tert-butyl (2S)-1-[(2S)-2-aminopropanoyl]pyrrolidine-2-carboxylate

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Di-t-butyl bromomalonate (15.9 g.) was added to a stirred solution of L-alanyl-L-proline t-butyl ester (26 g.) in toluene (30 ml.) which was cooled to 0° C., and the mixture was allowed to warm up to laboratory temperature during 2 hours and was stirred at that temperature for 14 hours. Diethyl ether (300 ml.) was added, the mixture was filtered to remove unreacted L-alanyl-L-proline t-butyl ester as the hydrobromide salt thereof, andthe filtrate was evaporated to dryness under reduced pressure. The residue was dissolved in ethyl acetate (400 ml.) and the mixture was washed successively twice with aqueous 0.5N-citric acid solution (100 ml. each time), once with water (100 ml.) and once with saturated aqueous sodium chloride solution (100 ml.), dried over magnesium sulphate and evaporated to dryness under reduced pressure. The residue was purified by chromatography on a silica gel column using a 9:1 v/v mixture of dichloromethane and ethyl acetate as eluant, and there was thus obtained as an oil N-[di-t-butoxycarbonylmethyl]-L-alanyl-L-proline t-butyl ester. to warm up to laboratory temperature during 2 hours the mixture was filtered to remove unreacted L-alanyl-L-proline t-butyl ester as the hydrobromide salt andthe filtrate was evaporated to dryness under reduced pressure The residue was dissolved in ethyl acetate (400 ml.) the mixture was washed successively twice with aqueous 0.5N-citric acid solution (100 ml each time), once with water (100 ml.) and once with saturated aqueous sodium chloride solution (100 ml.), dried over magnesium sulphate evaporated to dryness under reduced pressure The residue was purified by chromatography on a silica gel column a 9:1 v/v mixture of dichloromethane and ethyl acetate as eluant

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Dimethyl bromomalonate (6.6 ml.) was added to a stirred solution of L-alanyl-L-proline t-butyl ester (22.9 g.) in dichloromethane (40 ml.) which was cooled to 0° C., and the mixture was allowed to warm up to laboratory temperature and stirred at that temperature for 16 hours, then diluted with dichloromethane (150 ml.). The solution was washed successively twice with aqueous 0.5N-citric acid solution (150 ml. each time), water (200 ml.) and saturated aqueous sodium chloride solution (200 ml.), dried over magnesium sulphate and evaporated to dryness. The residue was purified by chromatography on a silica gel column using a 4:1 v/v mixture of dichloromethane and ethyl acetate as eluant, and there was thus obtained as an oil N-(dimethoxycarbonylmethyl)-L-alanyl-L-proline t-butyl ester. to warm up to laboratory temperature The solution was washed successively twice with aqueous 0.5N-citric acid solution (150 ml each time), water (200 ml.) and saturated aqueous sodium chloride solution (200 ml.), dried over magnesium sulphate evaporated to dryness The residue was purified by chromatography on a silica gel column a 4:1 v/v mixture of dichloromethane and ethyl acetate as eluant

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Ethyl 6,6-ethylenedioxy-2-oxo-6-(2-thienyl)-hexanoate (3 g.) and L-alanyl-L-proline t-butyl ester (2.5 g.) were added to a suspension of freshly activated 4 Å molecular sieve (15 g.) in ethanol (35 ml.) and the mixture was stirred at laboratory temperature for 1 hour. Acetic acid (1.25 ml.) was added, and sodium cyanoborohydride (0.625 g.) was then added in portions during 1 hour. The mixture was stirred at laboratory temperature for 48 hours and then filtered, and the filtrate was evaporated to dryness under reduced pressure. The residue was purified by flash chromatography on a silica gel column using a 40:1 v/v mixture of dichloromethane and methanol as eluant. There was thus obtained as an oil N-[1-ethoxycarbonyl-5,5-ethylenedioxy-5-(2-thienyl)pentyl]-L-alanyl-L-proline t-butyl ester. The mixture was stirred at laboratory temperature for 48 hours filtered the filtrate was evaporated to dryness under reduced pressure The residue was purified by flash chromatography on a silica gel column a 40:1 v/v mixture of dichloromethane and methanol as eluant

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: A solution of ethyl 2-bromo-9-phthalimidononanoate [prepared according to Chem. Pharm. Bull. 34, 2078 (1986)] (3 g), L-alanyl-L-proline tertiary butylester (1 g) and triethylamine (0.8 ml) in acetonitrile (200 ml) is subjected to 60 hours' reflux. Acetonitrile is distilled off under reduced pressure, and the residue is dissolved in ethyl acetate. The solution is dehydrated with anhydrous sodium sulfate, and thereafter is concentrated to obtain an oily residue. When this substance is applied to a silica gel chromatography (ethyl acetate:n-hexane=3:1) for separative purification, 310 mg of N-[1(R)-ethoxycarbonyl-8-phthalimidooctyl]-L-alanyl-L-proline tertiary butylester is obtained as a colorless oil from the first effluence. ' reflux Acetonitrile is distilled off under reduced pressure the residue is dissolved in ethyl acetate is concentrated to obtain an oily residue When this substance is applied to a silica gel chromatography (ethyl acetate:n-hexane=3:1) for separative purification

Conditions: temp:-20.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: L-alanyl-L-proline t-butylester (2.42 g, 10 mmole) was dissolved in methylene chloride (50 ml), and triethylamine (1.01 g, 10 mmole) was added thereto with cooling to -20° C and stirring. Methylene chloride (5 ml) solution of ethanesulfonylchloride (1.29 g, 10 mmole) was added dropwise to the solution while keeping the temperature thereof to -20° C. and then stirred overnight at room temperature. The solution was washed with 10% aqueous citric acid, water, 5% aqueous sodium bicarbonate and water in the order given, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure to give oily residue of N-ethanesulfonyl-L-alanyl-L-proline t-butylester(yield: 1.04 g, 31.1%). to -20° C. and then stirred overnight at room temperature The solution was washed with 10% aqueous citric acid, water, 5% aqueous sodium bicarbonate and water in the order dried over anhydrous magnesium sulfate The solvent was distilled off under reduced pressure

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Benzyloxycarbonyl-L-alanyl-L-proline (16.02 g, 50 mmole) was dissolved in methylene chloride (100 ml) solution of 98% sulfuric acid (0.5 ml), and isobutene gas was blown into the solution under cooling until the volume of the solution was increased by 50 ml. After standing it for 64 hours at room temperature, the solution was washed with 0.15M sodium carbonate. The methylene chloride layer was further washed with 10% aqueous citric acid, water, 5% aqueous sodium bicarbonate and water in the order given, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure and the oily residue thus obtained was dissolved in t-butanol (90 ml). Hydrogen gas was passed through the solution in the presence of 5% palladium-carbon (6.5 g) as a catalyst for 2 hours at room temperature. The catalyst was separated by filtration and the solvent was distilled off under reduced pressure to give colourless oily L-alanyl-L-proline t-butylester (yield: 10.36 g, 85.5%). under cooling until the volume of the solution was increased by 50 ml for 64 hours at room temperature the solution was washed with 0.15M sodium carbonate The methylene chloride layer was further washed with 10% aqueous citric acid, water, 5% aqueous sodium bicarbonate and water in the order dried over anhydrous magnesium sulfate The solvent was distilled off under reduced pressure the oily residue thus obtained The catalyst was separated by filtration the solvent was distilled off under reduced pressure

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Benzyloxycarbonyl-L-alanyl-L-proline (16.02 g, 50 mmole) was dissolved in methylene chloride (100 ml) solution of 98% sulfuric acid (0.5 ml), and isobutene gas was blown into the solution under cooling until the volume of the solution was increased by 50 ml. After standing it for 64 hours at room temperature, the solution was washed with 0.15M sodium carbonate. The methylene chloride layer was further washed with 10% aqueous citric acid, water, 5% aqueous sodium bicarbonate and water in the order given, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure and the oily residue thus obtained was dissolved in t-butanol (90 ml). Hydrogen gas was passed through the solution in the presence of 5% palladium-carbon (6.5 g) as a catalyst for 2 hours at room temperature. The catalyst was separated by filtration and the solvent was distilled off under reduced pressure to give colourless oily L-alanyl-L-proline t-butylester (yield: 10.36 g, 85.5%). under cooling until the volume of the solution was increased by 50 ml for 64 hours at room temperature the solution was washed with 0.15M sodium carbonate The methylene chloride layer was further washed with 10% aqueous citric acid, water, 5% aqueous sodium bicarbonate and water in the order dried over anhydrous magnesium sulfate The solvent was distilled off under reduced pressure the oily residue thus obtained The catalyst was separated by filtration the solvent was distilled off under reduced pressure

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: A solution of 0.57 g of sodium cyanoborohydride in 40 ml of ethanol is added dropwise to a stirred mixture of 2 g of N-(L-alanyl)-L-proline tert-butyl ester.oxalate, 1 g of sodium acetate, 0.72 g of acetic acid, 10 g of molecular sieves 3 A, 3.4 g of ethyl 6-(1-benzyloxycarbonyl-4-piperidyl)-2-oxohexanoate and 50 ml of ethanol at room temperature over a period of 2 hours. The reaction mixture is allowed to stand overnight and filtered, and the filtrate is concentrated under reduced pressure. The resulting oily material is purified by silica gel column chromatography (hexane:ethyl acetate=1:1 to hexane:acetone=1:1) to give 0.66 g of N-[N-[5-(1-benzyloxycarbonyl-4-piperidyl)-1-ethoxycarbonylpentyl]-L-alanyl]-L-proline tert-butyl ester as a colorless oil. filtered the filtrate is concentrated under reduced pressure The resulting oily material is purified by silica gel column chromatography (hexane:ethyl acetate=1:1 to hexane:acetone=1:1)

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: 1.65 g of Z-Gly-ONP and 1.09 g of H-Ala-Pro-OtBu in 20 ml of ethyl acetate are allowed to stand for one hour at 0° C. and 20 hours at room temperature. The mix is then diluted with ethyl acetate, washed with 50% saturated potassium carbonate solution and with water, dried over sodium sulphate and evaporated. 2.06 g of an oil which is homogeneous according to thin layer chromatography result. Rf = 0.8 in the system chloroform-methanol (1:1) on silica gel. washed with 50% saturated potassium carbonate solution and with water dried over sodium sulphate evaporated result

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: 9.36 g of Z-Ala-Pro-OtBu are hydrogenated in 100 ml of methanol in the presence of 500 mg of Pd on charcoal (10%) at room temperature. The hydrogen uptake has ended after 90 minutes. The solution is freed of catalyst by filtration and evaporated in a waterpump vacuum at 30° C. to give an oil. This proves to be homogeneous in a thin layer chromatogram on silica gel. Rf = 0.35 in the system chloroform-methanol (1:1). The solution is freed of catalyst by filtration evaporated in a waterpump vacuum at 30° C. to give an oil

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: A solution of L-alanyl-L-proline t-butyl ester (1.0 g, 4.13 mmol), N-[(t-butyloxy)carbonyl]-L-phenylalanine chloromethyl ketone (1.2 g, 4.03 mmol), sodium bicarbonate (370 mg, 4.4 mmol) and sodium iodide (300 mg, 2 mmol) in dimethylformamide (7 ml) was stirred overnight at room temperature. The reaction mixture was evaporated. Dioxane (6 ml) was added to the residue followed by benzyloxycarbonyl chloride (0.75 ml, 5.24 mmol) and sodium bicarbonate (500 mg, 5.95 mmol). After stirring overnight, the reaction mixture was evaporated, the residue taken up in ethyl acetate and washed neutral (water, saturated sodium bicarbonate solution, water, 10% potassium bisulfate solution and water). The crude material (2.6 g) was chromatographed over silica gel (200 g) using the solvent system ethyl acetate:benzene-1:3 to obtain 1.32 g of the title compound. The reaction mixture was evaporated Dioxane (6 ml) was added to the residue the reaction mixture was evaporated washed neutral (water, saturated sodium bicarbonate solution, water, 10% potassium bisulfate solution and water) The crude material (2.6 g) was chromatographed over silica gel (200 g)

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: A mixture of (S)-β-(benzoylamino)-α-[[(4-methoxyphenyl)methyl]thio]-benzenebutanal (isomer A) (0.45 g., 1.07 mmole), L-alanyl-L-proline, 1,1-dimethylethyl ester (0.78 g., 3.21 mmole), and crushed 3A° molecular sieves (2 g.) in tetrahydrofuran (5 ml.) and absolute ethanol (5 ml.) is stirred at room temperature under argon. After 2.5 hours, sodium cyanoborohydride (0.20 g., 3 eq.) is added and stirring continued for 15 hours. The reaction mixture is filtered to remove the sieves and the filtrate is diluted with ethyl acetate and washed with water, 1N hydrochloric acid (twice), 1N sodium bicarbonate, and brine. After drying over anhydrous MgSO4, the solvent is removed to give the desired product, as a mixture of diastereomers, as a pale yellow oil. TLC (silica gel, benzene:acetone, 4:1) isomer A at Rf =0.30 and isomer B at Rf =0.13. Flash chromatography (silica gel LPS-1; benzene:acetone, 7:3) affords impure product (isomer A). Repeated flash chromatography (silica gel LPS-1; ethyl acetate:benzene, 6:4) gives 0.20 g. of 1-[N-[(3S)-3-(benzoylamino)-2-[[(4-methoxyphenyl)methyl]thio]-4-phenylbutyl]-L-alanyl]-L-proline, 1,1-dimethylethyl ester (isomer A) as a nearly colorless oil. continued for 15 hours The reaction mixture is filtered to remove the sieves the filtrate is diluted with ethyl acetate washed with water, 1N hydrochloric acid (twice), 1N sodium bicarbonate, and brine After drying over anhydrous MgSO4 the solvent is removed