NO156408B - ANALOGY PROCEDURE FOR THE PREPARATION OF THERAPEUTIC ACTIVE L-PROLINE DERIVATIVES. - Google Patents

Description

The present invention relates to an analogue method for the production of therapeutically active L-proline derivatives of formula (I)

wherein R is an acyl group attached to the o-amino group of an amino acid and is selected from the group consisting of cyclopropanecarbonyl, cyclohexanecarbonyl and adamantanecarbonyl, A is a glycine, sarcosine or o<-D-amino acid residue with a ©C-carbonyl group which forms a thiol ester bond with the sulfur atom (S),

and in which the 2-methyl-propanoyl part has the D-configuration, or pharmaceutically acceptable salts thereof and the characteristic feature of the method according to the invention is that

a) either a compound of formula (II) is reacted

wherein A' is a glycine, sarcosine or a protected or

unprotected o£ -D-amino acid residue, and R has the above meaning, or a reactive derivative thereof, with a connect-

else with formula (III)

in which R' is a hydrogen atom or a carboxyl protecting group or a reactive derivative thereof, in an inert organic solvent at a temperature between -50 and 20°C, any protecting group is removed from the reaction product and, if desired, the reaction product is converted into a pharmaceutically acceptable salt, or

b) a compound of formula (IV)

wherein A1 is a glycine, sarcosine or a protected or unprotected oC-D amino acid residue, and R has the above meaning, or a reactive derivative thereof, is reacted with a compound of formula (V)

in which R' is a hydrogen atom or a carboxyl protecting group or a reactive derivative thereof, in an inert organic solvent at a temperature between -50 and 20°C, any protecting group is removed from the reaction product and, if desired, the reaction product is converted into a pharmaceutically acceptable salt. These features of the invention appear in the patent claim.

Compounds similar to the compounds represented by formula (I), e.g. compounds in which R is a benzoyl, acetyl or t-butyloxycarbonyl group and A is an L-o(-amino acid are described in UK patent application

No. 2,050,359A. European Patent Application (EP) 9898A1

describes compounds in which R is benzoyl, acetyl, t-butyloxycarbonyl, cyclopentanecarbonyl, etc., and A

is L-fl-amino acid. EP 35.38 3A1 describes compounds in which R is a benzoyl group and A is a D-phenylalanyl group. The compounds of formula (I) are thus new. The compounds are indicated for pharmaceutical use.

The amino acid residue represented by A in formula (I) is, as mentioned, glycine, sarcosine or an o<-D-amino acid residue. Examples of o<-D-amino acids can be mentioned neutral, acidic or basic amino acids, or aliphatic, aromatic, heterocyclic or alicyclic amino acids such as D-alanine, D-leucine, D-asparagine, D-methionine, D-glutamine, D- phenylalanine, D-tryptophan, D-ornithine, D-phenylglycine,

D-threonine, D-glutamic acid, D-arginine, D-cysteine, D-aspartic acid, D-histidine, D-isoleucine, D-proline, D-lysine, D-serine, D-tyrosine and D-valine. Functional groups in these amino acids such as hydroxyl, mercapto, amino and carboxyl may be substituted with lower alkyl, benzyl and lower alkanoyl. Preferred examples of A include glycine, sarcosine, D-alanine, D-leucine, D-methionine, D-glutamine, D-phenylalanine,

D-tryptophan and D-phenylglycine.

the group in formula (D has an asymmetric carbon atom so that the group can be in the D-form, L-form or DL-form, i.e. a mixture of the D- and L-forms. In accordance with the present invention, the D-form is used on because of its biological activity The proline residue in formula (I) may be in the D-form, L-form or .DL-form, but in accordance with the present invention the L-form is used because of its biological activity.

Illustrative pharmaceutically acceptable acids of the proline derivatives of formula (I) include salts with alkali metals such as sodium and potassium, salts with alkaline earth metals such as calcium and magnesium, and salts with basic amino acids such as arginine and lysine. The calcium salts and the lysine salts are preferred.

If A in formula (I) has a free functional group which should not be included in the reaction, such as e.g. mercapto, hydroxyl, amino or carboxyl, the compound of formula (II) means a derivative in which such a group may be protected.

A functional group in A' which should not enter the reaction can be protected with such groups which are conventionally used within this technique of peptide syntheses and which can be removed under relatively mild conditions. E.g. a mercapto group can be protected with an aralkyl group such as e.g. trityl, benzyl or p-methoxybenzyl; a hydroxyl group may be protected with a benzyl group, an amino group may be protected with a t-butyloxycarbonyl group and a carboxyl group may be protected with a t-butyl group. These protective groups can be removed by treatment with hydrogen fluoride, trifluoroacetic acid or hydrogen chloride. Alternatively, a carboxyl group is protected with a lower alkyl-substituted silyl group, such as e.g. a trimethylsilyl group, which is then removed by treatment with water. When R' in formula (III) is a carboxyl protecting group, this can be a t-butyl group or a lower alkyl-substituted silyl group and can be removed by treatment with hydrogen fluoride, trifluoroacetic acid, hydrogen chloride or water.

The reactive derivative of the compound with formula (II) is such that the carboxyl group that is included in the reaction is activated. The carboxyl group can be activated in any of the forms usually used within the technique of peptide synthesis, such as e.g. activated amides, acid halides, activated esters and mixed acid anhydrides. Among these forms, activated esters with N-hydroxysuccinimide, mixed acid anhydrides with carboxylic acid monoesters, and activated amides with carbonyldiimidazole are preferred. Carbodiimides such as dicyclohexylcarbodiimide can be used as a condensing agent to form an amide bond between the carboxyl group and the imino group.

The reactive derivatives of the compound of formula (III) are such that the imino group in the compound is activated. The imino group can be activated using any of the methods conventionally used in the field of peptide synthesis, e.g. the method of introducing a silyl group such as e.g. the trimethylsilyl group, the 'phosphazo method' using a phosphorus compound such as phosphorus trichloride (Ann. Chem., 572 96 (1951)), the phosphoric acid ester method using a phosphoric acid ester such as eg the tetraethyl ester of pyrophosphoric acid, or the "N-carboxylic anhydride method" (NCA method).

The reaction can be carried out in an inert organic solvent such as e.g. tetrahydrofuran, dioxane, dimethylformamide, hexamethylphosphoric triamide, chloroform, dichloromethane or acetonitrile. The reaction is usually carried out under cooling or at room temperature (-50

to 20°C), preferably from -30 to 10°C. The reaction period varies with the reaction temperature, the compounds to be reacted and the solvent, and is usually between 0.5 and 48 hours, preferably between 1 and 6 hours.

After the amide formation reaction, protecting groups in the reaction product are removed by any of the methods indicated above for the respective protecting groups.

The final product can be isolated from the reaction mixture and purified by any conventional method, e.g. by different types of chromatography using silica gel, dextran-crosslinked polymer and porous polymers such as e.g. styrene-divinylbenzene or acrylic acid esters. A suitable developing solvent may be selected from chloroform, ethyl acetate, methanol, ethanol, tetrahydrofuran, benzene, water and acetonitrile. Alternatively, the end product can be obtained by first isolating the reaction product in the form of an organic salt such as a dicyclohexylamine salt which is then treated with an acid such as hydrochloric acid or potassium hydrogen sulfate.

The compound of formula (I) obtained in this way has a carboxyl group at the proline site, so that it can form salts with various basic materials if desired. Salts with pharmaceutically acceptable basic materials are particularly important. These salts can be prepared by a conventional method, ie by treating the carboxyl group with an equimolar amount of any of the above stated bases, e.g. alkali metals, alkaline earth metals and basic amino acids.

The compound with formula (II) which is used as starting material in the analogical method according to the invention can be prepared using the following method:

A compound of formula (VI):

in which R has the same meaning as previously indicated, or a reactive derivative thereof in the carboxyl group, is reacted with an amino acid of formula (VII)

wherein A" is an amino acid residue equivalent to A'minus the group NH or C=0; R" is a hydrogen atom or a carboxyl protecting group, or a reactive derivative thereof, followed by removal of R" if this is a protecting group, to to form a compound with formula

(VIII)

wherein R and A" have the previously stated meaning, upon which the compound of formula (VIII) or a reactive derivative thereof is reacted with a compound of formula (IX)

wherein R''' is a hydrogen atom or optionally a carboxyl protecting group, followed by removal of R''' if this is a protecting group.

Examples of reactive derivatives in the carboxyl group of the compound of formula (VI) include those commonly used in the art of peptide synthesis, such as e.g. activated amides, acid halides, activated esters and mixed acid anhydrides.

If R" in the amino acid of formula (VII) is a protecting group, advantageous examples include a t-butyl group which can be removed by hydrogen fluoride, trifluoroacetic acid or hydrogen chloride, a benzyl group which can be removed by catalytic reduction with palladium or the like and lower alkyl groups such as methyl and ethyl which can be removed by hydrolysis under alkaline conditions.

The reactive derivative of the amino acid of formula (VII) is such that the amino group is activated, and such a derivative is prepared using any of the activation methods conventionally used in the area of peptide syntheses, such as e.g. introduction of a silyl group, e.g. a trimethylsilyl group, or the "phosphazo method", the phosphoric acid ester method and the "N-carboxylic anhydride method".

A carbodiimide such as dicyclohexylcarbodiimide can be used as a condensing agent to form an amide bond between the compound of formula (VI) and the amino acid of formula (VII).

The reaction can be carried out in an inert organic solvent such as e.g. tetrahydrofuran, dioxane, dimethylformamide, hexamethylphosphoric triamide, chloroform, dichloromethane or acetonitrile. The reaction is generally carried out under cooling or at room temperature (-50 to 20°C), preferably at from -30 to 10°C. The reaction period varies with the reaction temperature, the compounds that are reacted and the solvent, and is usually between 0.5 and 48 hours, preferably between 1 and 6 hours.

If R" in formula (VII) is a protecting group, this can be removed from the reaction product by any of the conventional methods described above.

The compound of formula (VIII) can be isolated from the reaction mixture and purified by any conventional method, e.g. by recrystallization from ethyl acetate, n-hexane, acetone or water, or by different types of chromatography as described in connection with the isolation and purification of the compound of formula (I), or by cleavage of an organic salt of the reaction product with an acid.

The compound of formula (VIII) can be reacted with the compound of formula (IX) by reacting the carboxyl group in the former compound with the mercapto group in the latter compound in the presence of a condensing agent such as carbodiimide; e.g. dicyclohexylcarbodiimide. A reactive derivative of the compound of formula (VIII) can be reacted with the compound of formula (IX). Examples of a reactive derivative of the compound of formula (VIII)

include activated amides, acid halides, activated esters and mixed acid anhydrides, activated amides with carbonyldiimidazole being preferred.

The carboxyl group in the compound of formula (IX) may optionally be protected. Preferred protecting groups are t-butyl and other groups which can be removed under acidic conditions.

The thiol ester-forming reaction is carried out in an inert organic solvent such as e.g. tetrahydrofuran, dioxane, dimethylformamide, hexamethylphosphotriamide, chloroform, dichloromethane and acetonitrile. The reaction is usually carried out under cooling or at room temperature (-50 to 20°C), preferably from -30 to 10°C when the compound of formula (VIII) is used in the form of an activated amide or acid halide, and from -10 to 10°C when the compound is used in the form of an acid anhydride. The reaction period varies with the reaction temperature, the compounds to be reacted and the solvent, and is usually between 0.5 and 48 hours, preferably between 1 and 6 hours. After the thiol ester formation reaction, R'" is removed from the reaction product if it is a protecting group. If the protecting group is a t-butyl group, it can be removed by a conventional method, i.e. by reaction with hydrogen fluoride, trifluoroacetic acid or hydrogen chloride .

The final compound can be isolated from the reaction mixture and purified using conventional methods, such as e.g. recrystallization from an organic solvent such as ethyl acetate or n-hexane, and various types of chromatography as described in connection with the isolation and purification of the compound of formula (I), or by cleavage of an organic salt of the reaction product with an acid.

If A in formula (I) has a free functional group which should not be included in the reaction, such as e.g. mercapto, hydroxyl, amino or carboxyl, the compound of formula (IV) means a derivative in which such a group may be protected.

Any functional group in A' which should not be included in the reaction is protected by means of the groups which are conventionally used in the area of peptide syntheses and which can be removed under relatively mild conditions. E.g. a mercapto group can be protected with an aralkyl group such as e.g. trityl, benzyl or p-methoxybenzyl,

a hydroxyl group can be protected with a benzyl group,

an amino group can be protected with a t-butyloxycarbonyl group and a carboxyl group can be protected with a t-butyl group. These groups can be removed by treatment with hydrogen fluoride or hydrogen chloride.

Alternatively, a carboxyl group is protected with a lower alkyl-substituted silyl group, such as e.g. the trimethylsilyl group, which is then removed by treatment with water. When R'

in formula (V) is a carboxyl protecting group, this can be a t-butyl group or a lower alkyl-substituted silyl group and can be removed by treatment with hydrogen fluoride, trifluoroacetic acid, hydrogen chloride or water.

The reactive derivative of the compound with formula (IV) is such that the carboxyl group that is included in the reaction is activated. The carboxyl group can be activated in a suitable form such as e.g. activated amides, acid halides, activated esters or mixed acid anhydrides. Activated amides with carbonyldiimidazole are preferred. Carbodiimides such as dicyclohexyl carbodiimide can be used as a condensing agent to form an amide bond from the carboxyl group and the mercapto group.

The reaction between the compound of formula (IV) or its reactive derivative and the compound of formula (V) or its reactive derivative is carried out in an inert organic solvent such as e.g. tetrahydrofuran, dioxane, dimethylformamide, hexamethylphosphotriamide, chloroform, dichloromethane or acetonitrile. The reaction is usually carried out under cooling or at room temperature (-50 to 20°C) and preferably at between -30 and 10°C when the compound of formula (IV) is used in the form of an activated amide or acid halide, and at between -10 and 10 °C when the compound is used in the form of an acid anhydride. The reaction period varies with the reaction temperature, the compounds that are reacted and the solvent, and is usually between 0.5 and 48 hours, preferably between 1 and 6 hours. After the thiol ester formation reaction, protecting groups are removed from the reaction product by any of the methods specified for the respective protecting groups.

The final compound can be isolated from the reaction mixture and purified by any of the conventional methods mentioned above, such as e.g. different types of chromatography using silica gel, dextran-crosslinked polymer and porous polymers such as e.g. styrene-divinylbenzene or acrylic acid esters. A suitable developing solvent may be selected from chloroform, ethyl acetate, methanol, ethanol, tetrahydrofuran, benzene, water and acetonitrile. Alternatively, the final compound can be obtained by first isolating the reaction product in the form of an organic salt such as e.g. the dicyclohexylamine salt which is then treated with an acid such as hydrochloric acid or potassium hydrogen sulfate.

The final compounds according to the invention, namely the L-proline derivative of formula (I) and pharmaceutically acceptable salts thereof, prevent the formation of angiotensin II from angiotensin I by inhibiting the activity of the angiotensin-converting enzyme. The compounds are therefore useful for the treatment of hypertension due to angiotensin II and as an agent for the treatment of heart failure.

The activity of some of the final compounds prepared in accordance with the invention for inhibiting the angiotensin-converting enzyme was measured.

(1) Method

An angiotensin-converting enzyme was extracted from a rabbit lung. A 0.111M boric acid-Na^O^ buffer solution

(pH 8.3, 0.6 ml), 0.2 ml of 0.111M boric acid-Na2CO3 buffer solution (pH 8.3) containing 25 mM benzoylglycylhistidyl-leucine (substrate) and 0.1 ml of 0.111M boric acid -Na-CO..-— 8 — 3 buffer solution (pH 8.3) containing from 10 to 10 M of the test compounds (13 final compounds prepared in accordance with the invention as indicated in the following table) were placed in test tubes and preincubated at 37°C for from 5 to 10 min. Then 0.1 ml of a solution of the enzyme (acetone powder) was added to each test tube and each mixture was incubated at 37°C for 30 min. The benzoyl-glycine produced by the enzyme was extracted with ethyl acetate in the presence of hydrochloric acid and the amount was determined by UV absorption at 228 nm. The enzyme activity in the presence of the test compounds relative to that in the absence of the test compounds (100) was determined. The concentration of each test compound at which the relative activity of the enzyme was 50% was referred to as the activity of each test compound that inhibited the activity of the enzyme and was

indicated as a I^^ value.

(2) Results

Compound No. 1: N-(3-(N-cyclohexanecarbonyl-D-alanyl-thio)-2-D-methylpropanoyl7-L-proline

No. 2: N-(3-(N-cyclopropanecarbonyl-D-alanyl-thio)-2-D-methylpropanoyl7-L-proline

No. 3: N-[3-(N-cyclohexanecarbonylglycylthio)-2-D-methylpropanoyl7-L-proline

Compound No. 4: N-(3-(N-cyclohexanecarbonyl-N-methyl-glycylthio)-2-D-methylpropanoyl/-L-proline

No. 5: N-[3-(N-cyclopropanecarbonylglycylthio)-2-D-methylpropanoylJt7-L-proline

No. 6: N-[3-(N-adamantanecarbonylglycylthio)-2-D-methylpropanoyl7-L-proline

No. 7: N-/TJ-(N-cyclohexanecarbonyl-D-phenyl-alanylthio)-2-D-methylpropanoyl7-L-

proline

No. 8: N-^3-(N-cyclohexanecarbonyl-D-leucyl-thio)-2-D-methylpropanoyL7-L-proline

No. 9: N-/3-(N-cyclohexanecarbonyl-D-tryptophyllthio)-2-D-methylpropanoyl/-L-proline

No. 10: N-/3-(N-cyclohexanecarbonyl-D-phenyl-glycylthio)-2-D-methylpropanoyl7-L-proline

No. 11: N-/3-(N-cyclohexanecarbonyl-D-methionyl-thio)-2-D-methylpropanoyl/-L-proline

No. 12: (N-cyclohexanecarbonyl-D-glutaminyl-thio)-2-D-methylpropanoyl/-L-proline

No. 13: N-(5-(N-adamantanecarbonyl-D-alanylthio)-2-D-methylpropanoyl7-L-proline

The activity of the final compounds produced in accordance with the invention lasts longer than known compounds with a similar activity, such as e.g. 3-mercapto-2-D-methylpropanoyl-L-proline (conventionally called "Captopril"), so that the desired control of blood pressure can be achieved with fewer daily injections. The mentioned compound "Captopril" and other known compounds cause a sudden drop in blood pressure during the initial administration period and this can develop into orthostatic hypotensive asthenia

(Lancet, vol. 1, no. 8, 115, page 557, March 10, 1979), but the final compounds prepared in accordance with the present invention have only a mild hypotensive effect during the initial period of administration and have little chance of developing orthostatic hypotensive asthenia. Medicines such as "Captopril" which has a free mercapto group causes various side effects due to the said mercapto group. These include disturbances in the sense of taste, production of albumin in the urine, agranulocytosis and dermal disorders accompanied by fever, which have been reported in Lancet, vol. 1, no. 8160, page 150 (January 19, 1980), supra, vol. 2, no. .8186, page 129

(19 July 1980) and South African Medical Journal, Vol 58, 172 (1980). On the other hand, the 1.0 ester bond in the final compounds prepared in accordance with the present invention is not subject to hydrolysis in vivo and is therefore unlikely to produce a mercapto group. The end products produced in accordance with the present invention therefore have little chance of causing the above-mentioned side effects due to the mercapto group.

The method in accordance with the invention for producing the end products is now described with the help of the following examples.

Example 1:

Preparation of N-substituted amino acids

(a) D-alanine (4.5 g) was dissolved in 230 mL aq

IN Na2CC>3 with stirring. To the solution was added dropwise at 5 - 10°C 100 ml of tetrahydrofuran containing 9.0 g of cyclohexanecarbonyl chloride and the mixture was stirred at this temperature for 30 min. and then stirred at room temperature for 1.5 hours. Then 2N HCl was added to the reaction mixture to adjust its pH to between 1 and 2. The reaction mixture was extracted with ethyl acetate and the organic layer was washed with saturated NaCl solution and dried over magnesium sulfate. The filtrate was evaporated under vacuum to give a crude compound. Recrystallization from ethyl acetate/n-hexane gave 4.65 g of N-cyclohexanecarbonyl-D-alanine. £' a3jT +26.6°.

(b) An ester of adamantane carboxylic acid and N-hydroxysuccinimide (3.0 g) was dissolved in 30 ml of tetrahydrofuran. To the solution was added water (5 ml) containing 0.89 g of D-alanine and 1.1 g of triethylamine was added and the mixture was stirred at 5°C overnight. After removal of tetrahydrofuran, water was added to the residue and 2N HCl was further added to the mixture to adjust its pH to between 1 and 2. The mixture was then treated as under (a) above to give 0.38 g of N-adamantane carbonyl -D-alanine. f~ g ^= +11.6°

(C=1.0 MeOH)

The compounds given in the following table were prepared by one or the other of the methods described under (a) and (b).

Example 2

(a) 3-frr-cyclohexanecarbonyl-D-alanylthio)-2D-methyl-propanoic acid

N-cyclohexanecarbonyl-D-alanine (5.97 g) was dissolved in

dry tetrahydrofuran (100 ml). To this solution was added carbonyldiimidazole (5.84 g) at between -20 and

-15°C and the mixture was stirred at this temperature for 1 hour. Then 3-mercapto-2-D-methylpropanoic acid (3.60 g) was added and the mixture was stirred at between -15

and -9°C for 1 hour and then at room temperature for another 1 hour. The mixture was evaporated under vacuum to

remove the solvent. To the residue was added water (40 ml)

added and 2N HCl was added to the mixture to adjust its pH to between 1 and 2. The mixture was extracted with ethyl acetate and the organic layer was washed with saturated NaCl solution twice and dried over magnesium sulfate. The filtrate was evaporated under vacuum to give a crude compound. Recrystallization from ethyl acetate/n-hexane gave a colorless prismatic crystal of the title compound (7.50 g, 83%).

Temp. 149 - 152°C

[α]D = +46.4° (C=.1.07, MeOH)

NMR (CD 3 OD, 6): 1 20 (3H, d), 1.35 (3H, d),

1.20 - 2.0 (11H, m), 2.40 - 2.80 (1H, m),

3.05 (2H, m), 4.50 (1H, m)

(b) 3-(N-cyclohexanecarbonyl-D-phenylglycylthio)-2-D-methylpropanoic acid

The procedure under (a) was repeated using N-cyclohexanecarbonyl-D-phenylglycine (2.61 g), carbonyldiimidazole (1.95 g) and 3-mercapto-2-D-methylpropanoic acid (1.20 g).

A yellow oil of the title compound

was prepared (2.07 g, 57%).

[α]D = -48.7° (C=1.19, MeOH)

NMR (CDCl 3 , 6): 1.24 (3H, d), 1.20 - 2.10 (11H, rn),

2.40 - 2.80 (1H, m) , 3.10 (2H,. m) , 5.75 (1H, d) ,

6.40 - 6.80 (1H, m), 7.50 (5H, S), 8.90 (1H, S)

(c) 3-(N-cyclohexanecarbonyl-D-leucylthio)-2-D-methyl-•propanoic acid

The procedure under (a) was repeated using N-cyclohexanecarbonyl-D-leucine (2.41 g), carbonyldiimidazole (1.95 g) and 3-mercapto-2-D-methylpropanoic acid (1.20 g). An oil of the title compound was prepared (2.40 g, 70%).

[oc]D = +12.2° (C=1.03, MeOH)

NMR (CDCl3, 6): 0.95 (6H, d), 1.28 (3H, d),

1.20 - 2.10 (11H, m), 2.40 - 2.80 (1H, m),

3.10 (2H, m), 4.70 (1H, m), 6.00 - 6.30 (1H, m),

9.20 (1H, S)

(d) 3-(N-cyclohexanecarbonyl-D-glutaminylthio)-2-D-methylpropanoic acid

The procedure under (a) was repeated using N-cyclohexanecarbonyl-D-glutamine (2.56 g), carbonyldiimidazole (1.95 g) and 3-mercapto-2-D-methyl-propanoic acid (1.20 g). A 0.70 g sample of the title compound was prepared (20%).

M.p. 146 - 149°C

[a]D= +10.8° (C=1.05, MeOH)

NMR (CD3OD, 6): 1.20 (3H, d), 1.20 - 2.10 (11H, m),

2.30 (4H, m), 2.40 - 2.80 (1H, m), 3.05 (2H, m),

4.50 (1H, m)

(e) 3-(N-cyclopropanecarbonylglycylthio)-2-D-methylpropanoic acid

The method under (a) was repeated using N-cyclopropanecarbonylglycine (0.78g), carbonyldiimidazole (1.06g) and 3-mercapto-2-D-methylpropanoic acid (0.66g). An oil of the title compound was formed (0.70 g, 52%).

[α]D = -33.8° (0=1.01, MeOH)

NMR (CDCl3, 6): 0.70 - 1.20 (4H, m) , 1.25 (3H, d) ,

1.30 - 1.80 (1H, m), 2.40 - 2.80 (1H, m), 3.14

(2H, m), 4.20 (2H, d), 7.30 (1H, S)

(f) 3-(N-adamantanecarbonylglycylthio)-2-D-methylpropanoic acid

The original method (a) was repeated using N-adamantanecarbonylglycine (2.38g), carbonyldiimidazole (1.95g) and 3-mercapto-2-D-methylpropanoic acid (1.20g). A 1.08 g sample of the compound mentioned in the title was formed (32%).

Temp. 132°C (decomposition)

[<Q>]D<=> -8.3° (C=1.01, MeOH)

NMR (CD3OD, 6): 1.25 (3H, d), 1.65 - 2.30 (15H, m),

2.40 - 2.80 (1H, m), 3.14 (2H, m), 4.06 (2H, d)' (g) 3-(N-cyclohexanecarbonyl-D-phenylalanylthio)-2-D-methylpropanoic acid

The method under (a) was repeated using N-cyclohexanecarbonyl-D-phenylalanine (1.37 g), carbonyldiimidazole (1.0 g) and 3-mercapto-2-D-methylpropanoic acid (0.60 g). An oil of the title compound was formed (1.5 g, 19%).

[a]D = +13.9° (C=1.02, MeOH)

NMR (CDCl 3 , 6): 1.25 (3H, d), 1.20 - 2.0 (10H, m),

2.1 - 3.0 (2H, m), 3.0 - 3.4 (4H, m), 4.95 (1H, t),

6.19 (1H, d), 7.24. (5H, S) , 10.12 (1H, S) ,

(h) 3-(N-cyclohexanecarbonyl-D-methionylthio)-2-D-methylpropanoic acid

The method under (a) was repeated using N-cyclohexanecarbonyl-D-methionine (0.325 g), carbonyldiimidazole (0.25 g) and 3-mercapto-2-D-methylpropanoic acid (0.15 g). An oil of the title compound was formed (0.29 g, 64%).

[a]D = -7.9° (C=1.0, MeOH)

NMR (CDCl 3 , 6): 1.26 (3H, d") , 1.2 - 2.0 (10H, m),

2.09 (3H, S), 2.0 - 3.2 (8H, m), 4.5 - 5.0 (1H, m),

6.62 (1H, d), 10.07 (1H, S)

(i) 3-(N-cyclohexanecarbonyl-D-tryptophyllthio)-2-D-methylpropanoic acid

The method under (a) was repeated using N-cyclohexanecarbonyl-D-tryptophan (0.157 g), carbonyldiimidazole (0.1 g) and 3-mercapto-2-D-methylpropanoic acid

(0.06g). A gummy sample of the title compound was prepared (0.17 g, 82%).

[a]D = -7.1° (C=1.0, MeOH)

NMR (CDCl3, «):' 1.26 (3H, d) , 1.2 '-"2.4 (11H, .m) ,

2.5 - 2.9 (1H, m), 3.0 - 3.3 (2H, m), 3.35 (2H, d),

<4.87> (1H, t), 6.4 - 6.9 (2H, wide s)f 6>9 _ 7 Q

(5H, m), 10.5 (1H, S)

Example 3

(a) N-(3-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methylpropanoyl/-L-proline

3-(N-cyclohexanecarbonyl-D-alanylthio)-2D-methyl-

propanoic acid (1.51 g) was dissolved in dry tetrahydrofuran (40 ml). Triethylamine (0.61 g) and ethyl chloroformate (0.65 g) were added to the solution at -5°C with stirring. Five minutes later, a solution containing L-proline (0.58 g) and triethylamine (0.61 g) dissolved in water (5 ml) was added and the mixture was stirred at 0°C for 1 hour and then at room temperature for 30 my. The mixture was evaporated under vacuum to remove the solvent. After adding water to the residue, 2N HCl was added to the mixture to adjust its pH to between 1 and 2. The reaction mixture was extracted with ethyl acetate and the organic layer was washed with saturated NaCl solution twice and dried over magnesium sulfate. The filtrate was evaporated under vacuum to remove ethyl acetate and the residue was subjected to chromatography on silica gel using a mixture of chloroform and methanol (100:1 to 100:2) as eluent. Fractions containing the final compound were collected and evaporated under vacuum to give a gummy sample of the title compound (0.3 g). (b) The same compound can be prepared by the following method.

3-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methylpropanoic acid (6.03 g) and N-hydroxysuccinimide (2.3 g) were dissolved in dry tetrahydrofuran (50 ml). To this solution was added dicyclohexylcarbodiimide (4.3 g) at between 0 and 5°C and the mixture was stirred overnight at this temperature. After the reaction, the precipitate was filtered off and insoluble matter was washed with a small amount of tetrahydrofuran. The filtrate and washing solutions were combined and evaporated under vacuum. To the residue was added ethyl acetate and the mixture was filtered. The ethyl acetate solution was washed with 0.5N HCl, water, aqueous Na2CO.j and saturated NaCl solution in the order mentioned. The solution was dried over magnesium sulfate, evaporated under vacuum, and the residue solidified by adding a mixture of ethyl acetate and hexane (1:10). The yield of solid product was 6.90 g (87%).

Temp. 113 - 116°C

Co- Cf +14.2° (C=1.05, MeOH)

NMR (CDCl 3 , $ ): 1.38 (6H, d), 1.20 - 2.20 (11H, m), 2.82 (4H, S), 2.8 - 3.30 (3H, m) 4.75 (1H, m),

6, 30 (1H, wide d)

The activated ester (3.98 g) was dissolved in tetrahydrofuran (40 ml). To the solution was added water (5 ml) with L-proline (1.15 g) dissolved therein and N-ethylmorpholine (1.26 ml)

was added and the mixture was stirred overnight. The mixture was evaporated under vacuum to remove the solvents and water was added to the residue. To the mixture was added 2N HCl to adjust its pH to between 1 and 2. The reaction mixture was extracted with ethyl acetate and the organic layer was washed with saturated NaCl solution and dried over magnesium sulfate. The filtrate was evaporated under vacuum. The residue was subjected to column chromatography on silica gel using a mixture of

chloroform and methanol (100:1 to 100:2) as eluent. The product obtained by evaporating the eluate (1.45 g) under vacuum was found to be identical to the title compound by NMR analysis. (c) The compound mentioned in the title can also be prepared using the following method. 3-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methylpropanoic acid (4.52 g) was dissolved in thionyl chloride (5 mL). To the mixture was added a drop of dimethylformamide and the mixture was stirred at room temperature overnight. The mixture was evaporated under vacuum to remove the solvents and to the residue dry toluene (2 ml) was added and the mixture was re-evaporated under vacuum to remove the solvent. To the residue was added dry tetrahydrofuran (10 ml) with stirring. The tetrahydrofuran solution containing the acid chloride was added dropwise at a temperature between 5 and 10°C with stirring to water (25 ml) with L-proline (2.3 g) and Na 2 CC> 3 (2.5 g) dissolved therein. The mixture was stirred at this temperature for 1 hour and at room temperature for 2 additional hours. The reaction mixture was subjected to extraction with chloroform. To the aqueous layer was added 2N HCl to adjust its pH to between 1 and 2. After extraction with chloroform, the chloroform layer was washed with saturated NaCl solution twice and dried over magnesium sulfate. After removal of chloroform, the residue was subjected to column chromatography on silica gel using a mixture of chloroform and methanol (100:1 to 100:2) as eluent. After evaporation under vacuum, a gummy substance (1.6 g) was obtained. This substance was found to be identical to the compound mentioned in the title by means of NMR analysis. (d) The compound mentioned in the title can also be prepared by means of a further method described in the following. 3-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methylpropanoic acid (3.01 g) and t-butyl ester of L-proline (1.71 g) were dissolved in dry dichloromethane (50 ml) and dicyclohexylcarbodiimide (2.15 g) was added to the solution while stirring and cooling with ice. The mixture was stirred at the same temperature for 1/2 hour and set aside overnight at 5°C. The reaction mixture was filtered and insoluble matter was washed with dichloromethane. The filtrate and washings were combined and washed with 1N HCl, water, 1N NaHCO-j and saturated NaCl solution in the order mentioned, dried over MgSO^ and evaporated under vacuum to give a gummy sample of the t-butyl ester of N- /3-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methyl-propanoyl7-L-proline (4.3 g). A 4.0 g sample of this ester was dissolved in anisole (30 mL) and trifluoroacetic acid (10 mL) was added to the solution. The mixture was stirred at room temperature for 1 hour and evaporated under vacuum to remove excess trifluoroacetic acid. The residue was subjected to column chromatography on silica gel (column: 2 x 35 cm) using a mixture of methanol and chloroform (1:100 to 3:100) as eluent. Fractions containing the final compound were collected and evaporated under vacuum to give a gummy substance (3.4 g). This substance was found to be identical to the compound mentioned in the title by means of NMR analysis and thin-layer chromatography.

The reaction components, the reaction solvents and the methods used for the preparation of the compound mentioned in the title in Example 3 are reproduced in the following table together with data for its physical properties. Other compounds were prepared using one of the methods described in Example 3 (a) to (d) and details of the specific method are given in the following table together with data for the physical properties of the final compounds. In the table, the numbers following the numbers after (1) and (2) in the "Rf value" column indicate the Rf values obtained by the thin layer chromatography using CHCl^/MeOH/AcOH (2:1:0.003) and n-BuOH/AcOH/H 2 O (4:1:1) as respective eluents.

Example 4

Various salts were prepared from the samples of N-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methylpropanoylZ-L-proline prepared in Examples 3 and 5.

(a) Ca salt

A 3.98 g sample of the compound prepared in Example 3* or 5 was dissolved in methanol (40 ml). To this solution was added calcium acetate hydrate (0.84 g) and the mixture was stirred under reflux for 1 hour. Insoluble substance was filtered off and the filtrate was evaporated under vacuum. To the residue was added chloroform and the mixture was filtered and evaporated under vacuum. To the residue was added diethyl ether and the mixture was filtered and dried with air to give a calcium salt of the compound of Example 3 or 5 (3.40 g). .2°

(C=1.0, MeOH)

(b) Mg salt

The method under (a) was repeated using 3.09 g of the compound prepared in Example 3 or 5 and 0.772 g of magnesium acetate tetrahydrate. A magnesium salt of the compound of Example 3 or 5 was formed (2.42 g).

£aj= -46.6° (C=1.0, MeOH)

(c) Lysine salt

A 1.19 g sample of the compound prepared in Example 3 or 5 was dissolved in methanol (22 ml). To this solution was added lysine (0.416 g) and the mixture was stirred at room temperature for 1 hour and evaporated under vacuum. To the residue was added chloroform and the mixture became

filtered and evaporated under vacuum. To the residue was added dimethyl ether and the product was filtered off to give a lysine salt of the compound in example 3 or 5

(1.54g) . £VJ£= -23.1° (C=1.0, MeOH)

(d) Na salt

A 2.27 g sample of the compound prepared in Example 3 or 5 was dissolved in methanol (25 ml). To this solution 0.514 g of sodium acetate was added and the mixture was stirred at room temperature for 30 min. and evaporated under vacuum. To the residue was added 200 ml of a mixture of methanol and chloroform (3:100 v/v) and the resulting mixture was filtered and evaporated under vacuum. The residue was dissolved in methanol and the solution was filtered and evaporated under vacuum. Ethyl acetate was added to the residue and the product was filtered off to give a Na salt of the compound of Example 3 or 5

(1.85g) . £V|f -26.7° (C=1.0, MeOH)

(e) Dicyclohexylamine salt

A 13.1 g sample of the compound from Example 3 or 5 was dissolved in 120 ml of acetonitrile. To the solution was added dicyclohexylamine (6 ml) with stirring. The mixture was stirred further for 30 min. and then put away overnight. The precipitate was filtered off with suction and dried in air. The resulting crude crystals were suspended in 300 ml of acetonitrile and the suspension was heated under reflux for 30 min. The suspension was cooled and the crystals were filtered with suction and dried in air to give a dicyclohexylamine salt of the compound of Example 3 or 5 (12.2 g). /"o7=-24.5° (C=1.0, MeOH)

^D

Example 5: N-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methylpropanoyl7-L-proline

N-cyclohexanecarbonyl-D-alanine (5.98 g) was dissolved in dry tetrahydrofuran (80 mL). Carbonyldiimidazole (5.84 g) was added at -18° with stirring to the solution while the mixture was cooled with ice. The mixture was stirred at this temperature for 1 hour and then N-(3-mercapto-2-D-methylpropanoyl)-L-proline (6.29 g) was added to the mixture followed by stirring at -18°C for 30 min. and then at room temperature for 1 hour. After completion of the reaction, the mixture was evaporated under vacuum to remove the solvent. To the residue was added water (50 mL) and 2N HCl was added to the mixture to adjust its pH to between 1 and 2. The mixture was extracted with ethyl acetate and the ethyl acetate layer was washed with saturated NaCl solution, dried over MgSO 4 and evaporated under vacuum . To the residue was added acetonitrile (120 ml) and then 6 ml of dicyclohexylamine (DCHA). The mixture was stirred at room temperature for 1 hour. After standing overnight, the precipitate was filtered off and dried in air to give a crude DCHA salt (14.35 g).

The crude salt was suspended in acetonitrile (300 mL) and the suspension was heated under reflux for 30 min. After cooling, the precipitate was collected by suction and dried in air to give a white DCHA salt (12.20 g). The DCHA salt (12.20 g) was suspended in ethyl acetate (90 ml) and to the suspension was added 0.5 N aqueous KHSO 4 (60 ml) and the mixture was shaken. The organic layer was washed with distilled water, dried over MgSO 4 and evaporated in vacuo to give a gummy substance (8.64 g). This substance was found to be identical to the compound mentioned in the title by means of NMR analysis and thin-layer chromatography.

Example 6: N-(N-cyclohexanecarbonyl-glycylthio)-2-D-methylpropanoyl7-L-proline

N-Cyclohexanecarbonylglycine (1.02 g) was dissolved in dry tetrahydrofuran (10 mL). To the solution was added 20 ml of dry tetrahydrofuran containing 1.07 g of carbonyldiimidazole at -20°C with stirring while the mixture was cooled with ice. The mixture was stirred at this temperature for 1 hour and then 6 ml of dry tetrahydrofuran containing 1.09 g of N-(3-mercapto-2-D-methyl-propanoyl)-L-proline was added followed by stirring at -20 C for 30 my. and then at room temperature for 1 hour. After completion of the reaction, the mixture was evaporated under vacuum to remove the solvent. The residue was subjected to column chromatography on silica gel (column: 2x35 cm) using a mixture of methanol and chloroform (1:100 to 3:100) as eluent. Fractions containing the final compound were combined and evaporated under vacuum to give a gummy substance (1.16 g). The substance was found to be identical to the compound mentioned in the title by means of NMR analysis and thin-layer chromatography.

Example 7: N-(N-cyclopropanecarbonyl-D-alanylthio)-2-D-methylpropanoyl/-L-proline

N-cyclopropanecarbonyl-D-alanine (635 mg) and triethylamine (0.70 mL) were dissolved in dry tetrahydrofuran (14 mL).

To the solution was added ethyl chloroformate (0.48 mL) at -15°C with stirring while the mixture was cooled with ice. The mixture was stirred at this temperature for 15 min. and then N-(3-mercapto-2-D-methylpropanoyl)-L-proline (1.09 g) and dry tetrahydrofuran (10 ml) containing triethylamine (0.70 ml) were added followed by stirring at -15°C for 15 min. and then at 5°C overnight. After completion of the reaction, the mixture was evaporated under vacuum at 30 to 35°C to remove the solvent. To the residue was added water (10 mL) and 2N HCl was added to the mixture to adjust its pH to between 1 and 2. The mixture was extracted with chloroform and the chloroform layer was washed with saturated NaCl solution, dried over MgSO 4 , and evaporated under vacuum . The residue was subjected to column chromatography on silica gel (column: 2 x 35 cm) using a mixture of methanol and chloroform (1:100 to 3:100) as eluent. Fractions containing the final compound were combined and evaporated under vacuum to give a gummy substance (395 mg). This substance was found to be identical to the compound mentioned in the title by NMR analysis and thin layer chromatography.

Example 8: N-(3-(N-cyclohexanecarbonyl-N-methylglycyl-thio)-2-D-methylpropanoyl7-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonyl-glycine was replaced with N-cyclohexanecarbonyl-N-methylglycine (1.09 g), carbonyldiimidazole (1.09 g) and N-(3-mercapto-2-D -methylpropanoyl)-L-proline (1.09 g). A gummy substance (0.76 g) was formed and this was found to be identical to the title compound by NMR analysis and thin layer chromatography.

Example 9: N-(3-(N-cyclopropanecarbonylglycylthio)-2-D-methylpropanoyl/-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonylglycine was replaced with N-cyclopropanecarbonylglycine (0.96 g), carbonyldiimidazole (1.09 g) and N-(3-mercapto-2-D-methylpropanoyl)- L-proline (1.09 g). A gummy substance (0.4 g) was formed and this was found to be identical to the title compound by NMR analysis and thin layer chromatography.

Example 10: N-(3-(N-adamantanecarbonylglycylthio)-2-D-methylpropanoyl7-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonylglycine was replaced with N-adamantanecarbonylglycine (1.42 g), carbonyldiimidazole (1.09 g) and N-(3-mercapto-2-D-methylpropanoyl)- L-proline (1.09 g). A gummy substance (1.7 g) was formed and this was found to be identical to the title compound by NMR analysis and thin layer chromatography.

Example 11: N-(N-cyclohexanecarbonyl-D-phenylalanylthio)-2-D-methylpropanoyl7-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonyl-glycine was replaced with N-cyclohexanecarbonyl-D-phenylalanine (1.54 g), carbonyldiimidazole (1.09 g) and N-(3-mercapto-2-D -methylpropanoyl)-L-proline (1.09 g). A gummy substance (0.97 g) was formed and this was found to be identical to the title compound by NMR analysis and thin layer chromatography.

Example 12: N-(3-(N-cyclohexanecarbonyl-D-leucylthio)-2-D-methylpropanoyl/-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonyl-glycine was replaced with N-cyclohexanecarbonyl-D-leucine (1.20 g), carbonyldiimidazole (0.97 g) and N-(3-mercapto-2-D -methylpropanoyl)-L-proline (0.98 g). A gummy substance (1.17 g) was formed and this was found to be identical to that in

the title compound by NMR analysis

and thin-layer chromatography.

Example 13: N-(3-(N-cyclohexanecarbonyl-D-tryptophyllthio)-2-D-methylpropanoyl/-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonyl-glycine was replaced with N-cyclohexanecarbonyl-D-tryptophan (1.75 g), carbonyldiimidazole (1.09 g) and N-(3-mercapto-2-D -methylpropanoyl)-L-proline (1.09 g). A gummy substance (0.8 g) was formed and this was found to be identical to the compound mentioned in the title by NMR analysis and thin-layer chromatography.

Example 14: N-(3-(N-cyclohexanecarbonyl-D-phenylglycyl-thio)-2-D-methylpropanoyl7-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonyl-glycine was replaced with N-cyclohexanecarbonyl-D-phenylglycine (1.30 g), carbonyldiimidazole (0.97 g) and N-(3-mercapto-2-D -methyl-propanoyl)-L-proline (0.98 g). A gummy substance (0.35 g) was formed and this was found to be identical to the title compound by NMR analysis and thin layer chromatography.

Example 15: N-(3-(N-cyclohexanecarbonyl-D-methionylthio)-2-D-methylpropanoyl/-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonyl-glycine was replaced with N-cyclohexanecarbonyl-D-methionine (1.30 g), carbonyldiimidazole (1.09 g) and N-(3-mercapto-2-D -methyl-propanoyl)-L-proline (1.09 g). A gummy substance (0.59 g) was formed and this was found to be identical to the title compound by NMR analysis and thin layer chromatography.

Example 16: N-(3-(N-cyclohexanecarbonyl-D-glutaminyl-thio)-2-D-methylpropanoyl/-L-proline

The method in example 6 was repeated with the exception that N-cyclohexanecarbonyl-glycine was replaced with N-cyclohexanecarbonyl-D-glutamine (0.5 g), carbonyldiimidazole (0.41 g) and N-(3-mercapto-2-D -methylpropanoyl)-L-proline (0.41 g). A gummy substance (0.3 g) was formed

and this was found to be identical to the compound mentioned in the title by means of NMR analysis and thin-layer chromatography.

Example 17: N-(N-adamantanecarbonyl-D-alanylthio)-2-D-methylpropanoyl-17-L-proline

The method in Example 6 was repeated with the exception that N-cyclohexanecarbonyl-glycine was replaced with N-adamantanecarbonyl-D-alanine (1.2 g), carbonyldiimidazole (0.97 g) and N-(3-mercapto-2-D -methylpropanoyl)-L-proline (0.98 g). A gummy substance (1.02 g) was formed and this was found to be identical to that in

■the title compound by means of NMR analysis and thin-layer chromatography.

Example 18: Dicyclohexylamine salt of N-(3-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methyl-propanoyl/-L-proline

(1) 3-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methyl-propanoic acid (241 g) was suspended in dry dichloromethane

(1.3 liters). To the suspension triethylamine (117 ml) was added dropwise at -15°C with stirring and then pivaloyl chloride (104 ml) was added dropwise at a temperature below -5°C with stirring and the mixture was further stirred for 30 min.

(2) L-proline (101 g) was suspended in dry dichloromethane (1.2 liters). To this suspension trimethylsilyl chloride (112 ml) was added dropwise at -15°C with stirring and then triethylamine (122 ml) was added dropwise at a temperature below -5°C with stirring, and the mixture was further stirred at -15°C for 30 min. To the resulting mixture, the solution prepared under (1) was added and the mixture was stirred under cooling for 15 min. and then at room temperature for 1 hour. After completion of the reaction, cold distilled water (1 liter) was added to the reaction mixture under cooling and then concentrated hydrochloric acid (90 ml) was added dropwise. The mixture was stirred for 10 min. and the organic layer was separated, washed with saturated NaCl solution twice and dried over magnesium sulfate. Until the filtrate became

dicyclohexylamine (326 ml) and acetonitrile (800 ml)

added and the mixture was stirred for 30 min. Then four 800 ml portions of acetonitrile were added at intervals of 10 min. and the resulting mixture was allowed to stand overnight at 5°C. The precipitate was suctioned off on a filter and dried in air to give a crude sample of the compound mentioned in the title. The crude sample was dissolved in hot dichloromethane

(3 liters) and filtered. The filtrate was stirred at room temperature for 30 min. and four further 750 ml portions of acetonitrile were added at 10 min intervals. and the resulting mixture was allowed to stand overnight at 5°C. The precipitate was filtered off and dried in air to give a colorless sample of the title compound

(404g).

Temp. 190 - 191°C ( JZp3 = -24.5° (C=1.0, MeOH)

D

Example 19: Calcium salt of N-(3-(N-cyclohexanecarbonyl-D-alanylthio)-2-D-methylpropanoyl/-L-proline

A 404 g sample of the dicyclohexylamine salt prepared in Example 18 was suspended in ethyl acetate (2 liters) and to the suspension was added 0.5N KHSO 4 (2.5 liters) and the mixture was stirred for 10 min. The organic layer was separated, washed with distilled water and dried over magnesium sulfate. The filtrate was evaporated under vacuum and the residue was dissolved in acetone (1.2 liters). To the acetone solution, distilled water (1.2 liters) and calcium carbonate (36.6 g) were added and the mixture was stirred vigorously at 40°C for 1 hour. After cooling, the mixture was filtered and the filtrate was evaporated under vacuum. The resulting syrup was suspended in acetone (1 liter) and the solution was stirred for 5 hours. The precipitate was suction filtered, dried in air, then vacuum dried to give a white powder of the compound mentioned in the title (212 g).

OH. = -47.2° (C=1.0, MeOH)

D

Claims (1)

Analogous process for the preparation of therapeutically active L-proline derivatives of formula (I) in which R is an acyl group attached to the o(-amino group of an amino acid and is selected from the group consisting of cyclopropanecarbonyl, cyclohexanecarbonyl and adamantane carbonyl, A is a glycine, sarcosine or o-D-amino acid residue with an oC-carbonyl group which forms a thiol ester bond with the sulfur atom (S), and in which the 2-methyl-propanoyl part has the D configuration, or pharmaceutically acceptable salts thereof, characterized in that a) either a compound is reacted with formula (II) in which A' is a glycine, sarcosine or a protected or unprotected oC-D amino acid residue, and R has the above meaning, or a reactive derivative thereof, with a compound of formula (III) in which R' is a hydrogen atom or a carboxyl protecting group or a reactive derivative thereof, in an inert organic solvent at a temperature between -50 and 20°C, any protecting group is removed from the reaction product and, if desired, the reaction product is converted into a pharmaceutically acceptable salt, or b) a compound of formula (IV) in which A' is a glycine, sarcosine or a protected or unprotected 0<_-D-amino acid residue, and R has the above meaning, or a reactive derivative thereof, is reacted with a compound of formula (V) in which R' is a hydrogen atom or a carboxyl protecting group or a reactive derivative thereof, in an inert organic solvent at a temperature between -50 and 20°C, any protecting group is removed from the reaction product and, if desired, the reaction product is converted into a pharmaceutically acceptable salt.