NO148416B - ANALOGY PROCEDURE FOR THE PREPARATION OF THERAPEUTICALLY ACTIVE TIAZOLIDINE, MORPHOLINE AND TIAZAN DERIVATIVES - Google Patents

Description

This invention relates to a process for the production of new thiazolidine, morpholine and thiazan derivatives with the general formula

and basic salts thereof, where

R 1 and R 2 are each hydrogen or lower alkyl;

is hydrogen or lower alkyl,

R 4 is hydrogen, lower alkanoyl or benzoyl,

X is 0, S, SO or SO 2 , m being 2 and n being 1 when X is 0;

m is 1 or 2,

n is 0, 1 or 2 and

m + n is 2 or 3;

p is 0 or 1.

The stars denote centers of asymmetry.

The lower alkyl groups designated by one of the symbols include linear and branched hydrocarbon radicals from methyl to heptyl, e.g. methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl and isopentyl. The C-^-C^ groups, especially the C^- and C2- groups of all types are preferred.

The lower alkanoyl groups are the acyl radicals of the lower (up to 7 carbon atoms) fatty acids, e.g. acetyl, propionyl, butyryl and the like, acetyl being preferred.

The symbols have the meanings given above throughout the description.

Particular preference is given to those compounds of formula I where and R 2 is particularly hydrogen, methyl or ethyl,

especially hydrogen; R, is in particular hydrogen, methyl or ethyl; R 4 is in particular hydrogen, acetyl or benzoyl; X is sulfur or oxygen, especially sulfur; m is 1 or 2; n is 1 and p is 1.

The compounds of formula I and the preferred subgroups can be prepared by various synthesis methods.

According to the invention, the acid is acylated with the formula

where the symbols have the meanings given above, with an acid of the formula

by a known method in which the acid III is activated prior to reaction with the acid II, including formation of a mixed anhydride, symmetrical anhydride, acid chloride, active ester, Woodward's reagent K, N,N'-carbonylbisimidazole, EEDQ (N-ethoxy-carbonyl-2 -ethoxy-1,2-dihydroquinoline) or the like.

The acid of formula II can of course be acylated stepwise. E.g. a fragment of the acylating agent III can first be attached to the acid of formula II, e.g. by reacting the acid with a halogen-acyl-halide of the formula: where Hal means a halogen atom, preferably chlorine or bromine, e.g. the compound 3-bromopropanoyl chloride. This leads to a product with the formula:

Reaction of this intermediate with a thiol R 2 -SH then leads to the desired product of formula I. This stepwise acylation is illustrated in the examples.

Optionally, a compound of formula I is hydrolyzed where

R 4 is different from hydrogen, to form a compound where R 4 is hydrogen,

and/or optionally a compound of formula I where X is S is oxidized to a compound where X is SO or SO 2 , or a compound where X is SO to a compound where X is SO 2 .

The products with formula I have at least 1, and can have up to

4 asymmetric carbon atoms. These carbon atoms are denoted by an asterisk in formula I. The compounds thus exist in diastereoisomeric forms or in racemic mixtures thereof. All of these can be produced according to the invention. The syntheses described above can use the racemate or one of the enantiomers as starting material. When racemic starting material is used in the synthesis, the stereoisomers obtained in the product can be separated by chromatography or fractional crystallization in the usual way. In general, the L-isomer with respect to the carbon atom of the amino acid is the preferred isomeric form.

The compounds produced according to the invention form basic salts with various inorganic and organic bases, which are also produced according to the invention. Such salts include ammonium salts, alkali metal salts such as sodium and potassium salts (which are preferred), alkaline earth metal salts such as calcium and magnesium salts, salts with organic bases, e.g. 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 other salts are also useful, e.g. for isolation or purification of the product, as is the case with the dicyclohexylamine salt.

The salts are formed in the usual way by reacting the free acid form of the product with one or more equivalents of the appropriate base which provides the desired cation in a solvent or medium in which the salt is insoluble, and filtering, or in water, and the water is removed by freeze-drying . By neutralizing the salt with an insoluble acid such as

a cation exchange resin in hydrogen form (e.g. polystyrene sulphonic acid resin - "Dowex 50" (Mikes, Laboratory Handbook of Chromatographic Methods, Van Nostrand, 1961) page 256) or with an aqueous acid and extraction with an organic solvent, e.g. ethyl acetate, dichloromethane or the like, the free acid form can be obtained, and optionally another salt can be formed.

Additional experimental details are found in the examples which represent preferred embodiments and which also serve as models for the preparation of other members of the group.

The new compounds produced according to the invention inhibit the conversion of the decapeptide angiotensin I to angiotensin II and are therefore useful in reducing or alleviating angiotensin-associated hypertension. The action of the enzyme renin on angiotensinogen, a pseudoglobulin in blood plasma, produces angiotensin I. Angiotensin I is converted by angiotensin-converting enzyme (ACE) to angiotensin II. The latter is an active pressor substance which has been suggested to be the substance that induces different forms of hypertension in different mammalian species, e.g. rats, dogs, etc. The compounds produced according to the invention intervene in the angiotensinogen -»■ angiotensin I •> angiotensin II process by inhibiting the angiotensin-converting enzyme and reducing or eliminating the formation of the pressor substance angiotensin II.

Inhibition of the angiotensin-converting enzyme by compounds of formula I can be measured in vitro with isolated angiotensin-converting enzyme from rabbit lungs following the method described by Cushman and Cheung (Biochem. Pharmacol., 20, 1637 (1971)), and by examining on a

excised smooth muscle (E. 0'Keefe, et al., Federation Proe.

31, 511 (1972)), where these compounds have been shown to be strong inhibitors of the contractile action of angiotensin I and potentiators of the contractile activity of bradykinin.

Administration of a composition containing one or a combination of the compounds of formula I or a physiologically acceptable salt thereof to various species of hypertensive mammals alleviates or reduces angiotensin-associated hypertension. A single dose, or preferably two to four doses daily, given on the basis of approx. 5 to 1000 mg per kg per day, preferably approx. 10 to 500 mg per kg per day, is <p>assent to reduce blood pressure. The animal model experiments described by S. L. Engel, T. R. Schaeffer, M.H. Waugh and B. Rubin, Proe. Soc. Exp. Biol. With. 143, 483 (1973) serves

as a useful guide.

The active compound is preferably administered orally, but parenteral administration, such as subcutaneous, intramuscular, intravenous or intraperitoneal, can also be used.

The compounds produced according to the invention can be used to achieve a reduction of blood pressure by preparation in preparations such as tablets, capsules or elixirs for oral administration or in sterile solution or suspension for parenteral administration. About. 10 to 500 mg of an active compound or a mixture of compounds of formula I

or a physiologically acceptable salt thereof is mixed with a physiologically acceptable carrier, excipient, binder, preservative, stabiliser, flavoring etc., in unit dose form as required by assumed pharmaceutical practice. The amount of active substance in these preparations is such that a suitable dose is achieved in the specified range.

The following examples, which represent preferred embodiments, shall serve to further illustrate the invention. All temperatures are in °C.

Example 1

3-(3-benzoylthiopropanoyl)-4-L-thiazolidinecarboxylic acid

To a solution of L-4-thiazolidinecarboxylic acid (6.6 g)

in normal sodium hydroxide (50 ml) cooled in an ice bath, add 2N sodium hydroxide (25 ml) and 3-bromopropionyl chloride (8.5 g)

in this order with vigorous stirring. After 3 hours, a suspension of thiobenzoic acid (7.5 g) and potassium carbonate (4.8 g) in water (50 ml) is added. The reaction mixture is stirred overnight at room temperature and filtered. The filtrate is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is dried and concentrated to dryness. The residue is purified by silica gel chromatography (benzene:acetic acid, 7:1), and the purified material is crystallized from ethyl acetate-ether-hexane to give 3-(3-benzoylthiopropanoyl)-4-L-thiazolidinecarboxylic acid, m.p. 105-106°C.

This product is dissolved in water, and an equivalent amount of sodium hydroxide solution is added. The solution is then freeze-dried to give the sodium salt.

Example 2

3-( 3- mercaptopropanoyl)- L- 4- thiazolidine carboxylic acid

3-(3-Benzoylthiopropanoyl)-L-4-thiazolidinecarboxylic acid

(6.7 g) is dissolved in a mixture of water (15 ml) and concentrated ammonia (7.5 ml) under a blanket of argon. After 1 hour of storage at room temperature, the reaction mixture is diluted with water (20 ml) and filtered. The filtrate is extracted with ethyl acetate, acidified with concentrated hydrochloric acid and re-extracted with ethyl acetate. The second ethyl acetate extract is dried and concentrated to dryness. The residue, 3-(3-mercaptopropanoyl)-L-4-thiazolidinecarboxylic acid, is crystallized from ethyl acetate, m.p. 110-112°C.

Example 3

a) 3-acetylthio-2-methylpropanoic acid (starting material)

A mixture of thioacetic acid (50 g) and methacrylic acid (40.7 g)

heated on a steam bath for 1 hour and then stored at room temperature for 18 hours. The reaction mixture is distilled in vacuum, and the fraction with b.p. _ c min 128.5-131° is collected.

2. .6 mm L

The 3-acetylthio-2-methylpropanoic acid can also be isolated by

allow the reaction mixture to crystallize after dilution with hexane, m.p. 40-42°.

b) 3-(3-Acetylthio-2-methylpropanoyl)-2-thiazolidinecarboxylic acid methyl ester

2- Thiazolidine carboxylic acid methyl ester (CA. 53i, 12.281d)

(4.4 g) and 3-hydroxybenzotriazole (4.0 g) are dissolved in dichloromethane (40 ml), and the solution is stirred and cooled in an ice bath. Dicyclohexylcarbodiimide (6.2 g) dissolved in dichloromethane (15 ml) is added, immediately followed by a solution of 3-acetylthio-2-methylpropanoic acid (4.9 g) in dichloromethane (5 ml).

After stirring for 15 minutes in an ice bath and 16 hours at room temperature, the precipitate is filtered off, and the filtrate is washed neutrally. The organic layer is dried and concentrated to dryness in vacuo to give 3-(3-acetylthio-2-methylpropanoyl)-2-thiazolidinecarboxylic acid methyl ester. c) 3-(3-mercapto-2-methylpropanoyl)-2-thiazolidinecarboxylic acid 3-(3-acetylthio-2-methylpropanoyl)-2-thiazolidinecarboxylic acid methyl ester (2.9 g) is dissolved in methanol (30 ml) and N sodium hydroxide (30 ml) is added. The reaction mixture is stirred at room temperature, portions are taken out every hour and examined by paper electrophoresis after hydrolysis of the methyl ester. When this hydrolysis is complete (about 3 hours), the reaction mixture is neutralized, concentrated in vacuo to remove methanol, acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is dried and concentrated to dryness to give 3-(3-mercapto-2-methylpropanoyl)-2-thiazolidinecarboxylic acid, m.p. 96-101°C.

Calculated for CgH^<NO>^<:> C 40.83, H 5.57, N 5.95, S 27.25 Found: C 40.45, H 5.38, N 5.73, S 26, 97 SH titration 99.4%.

Example 4

3-( 3- mercaptopropanoyl)- 1, 3- thiazane- 4- carboxylic acid

By using 1,3-thiazan-4-carboxylic acid (J. Biol. Chem., 607 (1957) instead of 4-L-thiazolidinecarboxylic acid in the method according to example 1 and then subjecting the product to the method according to example 2, one obtains 3-(3 -benzoylthio-propanoyl)-1,3-thiazine-4-carboxylic acid and 3-(3-mercapto-propanoyl)-1,3-thiazine-4-carboxylic acid.

The final product (3-mercapto compound) was obtained as a pale yellow viscous syrup with an R^ of 0.32 (silica gel - 7:1 benzene:acetic acid) and an elemental analysis as follows: Calculated for CgH13N03S2: C 40.83, H 5.57, N 5.95, S 27.25 Found: C 40.96, H 5.54, N 6.02, S 26.94.

Example 5

3-( 3- mercapto- 2- methylpropanoyl)- L- 4- thiazolidine carboxylic acid

3-acetylthio-2-methylpropanoic acid chloride (5.4 g, prepared from 3-acetylthio-2-methylpropanoic acid and thionyl chloride, b.p. 80 * o) and 2N sodium hydroxide (15 ml) are added to a solution of L-4-thiazolidinecarboxylic acid (Z. Naturforschg., 17b, 765 (1962))

(5.2 g) in normal sodium hydroxide (30 ml) cooled in an ice-water bath. After stirring for 3 hours at room temperature, the mixture is extracted with ether, the aqueous phase is acidified and extracted with ethyl acetate. The organic phase is dried over magnesium sulfate and concentrated to dryness in vacuo to give 3-(3-acetylthio-2-methylpropanoyl)-L-4-thiazolidinecarboxylic acid.

3-(3-Acetylthio-2-methylpropanoyl)-L-4-thiazolidinecarboxylic acid (1 g) is dissolved in a mixture of water (3 ml) and concentrated ammonia (3 ml) under a blanket of argon. The mixture is stirred at room temperature for 30 minutes and acidified with concentrated hydrochloric acid. The organic layer is dried and concentrated to dryness in vacuo to give 3-(3-acetylthio-2-methyl-propanoyl)-L-4-thiazolidinecarboxylic acid (dicyclohexylammonium salt crystallized from acetonitrile, m.p. (synth. 130°) 172- 186°) and 3-(3-mercapto-2-methylpropanoyl)-L-4-thiazolidinecarboxylic acid (dicyclohexylammonium salt crystallized from ethyl acetate-hexane, m.p. (synth. 170°) 180-188°).

Example 6

4- ( 3- acetylthiopropanoyl)- L- 1, 4- thiazane- 5- carboxylic acid

L-4-thiomorpholine-3-carboxylic acid hydrochloride (6.6 g,

0.036 m) is dissolved in 150 ml of dimethylacetamide, and 3-acetylthio-

propanoyl chloride (5.97 g, 0.036 m) is added. The temperature rises to 28°. To this solution is added N-methylmorpholine (10.9 g, 0.108 m). The temperature rises to 42°, and a white precipitate immediately forms. The mixture is heated on a steam bath for 1 hour and allowed to stand overnight at room temperature. The solid is filtered off to give 9.7 g of 4-(3-acetylthiopropanoyl)-L-4-thiazane-5-carboxylic acid, m.p. 202-204°. The solvent is removed to give a viscous residue which is rubbed out with water and 20% hydrochloric acid. The precipitated oil is extracted with 3 x 150 ml of ethyl acetate, and the extracts are dried over magnesium sulphate. The solvent is removed and the viscous residue (7.5 g) crystallizes on standing. After recrystallization from acetone-hexane, the product has a constant melting point at 122-125°.

Example 7

4-( 3- mercaptopropanoyl)- L- 1, 4- thiazane- 5- carboxylic acid

Aqueous ammonia (13 ml of concentrated ammonium hydroxide

in 30 ml of water) is stirred under nitrogen for 15 minutes, and solid 4-(3-acetylthiopropanoyl)-L-1,4-thiazane-5-carboxylic acid (6.8 g,

0.024 m) is added. A clear solution forms quickly at 5-10°. The solution is stirred at room temperature under nitrogen for 1 hour. The solution is extracted with 100 ml of ethyl acetate, and the aqueous

layer is made strongly acidic with 20% hydrochloric acid. The precipitated oil is extracted with 3 x 150 ml of ethyl acetate. The extracts are combined and dried over magnesium sulfate, and then the solvent is removed to give 5.6 g of semi-crystalline mass which is found to contain a significant amount of starting material. The recovered material (5.6 g) is again hydrolyzed as above with 12 ml of concentrated ammonium hydroxide in 25 ml of water for a further 2

hours. This solution is acidified, and the precipitated oil is extracted with 3 x 150 ml of ethyl acetate. The extracts are combined and dried over magnesium sulfate, and then the solvent is removed to give 2.7 g (48%) of 4-(3-mercaptopropanoyl)-L-1,4-thiazane-5-carboxylic acid as a viscous mass after drying overnight at room temperature and 1 mm Hg.

Analysis:

Calculated for CgH-^NO^: N 5.95, C 40.82, H 5.56, S 27.25, SH 100% Found: N 6.13, C 40.85, H 5.46, S 27 .38, SH 96%.

Example 8

3-( 3- mercapto- 2- D- methyl- l- oxopropyl)- L- 4- thiazolidine carboxylic acid- l- oxide

(a) 3-(3-Acetylthio-2-D-methyl-1-oxopropyl)-L-4-thiazolidine carboxylic acid-1-oxide

2.76 g of 3-(3-acetylthio-2-D-methyl-1-oxopropyl)-L-4-thiazolidinecarboxylic acid was introduced into 20 ml of methylene chloride and 1.84 ml of formic acid with stirring at room temperature. 1.04 ml of 30% hydrogen peroxide was added to this mixture. After 3.5 hours, the mixture was concentrated to dryness in vacuo, refluxed with ethyl acetate, filtered and concentrated to dryness in vacuo to give 2.5 g of product.

(b) 3-(3-mercapto-2-D-methyl-1-oxopropyl)-L-4-thiazolidinecarboxylic acid-1-oxide

The material from part (a) was treated for 20 minutes with a cold solution of water (10 ml) and concentrated ammonium hydroxide (10 ml), acidified with AG50WX2 resin and chromatographed on AG50WX2 resin to give 1 g of product as a hygroscopic lyophilisate. , optical rotation [o]^ -183.1°

(c = 1.42, methanol).

Analysis for CgH^NO^ • 1/2 H20

Calculated: C 36.95, H 5.43, N 5.39, S 24.66

Found: C 36.94, H 5.45, N 5.45, S 24.82

Example 9

3-( 3- mercaptol- l- oxopropyl)- 2- methyl- L- 4- thiazolidine carboxylic acid

3-(Acetylthio)propionyl chloride (9.0 g, 0.07 m) was added slowly to a solution of 2-methyl-L-4-thiazolidinecarboxylic acid (8.0 g, 0.0544 m) and sodium carbonate (2.9 g) in 70 ml of distilled water, the pH value being kept at 9 at 6°C by the simultaneous addition of 24% sodium carbonate solution. The slurry was stirred for 30 minutes in an ice bath, after which 70 ml of water was added to give a clear solution. Stirring was continued at room temperature for an additional 1 hour, after which, after extraction twice with ethyl acetate (discarded) and cooling to 6-7°C, the mixture was acidified with concentrated hydrochloric acid to

pH 2.5. The reaction mixture was then saturated with sodium chloride and extracted twice with ethyl acetate. The ethyl acetate extracts were dried over magnesium sulfate and evaporated to dryness to give 13.32 g of 3-(3-acetylthio-1-oxopropyl)-2-methyl-L-4-thiazolidinecarboxylic acid which was then converted to the dicyclohexylamine salt, crystallized from isopropanol and converted to the free acid which, after crystallization from ethyl acetate-hexane, melted at 119-122°C.

The above product was added to a mixture of 12 ml of water and 8 ml of ammonium hydroxide in an ice bath and then stirred for 1 hour at room temperature, after which 40 ml of water was added, and the mixture was extracted twice with ethyl acetate which was discarded. The aqueous mixture was then cooled, acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The ethyl acetate layer was dried over magnesium sulfate and concentrated to dryness in vacuo to give 2.3 g of an oil which was then stirred in hexane for 72 hours to give 1.75 g of 3-(3-mercapto-1-oxopropyl)-2 -methyl-L-4-thiazolidinecarboxylic acid as a colorless material, m.p. 89-95°C.

Analysis for C8H13S2N03

Calculated: C 40.83, H 5.57, S 27.25, N 5.95

Found: C 40.96, H 5.60 S 27.39, N 5.89.

Examples 10 and 11

D, L- 3-[ 3-( acetylthio)- 1- oxopropyl]- 2- thiazolidine carboxylic acid and dicyclohexylamine salt ( 1:1)

(a) DL-thiazolidine-2-carboxylic acid

A mixture of 19.0 g (0.167 mol) of cysteamine hydrochloride and 14.9 g (0.201 mol) of glyoxylic acid hydrate was dissolved in 15 ml of water. After addition of 50 ml of absolute ethanol, the reaction mixture was allowed to stand at room temperature for 45 minutes and was then treated with 13.8 g (0.168 mol) of sodium acetate. The reaction mixture was allowed to stand sealed at room temperature overnight, and was then cooled for 1 hour and filtered using chilled 95% ethanol for washing. After drying, the crude product (16.4 g) showed a melting point of 178-180°C (dec.). Analysis showed that the raw product contained approx. 0.93 molar equivalents of sodium chloride.

Analysis for C4H7N02S•0.93 NaCl

Calculated: C 25.62, H 3.76, N 7.47, S 17.10, Na 11.40, Cl 1758 Found: C 25.79, H 3.99, N 7.32, S 17/36 , Na 11.59, Cl 18.21.

b) DL- 3-[ 3-( acetylthio)- 1- oxopropyl]- 2- thiazolidinecarboxylic acid- dicyclohexylamine salt

To a mixture of 56 ml of water and 63 ml of tetrahydrofuran cooled in an ice-salt bath was added 7 g (37.4 mmol) of thiazolidine-2-carboxylic acid (x 0.93 NaCl) and 9.1 g (112 mmol) sodium bicarbonate. To the well-stirred, cooled mixture was added dropwise 5.0 ml (37.4 mmol) of 3-acetylthiopropionyl chloride. During the addition, the pH value was monitored with a pH meter and kept close to pH 7 by the addition of solid sodium bicarbonate. Base consumption began immediately upon addition of the acid chloride. After approx. At 45 minutes the pH stopped changing and the reaction was quenched by addition of 1N HCl to pH 1 and the reaction mixture was extracted with ethyl acetate. The organic solution was washed with water and brine, dried with sodium sulfate and concentrated in vacuo to 10.0 g of crude product. Chromatography on 850 g of Baker silica gel eluted with CHCl 2 :MeOH:HOAc, 20:2:1 gave 9.2 g of crude product which by thin layer chromatography still showed traces of contamination and contained a small amount of solvent.

The chromatographed free acid (9.3 g) was dissolved in 6 ml of ethyl acetate and added to 300 ml of ether. About. 200 ml of ethyl acetate was added to give a clear solution which after 2 hours at room temperature yielded 13.3 g (89%) of analytically pure product, m.p. 195-196°C and neutralization equivalent 444 (calcd 445). Recrystallization from methanol/ether raised the melting point to 197-198°C.

The dicyclohexylamine salt from part (b) (3 g, 6.72 mmol) was partitioned between 67.2 mL of 0.2N aqueous H 2 SO 4 and 67 mL of ethyl acetate. The aqueous phase was further extracted with 60 ml and 30 ml portions of ethyl acetate, and the combined ethyl acetate solutions were washed with two 15 ml portions of water and 15 ml of saline. After drying over Na 2 SO 4 , removal of solvents in vacuo gave 1.9 g of crude product. The semi-solid product was dissolved in CH 2 Cl 2 and dispensed into ampoules, after which the solvent was removed as best as possible by concentration under a stream of nitrogen followed by a final drying in high vacuum. The product obtained was homogeneous by analytical thin-layer chromatography: Rf = 0.47 on silica gel; 12:2:1, CHCl3:MeOH:HOAc (visualization with iodine).

Example 12

DL- 4-( 3- mercaptol- l- oxopropyl)- 3- morpholine- carboxylic acid

(a) DL- 4-( 3- acetylthio- l- oxopropyl)- 3- morpholine- carboxylic acid

0.0207 mol of 3-morpholine carboxylic acid hydrochloride was

reacted with 0.023 mol of 3-acetylthiopropionyl chloride in 50 ml of water which originally contained 2.2 g of sodium carbonate, at ice bath temperature. The reaction mixture was maintained at pH 8-8.2 by adding a total of 17 ml of 25% aqueous sodium carbonate solution during the reaction. When the pH stopped changing, the reaction was quenched with hydrochloric acid and the reaction mixture was extracted with ethyl acetate. The ethyl acetate extract gave 5.0 g of a crude, syrupy product which was converted to the dicyclohexylamine salt in 50 ml of ethyl acetate, wt 7.1 g, m.p. 182-185°C with sintering at 170°C. Crystallization from 300 ml of acetonitrile gave 5.7 g of colorless salt, m.p. 187-189°C.

Analysis for c;lqh;l5N05S'ci2H23N

Calculated: C 59.70, H 8.65, N 6.33, S 7.25

Found: C 59.46, H 8.53, N 6.22, S 7.53.

The dicyclohexylamine salt was converted to the free acid using 50 ml of 10% aqueous potassium bisulphate solution, m.p. 110-112°C.

Analysis for C, _H, .-N0.-S:

J 10 15 5

Calculated: C 45.96, H 5.79, N 5.36

Found: C 46.26, H 6.01, N 5.51.

b) DL-4-(3-mercaptol-ol-oxopropyl)-3-morpholine-carboxylic acid 0.013 mol of the product from part (a) was hydrolyzed with 8 ml of concentrated ammonium hydroxide in 19 ml of water with stirring for 2 hours at room temperature after the solution was prepared. The resulting colorless syrupy product weighed 2.75 g.

Analysis for CgH13N04S•1/4 H20

Calculated: C 42.94, H 6.08, N 6.26, S 14.33

Found: C 42.84, H 6.22, N 6.30, S 14.12.

Example 13

3-( 3- mercapto- 2D- methyl- l- oxopropyl)- L- 4- thiazolidine carboxylic acid- 1, 1- dioxide

3-(3-Acetylthio-2D-methyl-1-oxopropyl)-L-4-thiazolidinecarboxylic acid 1,1-dioxide (2.9 g) was added to a rapidly stirred mixture of 100 ml of water, 30 ml of methanol and 100 ml of concentrated ammonium hydroxide which was cooled in an ice bath. After 45 minutes, the reaction mixture was quickly washed with 150 ml of ethyl ether and 50 ml of ethyl acetate and was then acidified to pH 1 with concentrated hydrochloric acid. The mixture was extracted with ethyl acetate,

and the oily product obtained from the ethyl acetate extract was dissolved in 50 ml of ethyl acetate and treated with 1.3 ml of dicyclohexylamine and then diluted with ether to give 2.1 g of product as dicyclohexylamine salt, m.p. 221-222°C,

[α]^<5> -71.4° (c = 1, methanol).

The dicyclohexylamine salt gave on neutralization 1.11 g of the free acid as a hygroscopic semi-solid.

Analysis for c8Hi3N05S2 ' 1// 2 H2°

Calculated: C 34.77, H 5.10, N 5.07, S 23.21

Found: C 34.92, H 5.16, N 9.85, S 23.50 Neutralization equivalent, calculated 276.3, found 279.

Example 14

DL- 3-(3-mercaptol-ol-oxopropyl)-2-thiazolidinecarboxylic acid and dicyclohexylamine salt

a) DL- 3-( 3- mercaptol- l- oxopropyl)- 2- thiazolidine carboxylic acid dicyclohexylamine salt

Argon was bubbled through 40 ml of water for 5 minutes. To this was added 35 ml of 58% aqueous NH^OH solution, and the mixture was quickly stirred and covered with a blanket of argon while 7.00 g (15.7 mmol) DL-3-(3-acetylthio-1-oxopropyl) -2-thiazolidinecarboxylic acid dicyclohexylamine salt was added portionwise as a solid. After stirring the mixture at room temperature for 20 minutes, the reaction mixture was cooled to ice bath temperature and extracted with 40 ml and 30 ml portions of Cl 2 Cl 2 . The aqueous phase was then rapidly cooled again under argon and acidified to pH 1 with concentrated HCl and then extracted with 100 ml plus 3 x 50 ml portions of ethyl acetate.

The combined organic solutions were washed with 25 ml of water,

25 ml of brine, dried (Na 2 SO 4 ) and concentrated in vacuo to give the crude oily product. The oil was dissolved in 30 ml of methanol and treated with 3.13 ml (15.7 mmol) of dicyclohexylamine. After addition of 250 ml of ether and standing for 3 hours at room temperature, a yield of 5.71 g (90%) of dicyclohexylamine salt was obtained, m.p.: sintered at 200°C, melting 206-210°C (dec.).

Analysis for C^H-^NO.^ • DCHA salt

Calculated: C 56.68, H 8.51, N 6.96, S 15.93

Found: C 56.41, H 8.37, N 6.80, S 16.10

SH titration: 99.3%.

b) DL-3-(3-mercaptol-1-oxopropyl)-2-thiazolidinecarboxylic acid The dicyclohexylamine salt (5.60 g, 13.9 mmol) was added to

a separatory funnel containing 100 ml of ethyl acetate and 140 ml of ice-cold, 0.3N l^SO^ solution which had been flushed with argon.

The layers were shaken, separated, and the aqueous layer was further extracted with three 50 mL portions of ethyl acetate. The combined ethyl acetate extracts were washed with 25 mL water, brine, dried (Na 2 SO 4 , under argon), and then concentrated in vacuo. Traces of ethyl acetate were removed by dissolving the oily concentrate three times in approx. 150 ml CH2C12 and evaporation to dryness. The final weight was above the quantitative due to dissolved solvent remaining in the oil. Analytical thin layer chromatography: R f = 0.37, silica gel, 60:20:6:11, EtOAc:pyridine:MeOH:H 2 O, visualization with iodine.

Claims (1)

Analogous method for the preparation of a therapeutically active compound of the formula: wherein R 1 and R 2 are each hydrogen or lower alkyl; R 3 is hydrogen or lower alkyl; R^ is hydrogen, lower alkanoyl or benzoyl, X is 0, S, SO or SO 2 , m being 2 and n being 1 when X is 0; m is 1 or 2, n is o, 1 or 2 and m + n is 2 or 3; p is 0 or 1, and basic salts thereof, characterized in that a compound with the formula is reacted with a compound with the formula or a reactive equivalent thereof, and optionally a compound of formula I where R^ is different from hydrogen is hydrolyzed to form a compound where R^ is hydrogen, and/or optionally a compound of formula I where X is S is oxidized to a compound where X is SO or SO 2 , or a compound where X is SO to a compound where X is SO 2 , and optionally the compound produced is converted into a basic salt.