml). After storage at room temperature for 1 hour, the solvent is removed in vacuo and the residue is doused in ether, washed several times with saturated sodium chloride, dried over magnesium sulfate and evaporated in vacuo to give the title compound. Rf 0.5 (Silica gel, benzene / acetic acid 3: 1) 0.5, (Silica gel, methyl ethyl ketone / acetic acid / pyridine / water 14: 1: 2: 1).

Y22 keld_LXXlV_ a) 3-Triphenylmethylthio-2-methylpropanoic acid.

A solution of 3-mercapto-2-methylpropanoic acid (1.2g) and trityl chloride (2.9g) in methylene chloride (50ml) is kept at room temperature for 2 hours. The mixture is heated in a steam bath for 20 min and then evaporated to dryness in vacuo and the residue is dissolved in saturated aqueous sodium hydrogen carbonate and the solution washed with ethyl acetate. The aqueous phase is acidified to a pH of 3 and extracted with ethyl acetate. The organic layer is dried and evaporated to dryness to yield the title compound. R ^ 0.8 (silica gel, benzene / acetic acid 3: 1).

8104046-31 - b) 1- / 3- (Triphenylmethylthio} -2-methylpropanoyl / L-proline tert-butyl ester.

By using 3-triphenylmethylthio-2-methylpropanoic acid 5 instead of 3- (4-methoxyphenyl) methylthio-2-methylpropanoic acid in the procedure of Example LXXb, the title compound is obtained.

Example LXXV

1 - / - 3- (Triphenylmethylthio) -2-methylpropanoyl / - L-proline.

10 3-Triphenylmethylthio-2-methylpropanoic acid (1.8g) and N.N'-carbonyldiimidazole (0.8g) are dissolved in tetrahydrofuran (10ml) with stirring at room temperature. After 20 min, the solution is added to a mixture of L-proline (0.6 g) and 15 N-methylmorpholine (1 g) in dimethylacetamide (20 ml). The resulting mixture is stirred at room temperature overnight, evaporated to dryness and the residue dissolved in a mixture of ethyl acetate and 10% aqueous potassium hydrogen sulfate. The organic layer is separated and dried and evaporated to dryness in vacuo to give the title compound. Rf: 0.4 (silica gel, benzene / acetic acid 3: 1), R ^ 1.0 (silica gel, methyl ethyl ketone / acetic acid / pyridine / water 14: 1: 2: 1).

a) 3- (Tetrahydropyran-2-ythio) -2-methylpropanoic acid.

25

Boron trifluoride etherate (2.8 g) is added to a solution of 3-mercapto-2-methylpropanoic acid (2.4 g) and freshly distilled 2,3-dihydro-4H-pyran (1.9 g) in benzene (60 ml). . After 2 hours, potassium carbonate (4 g) is added, the mixture is stirred and filtered. The filtrate is evaporated to dryness to yield the title compound.

b) 1- / 3- (Tetrahydropyran-2-ythio) -2-methylpropanoyl / L-proline.

By using 3- (tetrahydropyran-2-ylthio) -2-methylpro-35panoic acid instead of 3-triphenylmethylthio-2-methylpropanoic acid in the procedure of Example LXXV 8104046 r "V - 32" - the title compound is obtained Rf: 0/8 (silica gel, benzene / acetic acid 3: 1, Rf:

0.75 (silica gel, methyl ethyl ketone / acetic acid / pyridine / water 14: 1: 2: 1). Y ° ° £ keeld_LXXVII

A) 3-Acetamidomethylthio-2-methylpropanoic acid.

3-Mercapto-2-methylpropanoic acid (2.4 g) and N-hydroxymethyl acetamide (1.8 g) are dissolved in trifluoroacetic acid and the solution is stored for 1 hour at room temperature. The trifluoroacetic acid is removed in vacuo and the residue dried in vacuo over potassium hydroxide to give the title compound.

b) 1- / 3- (Acetamidomethylthio) -2-methylpropanoyl / L-proline.

By using 3-acetamidomethylthio-2-methylpropanoic acid instead of 3- (tetrahydropyran-2-ylthio) -2-methylpropanoic acid in the procedure of Example LXXVIb, the title compound is obtained, Rj0.2 (silica gel, benzene / acetic acid 3: 1) ), R ^ 0.3 (silica gel, methyl ethyl ketone / acetic acid / pyridine / water 14: 1: 2: 1).

20 Pre-image_LXXYIIX_ 1000 Tablets, each containing 100 mg of 1- (2-mer-captopropanoyl) -L-proline, are prepared from the following ingredients: 1.1 * - / dithiobis- (2-D-methyl-3-propa-25-noyl) Bis-L-proline 100 g

Corn starch 50 g

Gelatin 7.5 g

Avicel (microcrystalline cellulose) 25 g

Magnesium stearate 2.5 g. The 1- (2-mercaptopropanoyl) -L-proline and corn starch are mixed with an aqueous solution of the gelatin. The mixture is dried and ground into a fine powder. The Avicel and then the magnesium stearate are mixed with the granulate. The mixture is then compressed into tablets to form 1000 tablets, each containing 100 mg of the active ingredient.

8104046

Claims (6)

1. A process for the preparation of a therapeutic composition having an antihypertensive effect, in which a proline is brought into a form suitable for therapeutic use, characterized in that a compound of the formula 1 in which R is a hydroxy or amino group or a lower alkoxy group, R 1 and R 1 each separately a hydrogen atom, an optionally substituted phenyl group with a lower alkyl group or a phenyl group, R 1 represents a hydrogen atom or a hydroxy or lower alkyl group, and R 2 represents a lower alkyl or phenyl group or a substituted phenyl group, the substituent of which is a halo, a lower alkyl or alkoxy group, a phenyl-lower alkyl, diphenyl-lower alkyl or triphenyl-lower alkyl, a lower alkyl thiomethyl, phenyl-lower alkyl methyl-lower alkanoylamidomethyl group or the group Μ M (I ff 15. R5 -M-C, Rg-NH-C-, RgS or a group of the formula 29, wherein R 1. a lower alkyl, phenyl or phenyl-lower alkyl group and Rg is a lower alkyl or phenyl group or a substituted phenyl group with a halogen atom, a lower alkyl or alkoxy group, a hydroxy lower alkyl or amino (carboxy) substituent ) -lower alkyl group and M represents a B oxygen or sulfur atom, / and n = 0, 1 or 2 and m = 2 or a basic salt thereof. 2. Process according to claim 1, characterized in that 1.1'V dithiobis (2-D-methyl-3-propanoyl / bis-L-proline) is used. Process for preparing a compound suitable for use in the process according to claim 1, characterized in that a compound of the formula 1, wherein R, R 1, R 2, R 1, R 1, r and n have the same meaning as in claim 1, prepared in a manner customary for such compounds. 35 8104046 • Λ *. RaR'nr »i Π5.776 R4 R (RJ | Ri, 1, 1, 1. V Re-s - (CHI - CH — COOH X - (ch)„ CH — COX 16 17 Rj Μ MI II l | Ri-S- (CH2), -CH-COOH 18 Rr-MCX g Rr-.M -CS-Me ^ QM Ri, Rf Rs_ N = C = M II I. I 0 21 Rf NH —CS (CH) W CH COOH R_ | _S_R 22 <> 24. Lower alkyl-CO - NHCHaOH 0? 3 11 iS R ^ -SSR ^ Ris — xo 25 26 R4 ~ S ~ S ° 3H27 1¾— S - SCN (HC) CHl ^ * III R-OC— HC-N-CO-CH-fCI-ή —S (ol * * J * k / p 29 SR Squibb & Sons, Inc. of Princeton, New Jersey, 8104046 United States America r3 Reg.No .: 115,776 A ** R2 —S-fCH) -CH - CO —N— CH — COR * Ri, T1 r ^ i HN - CH— COR ^ Ri z1. I 4 R $ ——S— (ChJ ^ —CH— COOH HF * ΐ M1 x “(chL” ch — C00H 5 CH «· C - COOH g \ Ri - SH 1 * R4 R (H2C- (ch]“ 10 χ— (Ch] ^ - CH — CO- N-CH-COR r R4 R, Η, £ - (άψ. R; —S— / ch) „- CH - CO — N —CH — COR ψ 11 Ri , R, HeC-CCH) ^ HS— (CHJ-CH-CO-N-CH-COR R 12 I3 Ril Ri HjC (CH) W I I I I CH-C-CO-N-CH-COR 13 _ f1 C ^ / isOiO-iCHl-CH-MOH 'K Γ Ir 15 Rg - S - CH - CH - COOH E.R. Squibb & Sons, Inc. 8 1 0 4 0 4 6 in Princeton, New Jersey Ver. S t. America