CS201034B2 - Method of producing amino acid - Google Patents

Abstract

For preparing the novel bestatin compounds of the formula <IMAGE> having R1 and R2 in accordance with the preceding Patent Claim 1, an alpha -hydroxybeta -amino acid of the formula <IMAGE> is used as the starting material. The latter is condensed, optionally after having protected the functional groups which are not to participate in the subsequent reaction, with a beta -amino acid of the formula <IMAGE> using a peptide-linking method. Protective groups which may optionally be present are subsequently removed. The optically pure diastereoisomers can be readily obtained from the compounds of the formula (III). The alpha -hydroxybeta -amino acids can be prepared by hydrolysing a corresponding cyanohydrin whose amino group is optionally protected. The novel compounds inhibit certain enzymes which decompose bleomycin and in this way amplify the anti-tumour activity of the bleomycins.

Description

The present invention relates to a process for the preparation of a novel amino acid of formula (I)

R1-CH-CH-COOH

II (I)

NH 2 OH wherein R 1 is alkyl of 1 to 4 carbon atoms, cyclopentylmethyl, cyclohexylmethyl, cyclohepttylmethyl or a group of formula ry (CH ₂ ) _n wherein R 8 is hydrogen, chlorine, methyl, nitro, hydroxyl or amino and _n is 0 or 1.

These compounds are useful as starting compounds for the preparation of aminoopeptidase B, leucimminopeptidase and bleomycin hydrolase compounds which increase the antitumor activity of bleomycin and have antitumor activity of formula II

2 1 2'1 '

R1 — CH — CH — CO — NH — CH — COOH (II)

III

NH ₂ OH R ₃ * wherein R 1 and R 8 are as defined above, and

R 1 is a substituent selected from the group consisting of hydrogen, alkyl of 1 to 6 carbon atoms, hydroxyalkyl of 1 to 12 carbon atoms, alkoxymethyl of 2 to 3 carbon atoms, alkylthiomethyl of 2 to 3 carbon atoms, carboxamidoalkyl of 1 to 2 carbon atoms, and carboxyalkyl of 2 to 3 carbon atoms.

Of the compounds of formula (II), (2S, 3R) -3-amino-hydroxy-4-phenylbenzoyl-Cl (S-leucine) is called bestatin, and can be prepared from a nutrient medium of a fungus producing non-actinomycetes. This new compound has the above-mentioned physiological activity

The amino acid or derivatives of formula I can be prepared by acid hydrolysis of nitrile derivatives of formula III

R-CH-CH-CN ¹¹ | (III) nh ₂ OH wherein R is as defined above or amino-protected nitrile derivatives.

Typical examples of derivatives of nitriles of general formula III may be mentioned drill carbamic acid 3-amino-2-hydrkχsprkpionové acid nitrile, 3-aiino-2-hydrkxy-5-iethylhlxaokvé acid nitrile, 3-aiino 2-hydroxyethyl QyЗ-flnylprkpikOkvé _l acid nitrile phenylbutyric acid, 3-amino-2-hydroxybutyric acid, 4-phenylisilyl, 3-amino-2-hydroxybutyric acid nitrile, 4-phenylbutyric acid nitrile, 3-amino-2-hydroxybutyric acid nitrile 4-p-Methylphenylsilyl, p-methylphenylbutyric acid nitrile, p-hydroxyphenylbutyric acid nitrile and 3-amino-2-hydroxybutyl-4-p-benzylphenylbutyric acid nitrile.

Any amino protecting group as known in peptide chemistry can be used to protect the amino group.

Preferred examples of acyl groups for protecting the amino group include, but are not limited to, phenyl, acetyl, cycloalkyl, benzoyl, optionally substituted; Examples of uretime-type amino protecting groups include optionally substituted benzyloxycarbonyl, C7-C7alkoxycarbonyl, and cycloalkoxycarbonyl.

Preferred examples of other protecting groups useful for protecting the amino group include arylsulfocyl groups, optionally substituted, phthalyl, o-nitropheryl, or tri-yl.

Any of the acids commonly used in the hydrolysis of nitriles can be used in the hydrolysis of the present invention. Examples of inorganic acids are hydrochloric, hydrobromic or sulfuric, examples of organic acids include alkylsulfonic acids, such as methyl sulfonic acid, ethylsulfonic acid, and further sulfonic acids, such as benzenesulfonic acid and sulfonic acids.

The concentration of these acids can be varied in any order as long as it is suitable for the hydrolysis of conventional nitriles. However, concentrations above 1 N can always be said to be preferred.

For amino-protected compounds that are poorly soluble in aqueous acid solutions, a water-miscible organic solvent such as tetrahydrofuran, dioxane, lower alcohols, acetone, dimethylformamide, dimethylacetimide and dimethylsulfoxide can be added. This increases the solubility of the compounds in aqueous suspensions.

If a protecting group is used, then the protecting group is hydrolyzed to form electrode materials, and a catalyst binding agent such as saline is added.

The temperature of the hydrolysis may be in any range from room temperature to the boiling point of the reaction mixture under reflux. Any of the usual procedures for such isolation can be used to isolate the expected amino acids from the reaction mixture.

For example, if the hydrolysis is carried out by treatment with a volatile acid, the excess acid used may be filtered by concentrating the reaction mixture under reduced pressure, and then the residue is dissolved in water and neutralized with an aqueous solution of the base until the electrochemical point is reached.

Further, acetone is added to the reaction network, if desired, with stirring. or ethanol, thereby crystallizing the compound of formula I, which is then filtered off. Alternatively, using a non-volatile acid, the reaction mixture is diluted with water such that it is less than 1 N per acid and the solution is poured over a strongly acidic ion-exchange resin so that the desired aminocrile is adsorbed onto the ion-exchange resin. It is then eluted from the ion exchange resin with a volatile base such as amoonac diluted with water and the solution is concentrated under reduced pressure. If necessary, acetone, methanol or ethanol is added to aid in the crystallization of the product, i.e. the compound of formula I, which is then filtered.

In the above reaction, (2RS, 3R) -aminokinesin is obtained as the starting compound (Xtrin); (2RS, 3S) -aminox-saliva is obtained

The (2RS, 3RS) -4-amino acid is thus obtained by sensing the (RS) -tril derivative.

The compounds of the formula I obtained can be used in the crude state as starting materials in the following process. For example, 3-amino-2-hydroxy-4-phenylbutanoic acid, which is the starting material for the preparation of bestatin, is in the form of (2S, 3R). Thus, in order to obtain this compound, it is necessary to cleave (2RS, 3R, 3-amino) -2-hydroxy-4-phenylbutanonic acid into (2S, 3R) - and (2R, 3R) -.

The (2RS, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid is separated into the optical isomers in ethyl acetate by separating the diastereoisomer obtained first with benzyloxycarboxylic acid (2RS, 3R). 3-Amino-2-hydroxy-4-phen-1-butanoic acid is converted to benzyloxycarbonyl- (2RS, 3R) -3-amino-2-hydroxy-4-phenyl-butanoic acid and then the brucine salt is prepared.

More precisely, by heating in ethyl acetate, benzyloxycarbonyl- (2RS, 3R) -3-amino-22-yl is dissolved in x-4-phenylbutanoic acid and the corresponding equivalent of brucine, or a small excess of this, and the solution is filtered if necessary.

Upon cooling or cooling of the filtrate, crystals precipitate, which are separated off and, if necessary, recrystallization from ethyl acetate is repeated until an optically pure salt of brucine with benzyloxy-carbones is obtained (2S, m.p. 3R) 3-Amino-2-hydroxy-4-phenylbutane. Conventional treatment of this brucine salt further affords optically pure benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxyl-4-phenylbutanoic acid.

Alternatively, the pure benzyl-4-phenyl-4-phenyl-butanoic acid may be prepared as follows:

Brucine salt and benzyloxycarbonyl- (2), 3R) -3-amino-2-hydro-5X-4-phenylbutanoic acid salt. in ethyl acetate containing a small amount of benzyloxycarbonyl (2S, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid, the brucine is removed by shaking with a dilute hydrochloric acid solution, then the ethyl ester layer is added to the ethyl ester ^With acetic acid is dried with a deldatizing agent, such as anhydrous sodium sulfate, and the filtrate is flooded under reduced pressure. The residue was dissolved in a small amount of ethyl acetate-acetic acid. Crystallization was added by the addition of petroleum ether and filtration of the sub-portion gave pure benzyl-4-carbonyl-2- (2R, 3) -3-amino-2-hydroxy) -44-phenylbutanoic acid.

It was found that (2), 3R) -foima acid. Benzyl-4-carboone-3-amino-o-h-dO-n-phenylbutanoic acid is less soluble in organic solvents than the (2S, 3R) form.

Benzyloic acid: (2R, 3R) -3-Amino-2-hydroxx-4-phenylbutanoic acid is first dissolved in a solvent at room temperature or with heating and addition of an additional solvent to precipitate the dissolved ieroprazole, precipitates the crystal. which can be isolated by filtration. If necessary, this process is repeated until optically pure benzyloxycarbonyl-S- (2R, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid is obtained.

The first solvent in which the substance is dissolved may be one of the lower alcohols, such as methanol, ethanol and propanol, and one of the ethers, such as diethyl ether, aits-or-o-ether, tetrahsar- furan and aioxane, then esters such as methyl or ethyl acetate, ketones such as acetone and methyl ethyl ketone, halogenated hydrocarbon derivatives such as methylene chloride and chloroform, amides such as dimethylformamide and dimethylacetamide, and finally some nitriles , for example

The second solvent (which is first diluted to precipitate the product) includes petroleum hydrocarbon fractions such as petroleum ether, gasoline and ligroin, aromatic hydrocarbons such as benzene and toluene, alkanes such as pentane and hexane, and cycloalkanes such as cyclopentane and cyOlohexane. Using the diastereoisomer and bruoin process, impure benzyloxycarbonyl- (2S, 3R) -3-aminoo-2-hydrohydro-4-phthyll-butyric acid can be obtained from the filtrate and purified to give optically pure (2S, 3R) -3 acid. -amino221hrdoxoxi-4-phenylbutane.

Benzyloxycarbonyl-3-amino-2-hydroxyl-4-phenylbutanoic acid used in the above resolution to optical antipodes can be synthesized using the Schotten and Baumann procedure by reaction of (2RS, 3R) -3-amino-2-Jh] 4-phenylbutamic with benzyloxycarbonyl chloride in the presence of a base, or in the presence of a tertiary amine such as triethyl or Nm, in which case benzyloxycarbonylating agent benzyloxycarbonylronynp-n-thiopheneacetone, benzyloxycarbonyl, benzyloxycarbonylate, benzyloxycarbonylating agent,

benzyloxycarbonyl-N-pyrrolidinoimidonitrile; and benzyl-5-carbonyl-4,6-dimethyl-2-mercaptopyrimidine in a solvent which is a mixture of dioxane, tetrahydrofuran, acetontril or formamide with water.

Although the above-mentioned resolution to the optical antipodes is described with respect to 3-arino-2-1H-hydroxyl-14-phenylbutannic acid, a similar procedure or desirably for other amino acids may be employed.

The following is a process for preparing bestatin and related compounds of formula II using the compounds of formula I of the invention.

AAinnnolstina of formula II is condensed with a compound of formula IV \ H ₉ -CH-COOH I ² (IV) ^R 3 wherein R is as defined above for desert procedure is usually employed on sorts of amino acids in peptide chemistry, and then removes protecting groups which may have been used, if necessary, to protect functional groups that did not participate in the actual reaction.

Typical examples of aminonolsein include glycine, alanine, 2-arinnbutyric acid, valine, norvaline, leucine, norleucine, isoleucine, tert-leucine, 2-amino-heppenoic acid, 2-amino-5-rethylhtanannic acid, 2-aminoctanoic acid, 2-Amino-chloro-heptanoic acid, serine, threonine, allothrenine, cysteine, hnmooylSein, asparagine, glutamine, O-mrttylesrine, O-εNгу ^ ег ^, O-prooplserine, meth-l.oi ^^ n, ethionine, acid aspartic acid, glutaric acid, phenylglycine, p-reoxygeneenl-glycine, phenylalanine, tyrosine, p-mmtho3x16 ^ 1811, and p-nitrophenylalanine.

Methods for condensation of the amino kislinines of formula I with the amino isoisoline of formula IV include the carbodiimide procedure using either. diciclohexylcarbodimide or 1-ethyl-3- (3-diretyllarinopropyl) carbodiiride. Further, the azide process may be carried out using the Opsee nitrite or its all-ester, the mixed anhydride process may be carried out using the ethyl chloroformate or isobutplester of chloroformic acid, a process using active esters such as cesium, vinyl ester, an optionally substituted phenyl ester, thiophenyl ester or N-hydroxyethylsulfonamide, further a process with an O-acylhydroxylarine dihydrate such as O-acylacetoxime or O-acyl acetyl hexanone oxime, and a process with N-acylderivatives to use carbonyliiriazoles.

Organic solvents useful in the above reaction include ethers such as tetrahydrofuran or dioxane, esters such as ethyl ester or methyl ether, ketones such as acetone or methyl ethyl ketone, halogenated hydrocarbon derivatives such as methylene chloride or chloroform, amides such as dimethylformamide or dimethylacetamide and nitriles such as acetoniirr.

In the case of condensation of an amino-protected amino group of the formula I with an amino-protected carboxyl group and amino acids of the formula IV with an unprotected carboxyl group and using active ester processes, a solvent mixture of water and water-miscible solvent may be used as an organic base such as sodium hydrogen carbonate magnesium, or in the presence of an organic tertiary amine, such as triethyl amine or N-trichloride.

Possible protecting groups can be removed from the resulting product using procedures conventional in peptide chemistry such as catalytic hydrogenation on palladium, quenching in an alkali environment, acidolysis by treatment with hydrogen bromide in acetic acid, reaction with rimonosulfonic acid, hydrogen chloride in dioxane, tetrahydrofuran or ethyl acetate. acetic acid ester, reaction with liquid hydrogen fluoride, lydrazinolysis by treatment with hydrazines or reaction with sodium in liquid ammonium. The expected compound of formula (II) is thus obtained.

* ·

V. * · ·

The nitride derivative of the general formula (III), which is used as starting material in the process according to the invention, can be synthesized by reducing the alpha-aminoocylsulfonyl, whose amino group is. the protected carboxyl group is converted to the function of the amide group by reaction with. a secondary amine at a temperature below 0 ° C and a metal oxide in an ether such as diethyl ether or tetrahydrofuran; the amino-protected aldehyde is protected. This is converted to an adduct with sodium dihydrogen sulfite and then further reacted with an alkali metal cyanide or directly with hydrogen cyanide to form the cyanohydride or 3-amino-nitrile nitrile.

Examples of preferred secondary amines are aziridine,

N-carboxylic acid, carbazole, 3,5-di-m-thiopyrazole and imidazole. Jako pří; Preferred metal hydrides include hydride, hydride, hydride, hydride, and urea-bis ((melamine) oxy) amine hydride.

An example of a protecting group for an amino group is one of the following: of the protecting groups usually used in peptide chemistry. Preferably, urethane-type protecting groups may be used, especially a benzyloxycarbonyl protecting group is preferred.

According to a second possible method, the amino group in the lower alkyl alpha-amino acid ester is protected by a suitable protecting group such as any of the above, followed by reducing the amino-protected product at a temperature below -40 ° C in an aromatic hydrocarbon such as toluene or benzene. or one of ethers such as diethyl ether or trichlorofuran by treatment with a metal hydride such as lithium hydride, diisobutylaluminium hydride or urethi-bis- (2-retinoxylninyl) amino uride to form the aminoaldehyde, protected with an arinine.

Treatment of the aminoaldehyde as described above gives 3-amino-2-1,027-7.

The physiological efficacy of bestatin and the non-existent compounds of formula II are determined as follows:

(a) Inhibitory efficacy against arsenicidose B

Test procedure:

It has been modified by the method that popssai the V.K. Hopusu, the K.K. Mak strain, G.G. Glenner in Arch. Biochem. Biophys. 114, 557 (1966). To a mixture of 0.3 ml of a 1 mM solution of beta-naphthyl amide argimide and 1.0 ml of 0.1 M hydrochloride tris buffer (pH 7.0), 0.7 ml of distilled water is added with or without the resins. and everything is heated to 37 ° C for 3 minutes.

б

The reaction is initiated by the addition of 0.2 ml of aminopeptidase B solution prepared using Sephadex RG 100 chromatography as described by Hopusu et al. (see above).

After 30 minutes of storage at 37 ° C, 0.6 ml of 1.0 M acetate buffer (pH 4.2) containing 1.0 mg / ml o-aminoazotoluene diazonium salt and 1.0% Tween 20 are added. . After 15 minutes at room temperature, the absorption (a) was measured at 530 nm with a spectrophotometer. For control, the absorption (b) is measured in a similar manner after the reaction without the sample itself. The inhibition percentage is calculated from the equation:

(b-a) / b.100.

Percent inhibition at various concentrations was measured and 50% inhibition (ΙΏ ^ θ) was derived from the% inhibition found. The results are shown in Table 1.

Table 1

Inhibitory activity of bestatin and related compounds against aminopeptidase B.

substance no.

Ingredients .

2.

3.

4.

5.

6.

7.

8.

9.

10.

11.

12.

1.3.

14.

15 Dec

16.

17.

18.

19 Dec

20 May

21.

22nd

23.

24.

25.

26.

(2S, (2S, (2S, (2S, (2S, (2S, (2S, (2S, (2S, (2S, (2S, (2S, _.

(2RS, (2RS, (2RS, (2RS, (2RS, (2RS, (2S, 3R) -3-amino) 2-hydroxy-4-p-nitrophenylbutanoyl- (S) -leucine (2S, 3R)) - 3-Amino-2-hydroxy-4-p-aminophenylbutanoyl- (S) -leucine (2S, 3R) -3-amino-2-hydroxy-4-cyclohexylbutanoyl- (S) -leucine (2RS, 3RS) -3 -amino-2-hydroxy-4-p-hydroxyphenylbutanoyl- (S) -leucine (2S, 3R) -3-amino-2-hydroxy-4-p-hydroxyphenylbutanoyl- (S) -leucine

3R) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -leucine (bestatin) 3S) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -leucine 3R) -3-amino- 2-hydroxy-4-phenylbutanoyl- (R) -leucine

3S) -3-amino-2-hydroxy-4-phenylbutanoyl- (R) -leucine 3S) -3-amino-2-hydroxy-4-phenylbutanoylglycine 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- ( S) -valine 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -isoleucine 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -methionine 3R) -3-amino 2-hydroxy-4-phenylbutanoyl- (S) -glutamine 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -norvaline 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- ( S) -norleucine 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -phenylalanine 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -serine

3R) -3-Amino-2-hydroxy-4-phenylbutanoyl- (S) -glutamic acid 3R) -3-Amino-2-hydroxy-4-phenylbutanoyl- (RS) -2-aminoctanoic acid

3R) -3-amino-2-hydroxypropionyl- (S) -leucine

3R) -3-amino-2-hydroxy-5-methylhexanoyl- (S) -leucine 3R) -3-amino-2-hydroxy-3-phenylpropionyl- (S) -leucine 3RS) -3-amino-2- hydroxy-4-p-chlorophenylbutanoyl- (S) -leucine

3RS) -3-amino-2-hydroxy-4-o-chlorophenylbutanoyl- (S) -leucine (3RS) -3-amino-2-hydroxy-p-methylphenylbutanoyl- (S) -leucine ^ID 50 tpg / ml)

0.10

1,25

0.56

0.04

21.5

0.55

0.05

0.22

1,2

0.17

0.13

4.2

0.72

3.1

0.07

0.48

0.01

0.01

0,10, 05

0.10

0.007 (B) Distribution of bleomycin in organs in the presence of bestatin.

100 mg / kg bleomycin B ₂ is injected subcutaneously into each of the 3 male ICR mice (4 weeks of age) and similarly, each of the 3 same age mice is injected subcutaneously with 100 mg / kg bleomycin B _{2 and} 100 mg / kg simultaneously. bestatin.

One hour after the administration of these Fridays, all 6 ^{µg were} sacrificed and the amount of bleomycin remaining in serum and organs was determined using a thin layer on a Petri dish, using Bacillus subtilis PCI219 as the test microorganism. The results are shown in Table 2, the figures being average pg / ml.

Table 2

Effect of concomitant administration of bestatin and bleomycin

B2 on bleomycin distribution in mouse organs.

serum lung pokoSka liver kidneys spleen stomach brain bleimycin 100 mg / kg 110 11.2 . 4,5. 0 68.0 4.0 1.6 0 bleomycin 100 mg / kg + bestatin 100 mg / kg 140 21.2 42.0 0 84.0 5.8 2.5 2.4

(C) Anniincence properties.

Effect of bestatin against fertilization

Immediately after subcutaneous administration of 10 mg / kg of bestatin to female ICR JCL mice at pubescent age, these mice are housed together with males of similar age. Females are killed after mating 2 days after vaginal plug formation, and eggs are collected to determine whether or not they have been fertilized. Fertilized eggs were from a period of 8-aS 16 cells.

Group of mice after administration of bestatin female no. 1 No.2 total N = 2 corpus luteim, count 10 19 Dec 29 - eggs collected, number 10 19 Dec 29 fertilized eggs 4 (41 «) 6 (31.6%) 10 (34.5 «) unfertilized eggs 6 (59%) 13 (68.4%) 19 (65.5 «) Control corpus luteim, count 13 13 26 eggs collected, number 9 11 20 May fertilized eggs 6 (66.7 «) . 8 (72.7%) 14 (69%) unfertilized eggs 3 · (33.3%) 3 (27.3%) 6 (31 «)

From the experiment, it is apparent that fertilization of the eggs collected from the 4 and 16-hour copolymers 16 hours after bestatin administration corresponds to 34.5%, whereas the control group is 70%. These results show that Se is bestatin with anti-fertilization activity.

Further, there is an analysis of the procedures for preparing the compounds of the invention. With reference to the examples, it will be appreciated that any method for introducing a suitable protecting group, any method for removing a protecting group from a functional group, as well as any other method of sequencing and linking an amino group may be used. as known from peptide chemistry in addition to those explicitly described herein.

Holdfi Rf refers to silicone plates GF254 (Merck) using the n-butyl alcohol: acetic acid: water (4: 1: 1) developing solution.

Some of the 3-amino-2-hydroxycarboxylic acids as well as the peptides containing them have 2 Rf values because they are mixtures of threo- and erythro- configurations.

He did

Dissolve in a mixture of 300 ml of concentrated hydrochloric acid and 300 ml of dioxane

35.4 g of oily benzyloxycarbonyl- (2RS, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid, nitrile, and after addition of 21.2 g of anisole, the reaction mixture is refluxed for 12 hours.

Dioxane was distilled off under reduced pressure, the aqueous solution obtained was washed with ether, and the aqueous phase was concentrated to dryness under reduced pressure. 200 ml of water are added to the residue, the insoluble matter is filtered off, the same amount of acetone is added and the pH of the reaction mixture is adjusted to pH 5 by adding 5% ammonia solution. The reaction mixture is left in the refrigerator, the precipitated crystals are filtered to obtain 13.66 g of the expected (2RS, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid.

[.alpha.] + 16.8 DEG (c = 1.25, N HCl);

R f = 0.23 and 0.26

Calcd for C ₁₀ H ₁₃ NO ₃ : C, 61.52; H, 6.71; N, 7.18;

Found:% G, 61.06;% H, 6.55;% N, 6.80.

13.66 g of (2RS, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid are dissolved in 70 ml of 1 N sodium hydroxide solution. The solution was stirred vigorously with ice-cooling, and 15 ml of benzyloxycarbonyl chloride and 70 ml of 1 N sodium hydroxide solution were added in 3 portions over 30 minutes. Thereafter, the reaction mixture was stirred vigorously for 1 hour with ice cooling and for a further 3 hours at room temperature. After completion of the reaction, the pH of the reaction mixture was adjusted to 1 by addition of 6 N hydrochloric acid. This gave an oily product which was extracted twice into 100 ml of ethyl acetate.

The ethyl acetate phase was washed with water and dried over anhydrous magnesium sulfate. After filtration, the filtrate is concentrated under reduced pressure. Crystallization of the residue from a mixture of ethyl acetate and petroleum ether gave 8.48 g of optically impure benzyloxycarbonyl- (2R, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid. On the other hand, concentrating the mother liquor under reduced pressure and crystallizing the residue from petroleum ether yields 8.07 g of optically impure benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid.

Mp 142-143 ° C. [.alpha.] + 70.2 DEG (c = 1.44, acetic acid).

Mixture 8.00 g of optically impure benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenyl-butanoic acid and 10.35 g of brucine dihydrate are dissolved in a mixture of 300 ml of ethyl acetate and 10 ml of heating. of methanol. After filtration, the filtrate was cooled to precipitate the salt of brucine and benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid.

The precipitated crystals were filtered off and crystallized twice from ethyl acetate to give 13.51 g of pure brucine and benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid salt.

Mp 144 ° C, [α] + 33.8 ° (c = 1.12, acetic acid).

13.5 g of the above crystals are suspended in 150 ml of ethyl acetate and the brucine is removed by shaking the suspension into 30 ml of 1 N hydrochloric acid solutions. The ethyl acetate layer is washed with water and, after drying over anhydrous magnesium sulfate, the inorganic fraction is filtered off, and the ethyl acetate is distilled off in vacuo. Precipitation of the residue from a solution of ethyl acetate and petroleum ether gave 6.21 g of benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenyl-butanoic acid. _of

Mp 15-4.5 ° C, [alpha] ²⁴ + 84.5 ° (c = 1.34, acetic acid).

578 ^P ro ^C1 ₈ H ₁₉ ^N ₅ 0 Calculated: 65.64% C, 5.82% H, 4.25% N;

Found: C, 65.29; H, 5.80; N, 4.35.

Benzyloxyoxycarbonyl- (2S, 3R) -3-amino-2-lhydro-4-phenylbutyric acid is hydrogenated on palladium black in acetic acid. The catalyst is filtered, the filtrate is then concentrated under reduced pressure and the residue is triturated in acetone. (2S, 3R) -1-Amino-2-hydroxy-2-hydroxylbutanoic acid is obtained after filtration in quantitative yield.

g γ ₈ + 29.5 ° (c = 0.97, 1 N hydrochloric acid).

For C ₁₀ H ₁₃ O ₃ N ₁ / 3H ₂ O Calculated: C 59.77, H 6.85, N 6.96;

found: ´ 59.86% C, 6.70% H, 6.87% N. '

Example 2.

Benzyloxycarbonyl- (2RS, 3S) -3-amino-22-cyclohexyl) -4-fetylbutanoic acid was prepared by benzyloxy / carbonnylation of (2RS, 3S) -3-amino-2-hydroxy-4-phenylbutanoic acid according to the procedure of Example 1.

7.75 g of benzyloxycarbonyl (2RS, 3R) -3-amino-2-hydroxy-4-phenylbutyric acid and 6.69 g of dehydroabiethylamine are mixed in ether and the sparingly soluble dehydroabiethylmonium salt of benzyloxycarbonyl (2R, 3R) is added. 3-Amino-2-hydroxy-4-phthyl-butyric acid is separated.

The prefiltered ethereal solution was bent under reduced pressure and the residual oily material was treated with a mixture of ethyl acetate and 1 N hydrochloric acid to prepare crude benzyloxycarbonyl per- (2S, 3S) -3-amino-2-hydroxy-4-phenyl butanoic acid. .

When the crystals were precipitated twice from ethyl acetate and ethyl acetate, 2.16 g of pure benzyloxycarbonyl-4S, 3S) -3-amino-2-hydroxy-4-phenylbutanoic acid was obtained, m.p. 175-176 ° C.

[.alpha.] D @ 20 + 5.6 DEG (c = 1.39, acetic acid).

The bicyclo [2- (3S) -3-amino-2-hydroxy] -4-phenylbutanoic acid is hydrogenated on palladium black in acetic acid. The alkaliser was filtered and the filtrate was then concentrated under reduced pressure. The residue was taken from acetone.

Filtration gave (2S, 3S) -3-amino-2-hydroxy-4-phenylbutanoic acid in quantitative yield.

[.alpha.] D @ ₂₀ = -5.6 DEG (c = 0.99, 1 N hydrochloric acid).

% C, 61.52; H, 6.71; N, 7.18; Found: C, 60.77; H, 6.56; N, 6.95.

Example 3

When (2S, 3R) -3-aminoo-2-hydroxy} -44-phenylbutanoic acid is tert-butoxycarbonyl, it is as described in T. Nagasawa et al., BuUl. Chem. Soc. Japan 46, 1269 (1973) using tert-butyl S-4,6-dimethylpyrimidin-2-ylthiolcarbooate to give the dicyclohexylammonium salt of the target in 77% yield. butoxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutanoic acid, m.p. 158-159 ° C.

[alpha] = + ⁵¹ ° ^for 9 ^ 27 ^ 2 ^ 44θ5 calculated: 68.02% C Found: 67.83% C; ^(c = ^{0 8 9} kvseHna ^acid)

S, 31% H, 5.88% N;

H, 9.52; N, 5.68.

Example 4

If 9.0 g of benzyloxycarbonyl- (2RS, 3R) -3-aino-2-hydroxy-propionitrile was reacted as described in Example 1, 3.16 was obtained. g of (2RS, 3R) -methylamine.

^[ alpha] 22 ^g = - ^0.6 ° ⁽ c = ≥ 10 1 N hydrochloric acid). Rf = 0

For C 4 H 8 NO 3: C, 40.33; H, 7.62; N, 11.76.

found. H, 7.51; N, 11.95.

Example 5

2.76 g of oily benzyloxycarbonyl- (2RS, 3R) -3-amino-22-dihydro-5-ethylheanonitrile are dissolved in 30 ml of concentrated hydrochloric acid and 30 ml of dioxane. After addition of 2.16 g · anisole, the solution is heated to boiling for 4 hours. The dioxane was distilled off under reduced pressure and the remaining hydrochloric solution was washed with ether. The aqueous phase is concentrated to dryness under reduced pressure.

Water was added to the residue and the solution was adsorbed on a column packed with Dowex 50X4 (H type). After washing with water, the adsorbed material was eluted with 4 N aqueous ammonium hydroxide solution. The eluted solution is then _; thicken to dryness under reduced printing.

When acetone / water (1: 1) was added to the residue, crystals formed which were filtered off. 0.82 g of (2RS, 3R) -isobutylisossria is obtained.

[alpha] i? 2e = ^-8, 3 °. (c = ^ acid). Rf =. 0 ^, 18.

Example 6

If 3.0 g of benzyloxycarbonyl- (2RS, 3R) -3-amino-3-phenyl-3-phenylpropenitrile was reacted as described in Example 1, 0.80 g of (2RS, 3R) -phenylisoserine was obtained.

1

[elf] JDG - + 9.5 ° (h = 1.0, 1N cHorovodíková- Rf = 0.13 and 0.17. for C ₉ H _{3 N} MO calculated: 59.66% C, 6.12% H, 7.73% N; found: 59.18% C, 6.15% H, 7.67% N.

Example 7

When 10.3 g of Z- (2RS, 3KS) -33arnino-22hydrooy-4 _"C p hlorfenylbutyro" nitrile Necla reacted as described in Example 1, yielding 3.10 g · (2RS, 3RS) -3- amino- (cis -droxy-4-β-fluorophenebutanoic acid).

Rf = 0.25.

For C ₁₀ H ₁₂ NO ₃ Cl requires: C 52.29%, H 5.27%, N 6.10%;

Found: C 52.11, H 5.17, N 5.99.

Příklaa8

If 10.3 g of benzyloxycarbonyl-22RS, 3RS) -3-amino-32-pyridinyl-4-oochloophenylbutdroc nitrile was not reacted as described in Example 1, 4.82 g (2RS, 3RS) -3-aminoc2-lydro were obtained; o-Dlorophenyl butanoic acid ·

R f = 0.25 and 0.29.

For С ₁₀ И ₁ 2 ^) ₃ С1

calculated: 52% -9% C, 5% -7% H, 61% N; found: 52 _э 1% C 517% H% N 59H

Example9

When 11.2 g of Z- ^ RS, 3RS) -3 'íaiinno32hl _Γ - _ΓO oχd34hpmiehy-l'enylbutyronitrilu not respond as described in examples 1, yielding 3.41 g of (2RS, 3RR) -3-aminoз2зlddroxy-4зphiithylfend0but ^ lic acid ·

Rf = 0.20.

For C _n H ₁₅ NO ₃ Calculated: C% 6314, 7 ^ 3 Kt, 6,999% 'N;

Found:% C, 4% C, 7.01% H, 6.79% N.

EXAMPLE 10

22.0 g of oily benzyloxycarbonyl-22RS, 3RR-3-amino-2-hydroxy-4-pyridinylbutyloliril are dissolved in a mixture of 200 ml of concentrated hydrochloric acid and 200 ml of dioxane. After addition of 13.2 g of anisole, the reaction mixture is heated to boiling for 12 hours.

The dioxane was then distilled off under reduced pressure and the remaining aqueous solution was washed with ether. The aqueous phase is concentrated under reduced pressure and evaporated to dryness. Water (100 ml) was added to the residue and the insoluble material was filtered off. After addition of the same amount of acetone, the pH is adjusted to 5.5 with aqueous ammonia. The mixture was allowed to stand in the fridge. The precipitated crystals are filtered to give 6.73 g of the desired (2RS, 3RS) -3-amino-2-hydroxy-4-p-yersulfonylbutyl-butyric acid.

If 30 g of benzyloxycarbonyl-C2RS, 3R) -3-amino-2-hydroxy-4-p-hydroxybutylbutyrenesulfonyl were reacted as described in Example 10, 12.61 g (2RS, 3R) -3 were obtained. Loo-2-hydroxy-4-piperidinylphenyl butanoic acid.

Rf = 0.20.

^P ro C ^ H ^ NO ⁴ Calculated: 58.81% C, 5.92% H, 7.82% N;

Found:% C, 58.63;% H, 5.99;% N, 7.43%.

When (2RS, 3R) -3-dimos-2-hydroxy-4-p-hyeroxyphenylbutyric acid is benzyloxycarbonylated using benozon- (S) -4,6-dimethylpyrimidin-2-ylthiocarbamates, and then reacted with dicyclohexylminine, it is obtained benzyloxycarbonyl- (RS, 3R) -3- (4R) -2-hydroxy-4-p-hydroxyphenyl butanoic acid in the form of a dicyclohexylmonium salt.

15.22 g of crude dicyklohexylmoniové sooi 'crystallized from meth8umc> LU · lihylacltáis and petroleum ether to give 3.2 g of optically impure dicyklohexylmoniové sooi bennylo: xykarborn ^r l' (2R, 3R) -3-2-amios hydrsxy- 4-p-Lydroxyphenyl-butanoic acid in a first crystalline subclass.

The mother liquor was evaporated to dryness and the residue was precipitated three times from ethyl acetate and ether to give 5.02 g of the optically pure dicyclohexylimmonium salt of benzyloxycarbonyl- (2S, 3R) -. 3-amio-2-hyeroxy-4-p) yeroxyfenylbsitanové acid, mp 121-122 ° C, [al ^f a] = ⁺⁵ 5 ^»3 ° (c = 4.22, acetic acid).

^p ro ^ ⁴¹ ¾.

calculated: 69.46% C, 8.16% H, 6.17% N;

Found: C, 69.831; H, 8.35; N, 6.42.

Comparative Example 1

Synthesis of nitrile from aldehyde

The solution, seen from 20.8 g of sodium hydrogen sulphite and 50 g of water, is added to about 52.7 g of oily benoyloxycarbonyl, (R) -fluorobenzoate and the precipitated editorial product is separated and washed with water and ether. 77 g of crude sodium salt of 2-benzylcarbonylamino-1-hydroxy-3-phenylpoopene sulphonic acid are thus obtained. .

g of the resulting adduct is suspended in 250 ml of water and brought to 10-12 ° C. After addition of 500 ml of ether, a solution of 13 g of potassium cyanide in 100 ml of water is added dropwise over 30 minutes.

The mixture was then allowed to react for 3 hours at room temperature and the aqueous phase was discarded. The ether phase is washed with an aqueous sodium chloride solution and dried over anhydrous magnesium sulfate.

After filtering off the magnesium sulphate, the ether was distilled off under reduced pressure to give 49 g of oily benzyloxycarbonyl- (2RS, 3R) -3-amino-2-hydroxy-4-phenylbutyronitrile.

The other nitriles used in the present invention are prepared in a manner similar to the above.

F

Comparative Example 2

2.25 g of benzylo-4-carbornyl 4- (2S, 3 R) -4,4-amino-4- (2-carboxy-4-carboxy-4-carboxylic acid) - The acid is dissolved in 70 ml of tetrahyerofuran and after addition of 2.75 g of the benzyl ester salt by isolation with p-toluenesufoonic acid, the reaction mixture is not stirred for 0.98 ml and cooled to -5 ° C. 1.40 g of Ν, Ν'-decyclohexylcarbodiimie was added and the reaction mixture was allowed to stand for one night.

The tetrahydrofuran was then separated under reduced pressure, 200 ml of ethylene acetic acid were added to the residue, the insoluble matter was separated off, the filtrate was washed with 1 N sulfuric acid solution, water, 5% aqueous sodium hydrogen chloride solution and water (exactly in that order). The organic solution is dried over anhydrous magnesium sulphate. The filtrate was bent under reduced pressure, and the residue solidified upon addition of a mixture of acetic acid and petroleum ether. . Crystallization from the same mixture of solvents yielded (2S, 3R) -3-ethylamino-2H-diol-4-phenylbenzoyl- (S) -leucine.

^The substance has ^t . ^t . ¹²² 'and ^{from 12} 3 ° C, [Al ^f a] ^{2 g} + 3 H ^{2 (c} = ^1, kyseltoa octováh ^for с ^ _н зб и _{2 о} б calculated: 69.90% C, 6.1 - % H, 5.26% 3 N;

found: 69.66% C, 7.0% H - 513% N.

In 100 ml of methanol rozpuutí 3.22 gigabytes ^ j ^ s ^ j ^ yl ^ j ^ eenzyltχykarbonnУ- ester and (2S, 3R) ² -3hemiooh hhyeroχy fnnyleutenoylh-4 (S) -leucine and after addition of 100 mg of palladium black the solution is hydrogenated for 3 hours.

The catalyst was decanted off and the filtrate was bent under reduced pressure. Crystallization of the residue from a mixture of methanol and acetic acid gave 889 mg of (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyl-4 (S) -fluoric acid. Mp 208-213 ° C (dec.). .

^[ al ^f a ^] | ^g = -23. ⁵ DEG ^(c = ^{0, 48,} tyselina acid). Rf value = ⁰ 48th

^p ro ^ ¾ ⁴ ^ ⁴

calculated: found: 62 :) 1% c, 62.61% C, H, 7.85; N 9.09; H, 7.86; N, 8.83. Comparative example 3

A mixture of 85 mg of N -hydroxy-benzyl ester of benzyloxycarbonic acid (3S, 3R) -3-methyl-2-hydroxy-A-phenylbutyrate and 86 mg of (S) -Isulfurobenzyl ester salt with p-tolon-sulfonic acid was allowed to react for 12 hours in an environment of 5 ml dioxane at room temperature in the presence of 0.03 ml triethylamine.

The reaction mixture was quenched under reduced pressure, and the residue was extracted into ethylene acetic acid. The solution in this solvent is washed with 1 N sulfuric acid solution, then with water, 5% aqueous sodium hydrogencarbonate solution and water, respectively.

After drying over anhydrous magnesium sulfate, a solid residue is obtained by distilling off the solvent, which is recrystallized from ethyl acetate by addition of petroleum ether. 90 mg of benzyl ester of benzenesulfonylcarbonyl-2S, 3R) -3-amino-10-hydroxy-4-phenylbutenoolleucine are obtained.

Mp 122 ° C, [alpha] - + 15.1 ° (o = 0.77, acetic acid).

Similarly, as described in - Comparative Example 2 was obtained from 75 mg of benzyl benByloxykarboxnrl- (2S, R) -3-Amir ^ C ^ ^ O2 -] ^ h ^ d ^ dro ^ '^ --f ^of (^ (2S, 3R) -3-Amino-2-yl-2-yl-4-phenyl-butyiool- (S) -leucine 33 mg of (2S, 3R) -3-amino-2-yl-4-phenyl-butyiool. .

^La t ^k and a [Al ^{f a]} _2γ Θ -. 21.8 ° ^(c - ^{0, 45,} tyselina acid) '· ^f = ^0, the 48th

Comparative Example 4

A mixture of 330 mg of benzyloxycarbonyl-1- (2S, 3R) -34-amino-2-hydroxy-44-phenylbutyric acid, prepared similarly to that described in Example 1, 162 mg of N-hydroxybenzotriazole, 217 mg of methyl-γ-ester (S-leucine, 0.17) ml of tree oil and 206 mg of N, N4-dicyalohexylcarodiimide The reaction was carried out according to Example 1, whereby 293 mg of Oenzyloxy: α-amino-(2S, -3R) -34-amino-2-methyl-4-phenyl-butanoyl (phenyl) -butanoyl (phenyl) butanoyl (phenyl) butanoyl. -loucinu.

The substance has tt'12O DEG C. [alpha] | γ ₈ - 22J ⁰ (c = 1.09, Acetic kyseinaa).

^For calculated: 65.77% C, 7.07% H, 6.14% N; Found:% C, 65.91;% H, 7.16;% N, 6.28.

Dissolve 250 mg of meet (1S, 3R) -34 amino-42-amino-4-phenoxy-4-oxoyl-10-ol (S) -lstciot in 10 ml of methanol and add 0.6 ml of 1 N aqueous sodium hydroxide solution and the reaction mixture. is stirred for 6 hours at room temperature.

The methanol was further distilled off under reduced pressure, water was added to the remaining liquid, and the pH was adjusted to 2 by acidifying with 1N sulfuric acid. The precipitated portion was extracted into ethyl acetate, washed with water, then dried over anhydrous sulfate. magnesium and the solvent is removed by filtration after filtration. The residual solid was recrystallized from a mixture of ethyl acetate and petroleum ether.

Repeatedly ^, ^ and ^ 1 ^ i ^]. ^ I pupils gave 150 mg bθnzylo: χlaarbooylι · (2S, 3R) -3-amino-4-24hydrox feln ^г 10utanoyll (S) leucine 41.

Mp 209 [deg.] C. [.alpha.] D @ ₂₀ +27.2 DEG ( ^c = O @ 2 = acetic acid).

^P ro ^{C 4 H} 2 ^N 2 about ₃ ° ^6: calculated: 65.14% C, 6.75% H, 6.46% N;

Found: C, 65.29; H, 6.75; N, 6.33.

120 mg of Oenoylo [beta] -carbonyl- (2S, 3R) -3-amino-2-halo-44-ol / 4-butanalol-(S) -leuide were dissolved in 30 ml of methanol and, after addition of 50 mg of palladium black, the reaction mixture was hydrogenated for 3 hours.

The catalyst was filtered off and methanol was removed from the filtrate. 54 mg of (2S, 3R) -3-amino-42-hydroxy-4-phenylbotaaoyl-1 (S) -sucine (obtained from methanol and ethyl acetate) are obtained.

[alpha] | 7 ^g - 22.7 ° (c = 0.95, Acetic acid) ^Rf = 0.48.

Comparative Example 5 mg of benzyloxy / carbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyric acid N-hydroxysuccinimido is reacted with 54 mg of (S) -succin in 5 ml of dioxane and 5 ml of water, of 0.03 ml of triethylamine on standing for 2 days at room temperature

The reaction mixture is concentrated under reduced pressure, 30 ml of water are added to the residue, and the pH of the solution is adjusted to 2 by the addition of 1N sulfuric acid. The acidified solution is extracted into ethyl acetate (50 ml), solvent ue washed with 1 N sulfuric acid, water, 5% aqueous sodium hS <rr05 ^en УoUtiδitlnu solution and water in that order, after which the UE is dried over anhydrous SiR ^ em sulfate, filtration UE ethyl acetate by-ESI-accelerate, and the residual solid was recrystallized from ethyl acetate by addition of petroleum ether.

Using crystals from the same solvent combination, this yields 63 mg of the expected benzylcarbonyl- (2S, 3Ю-3-ynyl) -2H-dihydro-4-phenylbutal-4-ene (S) -leucine.

Mp ²⁰⁹ ° C [alpha] | ^{4 g +} 26, ^{7 (c} = ⁰ 5 ⁵ tyselina octováh

In addition, 45 mg of benzylcarbodine- (2S, 3R) -3-amino-22-hydroxy-44-phenylbutylbutyl (S) -leucine in a similar manner to that described in Example 4 yields 15 mg of besatin.

^L has ^the AT and ^{[Al f a] 24 g} -. 2 ² 3 ^(c = ^{0 5 2} tyseltoa acid ^Rf = ^0.48 ^ «

Comparative example 6

Similar to Example 2, 330 mg of B-leucine benyydestyrate salt was reacted with 330 mg of 3-amino-2-hydroxy-4-phenylbutyric acid. p-toluenesulfonic acid, 162 mg of N-hydroydenyl Criazole, 0.14 ml of methyl ethyl and 206 mg of Ν'-dicyclotexylcarbodilic acid, and to prepare 450 mg of benzyl ester of benzylcarbonyl- (2S, 3R) -3-amino-o-o-2- R) leucink.

^L atka has it that it Я9 to ⁸ ° C, ^[Al and ^f] |? ^{g + 40.0} ° ⁽ c = acetic acid)

When treated with 430 mg of benzyl ester benzylcarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-hydroxybutyl-4-benzyl-R-succin in a similar manner as described in Comparative Example 2, it yields 144 mg ( 2S, 3R) -3-Amino-22-nitrocyl-4-trifluorobuyl-(S) -leucine.

[elfl] 2 ^ + 0 ^g, 9 ° (c = ^{0 47 c} ct ^c kyselica VA). Rf = ^{0, 4,} 9 ' ^P ro ^C16H24NgO _4: calculated: 62.31% C, 7.85% H, 9.09% N;

Found:% C, 62.47;% H, 7.89;% N, 9.09.

Comparative Example 7 '

A mixture of 660 mg of benzylocarboxylic boron- (2S, 3S) -3-amino-2-hydroxy-4-phenylenedioic acid, 944 mg of benzyl ester (S-leucine as sodium for p-colloSulconic acid, 324 mg of N-hydroxybenzyl nitrile), 0.34 md of triethylamine and 412 mg ^of ric-ex-lot-lotdlotexlotexborlotborborbore were treated similarly to that described in Comparative Example 1 to give 710 mg of benzyldicarbodecarbonyl- (2S, SS) -3-amino-22'-dihydro-fen 4-phenylbutyl-succin (S).

Mp 128 ° C, [.alpha.] D @ ₂₀ = 38.0 DEG (c = 0.81, acetic acid).

From 500 mg of benzyloxycarbonyl- (2S, 3S) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -leucine benzyl ester, 270 mg of (2S, 3S) - was prepared similarly as described in Comparative Example 2 - 3-amino-2-hydroxy-4-phenylbutanoyl- (S) -leucine.

[α] D -61.3 ° (c = 0.56, acetic acid). Rf = 0.37.

For calculated: 62.31% C,

H, 7.85; N, 9.09;

Found:% C, 61.99;% H, 7.71;% N, 9.13.

Comparative example 8

A mixture of 165 mg of benzyloxycarbonyl- (2S, 3S) -3-amino-2-hydroxy-4-phenylbutyric acid, 237 mg of (R) -leucine benzyl ester with p-toluenesulfonic acid, 81 mg of N-hydroxybenzotriazole, 0.08 ml of triethylamine and 103 mg of N, N &apos; -dicyclohexylcarbodiimide was treated in a similar manner as described in Comparative Example 2 to prepare benzyloxycarbonyl- (2S, 3S) -3-amino-2-hydroxy-4-phenylbutanoyl- (R) benzyl ester. -leucine (180 mg).

Mp 128-129 ° C, [alpha] - 11.0 ° (c = 1.02, acetic acid).

Similarly, as described in Comparative Example 2, 72.1 mg (2S) was obtained from 155 mg benzyloxycarbonyl- (2S, 3S) -3-amino-2-hydroxy-4-phenylbutanoyl- (R) -leucine benzyl ester. 3S) -3-amino-2-hydroxy-4-phenylnutanoyl- (R) -leucine.

[α] - 31.4 ° (c = 0.47, acetic acid), R f = 0.52.

For C ₁₆ H ₂₄ N ₂ 0 ₄

calculated: % C, 62.31;% H, 7.85;% N, 9.09; found: H, 7.63; N, 8.80.

Comparative example 9

A mixture of 330 mg of benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyric acid,

405 mg of glycine benzyl ester with p-toluenesulfonic acid, 162 mg of N-hydroxybenzotriazole, 0.14 ml of triethylamine and 206 mg of N, N'-dicyclohexylcarbodiimide were treated in a similar manner to that described in Comparative Example 2.

There was thus obtained benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutanoylglycine oily benzyl ester. Thin layer chromatography testing revealed that the product was uniform and pure.

Treatment of the oily product according to the procedure of Comparative Example 2 gave 117 mg of (2S, 3R) -3-amono-2-hydroxy-4-phenylbutanoylglycine [α] D-7.4 ° (c = 0.49, acetic acid). Rf = 0.26.

For C ₁₂ H ₁₆ O ₄ N ₂ O ₃ OH Calculated: C 54.92, H 7.09, N 9.87;

Found: C 54.33, H 6.87, N 9.69.

Comparative example 10

A mixture of 330 mg of benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyric acid, 440 mg of (S) -benzylserine o-nitro-benzyl ester hydrochloride, 162 mg of N-tydro-4-benzenaziazole, 0.14 ml of triethylamine and 206 mg of Ν, Ν'-dicyclohexylcarbodiimide were treated similarly to that described in Comparative Example 2 to give 540 mg of benzyloxycarbonyl- (2S, 3R) -3-ethylene-2-hydroxy-4-nitrobenzyl ester. -butylbutano-1- (S) -benzylserine.

The substance had mp 121.5 ° C; [alpha] | ув ^{+1 7>} 7 DEG ^<c = 1'07 'acetic acid).

Treatment of 500 mg of benzyloxycarbonyl- (2S, 1R, 4-amino-4-phenyl-butanoyl- (S) -benzylserine p-nitrobenzyl ester) according to the procedure of Comparative Example 2 gives 147 mg of (2S, 3R) -. [.alpha.] .alpha.-pyrrole-.beta.-4- (1-enylbutnoyl) -4.0 < - &gt; 20 May

For ^C 13 ^H 15 O ₅ ^N 2 · ^H 2 ° calculated: C 51.99 (5.7 1H, N 9.33;

found: 5b69 # C, 6.74% H, 8.97% N.

Comparative example 11,

A mixture of 330 mg of benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyric acid, 400 mg of (S) -glutamine benzyl ester hydrochloride, 162 mg of N- (trifluoromethyl) -Oriazole _> 0.14 ml ti Ethyl amine and 206 mg of Ν, Ν-dicyclohexylcarbodiimide were treated analogously to Comparative Example 2.

There was thus obtained (2S, 3R) -3-amino-22-ethyl-yoyl-phenyl-butanoyl- (S) -glutimine benzyl ester of benzoylcarbonylcarbonyl ester.

^{L 1} atka has mp 55 ° ^C; [alpha] ⁺ 62.3 ° ^(c = ⁰ 7 ^5-acetic tyselina

180 mg of benzyloxy-4-carbonyl- (2S, 3R) -3-amino-22-hydroxy-2,2-phenylbutayoyl- (S) -glucamine benzyl ester were treated similarly as described in Comparative Example 2 to give 61 mg of (2S, 3R) -3- ( N-amino-2-pyrrolyl-enylbutyl- (S) -gin.

^L atka having ^[alpha] - ^9, 9 ° ^(c = ^0. 37 '^ ^ sellna acetic acid Rf = ^0.1 7th

H, 7.1;% N, 11.69. .

Found: 50> 64% C 6 _F 22% H ^ H IfcN.

Comparative example 12

A mixture of 330 mg of benzylcarbonyl- (2S, 3 R) -3-amino-2-carboxylic acid (2 S, 3 R) -3-amino] - (2 R, 3 R) - (2 R, 3 R) - (2 R, 3 R)). : Linear acids,

600 mg of p-lueysulfonic acid (S) -O-benzylglutamine benzyl ester salt, 162 mg of N-hydroxybenzofrazole, 0.14 ml of triethylamine and 206 mg of Ν, Ν-diclk-hexylcarbidimide are treated with work up similarly as described in Comparative Example 2c to give 605 mg of benzyl-4-carboryl- (2S, 3R) -3-amino-2 (trifluoryl) -phenylbenzyl C (O) -1- (S) (O-benzylglutamine) benzyl ester .

^The substance has -t. t ,. ^116, GB ¹ 2 ^{0 '[Al f} a] 28 + ⁶ ° ^(c = ^{1, 0} 4' ^ selinc acid).

Treatment of 250 mg of benzyl 4-carbonyl-1- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) -O-benzylglutmin benzyl ester according to Comparative Example 2 gives 166 mg of (2S, 3N) -3-Lmino-2-hydro: Q1,4-phenyl-utazoyl- (S) -glucidate.

^L atka having [alpha] ^{| 3} ₈ - 18.4 ° (c = ^ 51 ^alkyl Selina acid), ^R = ^0, the 27th

^{For C} 15 ^H 20 ^O 6 ^N >

calculated: 55.55%> C, <5.22% H, 8.64% N;

Found: 55 »20% C; 5.90% H;

Comparative example

A mixture of 330 mg of benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyric acid, 455 mg of (S) -valine bennylester salt with p-toluenesulfonic acid, 162 mg of N-hydroxybenzolriazole, 0.14 ml of methylbenzyl and 206 mg of mg, Ν'-dicyclin-1-carboxylic acid are treated similarly to that described in Comparative Example 2.

To give 397 mg of O-biennylesteru -enzyln: χrkarb-NNL "(2S, 3R) ^-3-r aminzo2 lhУ ^{ooyly-4-4fbnyl} utшlznl- (S Svalina.

L ^and T has Units. · I. ^96.5 to 97 ° C ^{[alpha] 19 +} 2 DEG ^(c = tyseltea acid).

When 376 mg of benzyl ester: β-carbonyl-nz- (2S, 3R) -3-amino-2-amino-4-phenyl-utanool- (S) -valizl was treated in a similar manner as described in Comparative Example 2, 104% of the title compound was obtained. mg of (2S, 3R) -3-aminZl2-hhldOly-44ffbzllbltnzyl- (S) -valizu [alpha] 5γ ₈ - 1.4 ° (c = 0.55, k _y Selina acetate) Rf = 0.42.

For C1 _{5 2} H 2 O ₄ N ₂ Calculated: C 61.20%,

H, 7.53; N, 9.52.

Found:% C, 60.98;% H, 7.59;% N, 9.28.

Comparative example

A mixture of 330 mg of -enzyl-1-carbonyl-N- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyric acid, 455 mg of benzyl ester with p-toluenesulfonic acid, 162 mg of N-hydroxy-2-hydroxybutyric acid. ^! 0.14 ml of triethyl-tris and 206 mg of Ν, Ν-dicycloxycarbodide are treated similarly to that described in Comparative Example 2.

There were obtained 310 mg of benzyl ester-benzyl- (2S, 5R) -3- (2S, 5R) -3-aminol-2-hydroxy-1,4-benzyl-utazinyl-11- (S) -Z-valvalizyl.

^{La to} t and tt is ¹¹⁴ and ^z. И5 C ^{[Al f a]} || ⁸ + 22 ^{0 (c} = ^{0, 49,} Selina ^alkyl acid).

Treatment of 250 mg of benzylbenzyl (12-carbo) nzl- (2S, 3R) -3-ambzo-2-hydrobutyl- ^4- benzyl-lthiazole- (S) -zalzole according to the procedure of Comparative Example 2 gives 110 mg ( 2S, 3R) -3-Aminol-2-hyddolyl- ⁴ -phenylbutan- ⁴ -yl- ¹ H -alpha.

[alpha] ^{] 8} - M, 5 ° ^(c = 50 ^ tysellna acid), ^R = ^{0, the 41st}

For C ₁₅ H 22 O 4 N 2 calculated: 59.33% C, 7.64% H, 9.23% N;

found: 59.42 C. 7 ^ 8 H. 9.99% N.

Comparative example

Dicyclohexylamine was removed from 952 mg of tert-butyloxycarbonyl-tert-3R) -3-amino-2-hydroxy-4-phenylbutyric acid by treatment with 1N sulfuric acid and ethyl acetate to give the oleic acid tert. butoxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyric.

This oily portion and 440 mg of (S) -methionine methyl ester hydrochloride were suspended in 20 ml of tetrahydrofuran and the reaction mixture was neutralized by the addition of 0.308 ml of triethylamine.

The solution thus prepared was cooled to -5 ° C, 412 mg of N, N-dicyclohexylcarbodiimide was added, the reaction mixture was stirred at room temperature for 4 hours, the precipitated crystals were filtered off and the mother liquor was concentrated under reduced pressure.

The residue was dissolved in 200 ml of ethyl acetate, washed with 1 N sulfuric acid, water, 5% aqueous sodium thyrogate and then again with water, in that order, after which the organic solution was dried over anhydrous magnesium sulfate.

The residue obtained by concentrating the mother liquor under reduced pressure is dissolved in a 2: 5 mixture of ethylene chloride and benzene (2: 5), and the solution is applied to a column of chromatography G (Meeck). ).

The fractions containing the expected material were collected, the solvent was removed under reduced pressure, and recrystallization of the solid residue from ether / kerosene gave 380 mg of tert-butoxy- (2S, 3R) -3-amino-trimethyl-4-ethyl ester. -phenolbbUthaliol- (S) -methionine.

Mp ¹¹⁸ ° C, [alpha] C ⁵ 1.4 ° (c = ^0.5 б ^, acetic acid)

When treated according to the procedure described in Comparative Example 4,280 mg of methyl ester (3R) -3-amino-22-oxo-14-fetylbutanol-(S) -methionine, 200 mg of tert-butoxy- (2S, 3R) was obtained. -3-Aminite / hexyl-phenolbutyl-(S) -methionine.

^L and let ^K has m. ^P. 16 ° C ^{(foaming [alpha]} 22 ^{g +} 44 ¹ ° (c = ^{0 5 4,} acetic acid).

To 150 mg of tert -butyl- (2S, 3R) -3-dimethylamino-phenoxybutyl- (S) -methionine was added 0.2 ml of titiglutic acid and a 50% aqueous solution of trifluoroacetic acid, and the reaction mixture was stirred hour at min.

The oily portion obtained by concentrating the reaction mixture under reduced pressure is dissolved in water and adsorbed on a 5 ml column packed with Dowex 50X4 resin (in a hydrogen cycle).

The adsorbed material was eluted with a 2 N aqueous solution of ammonium, and then concentrated to dryness to give a solid material from the eluate. Crystallization of this portion from water / acetone (1: 1) afforded 87 mg of (2S, 3R) -3-amino-2H-dioyl-4-methoxybenzamide (S) -methiol.

[Al ^{f a]} - ²⁰ · 4 ° ^(c = ^{0, 49,} tyselina acetic ^ ^f = ^0, the 37th

H, 65.11; N, 8.35.

Found:% C, 53.64;% H, 6.47;

Comparative Example 16

A mixture of 330 mg of 3-amino-2-hydroxy-2-hydroxy-4-phenylbutyric acid bonza [beta] -carbonyl-tert -butyl-4-phenylbutyric acid, 472 mg of (S) -isulfonic acid with p-toluenesulfonic acid, 162 mg of N hydroxybenzoic acid, 0.14 ml of triethylamios and 206 mg of Ν, Ν-ykl-cyclicylcarbseiimies were treated in a similar manner to that described in Comparative Example 2 to give 466 mg of benzoyloxycarbonyl-tS, 3R) -3-amino-2-sulfonoyl-sulfonoyl-sulfonoyl-sulfonic acid. ) -isslesiios.

The substance has a melting point of 100 to 101 ° C, [alpha] Ц + 23.6 ° (c = 1.0, acid

Treatment of 440 mg of the above substances with the procedure of Comparative Example 2 gave 191 mg of (2S, 3R) ^{"3-aminos22hyderoyy4--,} enoУbbSiaosl·L (S) -issllUiinu.

[alpha] | γ + ^{8 6} 4 ° ^(c = ^0.5, acetic kyselka 2) ³ · ^0, 4 ^8th

^For 5¾ ⁴ ° ^ ¾ ⁴

calculated: 62.1 · * C · 7 * 85 H · 9.09%> Ni found: 62.1 · C · 7.70% H · 8 ^ 9 * K. Comparative Example 17

A mixture of 330 mg of benzylo: cis-cacbosoll (2S, 3R) -3-amino-2-hydroxy-4-phenoxybutyrone, 472 mg of (S) -osulfonyl salt with p-toluenesulfonic acid, 162 mg of N-hydroxybenoisulfonyl 0.14 ml of trlethyamiosis and 206 mg of β, β-ε-cyclopropyl alcohol were treated similarly as described in Comparative Example · 2 to give 437 mg of benzoylcarbonyl-C 2 S, 3 R) -3-amino-2-benoyl ester. hyddos-4-phenolbsaaosII (S) -osrIISiiou.

^{Substances AM mp 114} ° C ^from an H5, [Aatami] ^ ⁸ + 23.3 ° ^(c = ⁰ »94 tyaelina acid).

Starting from 350 mg of the above compound, 160 mg of (2S, 3R) -1-amino-2-yl-2-yl- ^4- phenylbenzanosulfonyl (S) -sulfonate was obtained using the procedure of Comparative Example 2.

^Substances and has a specific rotation of [al and ^f] | YG - 8.9 ° ^(c = ⁰ 7 ⁰ tyseltaa acid) ^Rf = ⁰ · 47 ^T ro 0 ^ 0 ^ ² ° Calculated: 58.88% C · 8.03% Η · β, 58% N;

Found: 58.95% C · 7.8 ⁴ % H, β ^ ^% N.

Comparative example 18

A mixture of 330 mg of benzylsycarbosylic acid (2S, 3R) -3-amino-2-hydrosulphonyl-4-phenylbutyric acid, 505 mg of soylbenzoic acid salt (RS) -emooctanoic acid with p-tolueotulonic acid, 162 mg of N-hydroxysulphosulphazole. 14 ml of triethylamine and 206 mg of N, N ' -ethylsilylbodiimide were treated in a similar manner to that described in Comparative Example 2a to obtain 430 mg of benoyltrole of benoyloic acid: [ ₍ R) -carbosol- (2S, 3R) -3-amino-2-halo] 14-Flutylbutyl-(RS) -2-mlsulfonyl.

Mp 90 DEG-92 DEG C., [.alpha.] D @ 20 +32.9 DEG (c = 0.94, acetic acid).

Zpraccuee-n, 250 mg of the above compound Following the procedure of Reference Example 2 to give 74 mg · (2S, 3R) -3-amio-2-hydrs: χl ⁴ -fl0l1butlOΌyl- (IS>) 2 2cадlOnosktloové.

The substance has a [alpha] - 34.9 ° (c = 0.53, acetic acid), Rf = 0.58.

^{For C} 18 ^H 28 ° 4 ^M 2 calculated: 64.26% C, 8.39% H, 8.33% N;

Found:% C, 64.43;% H, 8.18;% N, 8.02.

Comparative example 19

A mixture of 330 mg of benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyric acid, 513 mg of (S) -phenylalanine benzyl ester salt of p-toluenesulfonic acid, 162 mg of N-hydroxybenzotriazole, 0.14 ml of triethylamine and 206 mg of Ν, Ν'-dicyclohexylcarbodiimide was treated similarly as described in Comparative Example 2 to give 378 mg of benzyloxycarbonyl- (2S, 3R) -3-amino-2-hydroxy-4-phenylbutanoyl- (S) benzyl ester -phenylalanine.

Fabric Mint. mp 111-112 ° C, [α] D 25 + 39.6 ° (c = 0.97, acetic acid).

Treatment of 355 mg of the title compound as described in Comparative Example 2 afforded 108 mg of (2S, 3R) -3-amino-2-hydroxy-4-phenylbutyl-(S) -phenyl-alanine.

The substance has a specific rotation of [alpha] + 0.4 ° (c = 0.46, acetic acid), Rf = 0.44.

For C ₁₉ H ₂₂ N ₄ O ₂ .CH ₃ OH: calculated: 64.15% C, 7.00% H, 7.48% N;

Found:% C, 63.58;% H, 6.45;% N, 7.57.

Comparative Example 20

When the mixture is treated with 1.10 g of oily benzyloxycarbonyl- (2RS, 3R) -methylisoquinone prepared by benzyloxycarbonylation of (2RS, 3R) -methylisoserine, 1.97 g of (S) -leucine benzyl ester salt with p-toluenesulfonic acid, 675 mg N -hydroxybenzotriazole, 0.70 ml of triethylamine and 948 mg of Ν, d'-dicyclohexylcarbodiimide as described in Comparative Example 2 to give oily benzyloxycarbonyl- (2RS, 3R) -methylisoaeryl- (S) -leucinue

According to evaluation by thin layer chromatography, the oily product is pure.

Working up the oily product as described in Comparative Example 2 gives 765 mg of (2RS, 3R) -3-amino-2-hydroxypropionyl- (S) -leucine.

The substance has a specific rotation of [alpha] γ-28.7 ° (c = 1.09, acetic acid).

For ^C _T O ^{N H} 2 O 2 ° 4: calculated: 51.70% C, 8.68% H,

12.06% N;

found:

C 51.49, H 8.42, N 11.96%

Comparative example

1.32 g of oily benzyloxycarbonyl- (2RS, 3R) -isobutylisoserine prepared by benzyloxycarbonylation of the (2RS, 3R) -isobutylisoserine obtained in the above process, 1.76 g of (S) -leucine benzyl ester salt with p-toluenesulfonic acid, 60 g of N-hydroxybenzotriazole, 0.63 ml of triethylamine and 0.92 g of Ν, Ν'-dicyclohexylcarbodiimide were treated as described in Comparative Example 2 to give oily benzyl oxycarbonyl- (2RS, 3R) -iso-butyl-iso-eryl- (S) -leucine. Working up in a similar manner to that described in Comparative Example 2, 0.37 g was obtained. (2RS, BRJ-β-Aminoacetyl-droyil-methylheeananil) SS-leuccin.

^And has an speeificlcou rotation [al and ^f] | jQ - 26.6 ° (c = 1.0 tyselina octováRf = ^0, 43 and 0.49.

^{° C} 13 ^H 26 ^N 2 ° 4

calculated: 56,9 1 % c, 9.Э5% H, 10: 11% N; ycyezeoo: 57.01 «C, 9.00% H, 10.14% N. Comparative example 22nd

A mixture of 2-0-mg benzyloyykarbonnl C2RS, 3R) -phenylisoserinate mp 170-171 ° C, ral, and ^{f] | γ} _eighth + 18.5 ° ^(c = ⁰ 47 tyselina acid - ^ benzyloyytorbonylovtaím obtained ^(2RS, 3R) phenylisoserine bennylesteru salt 394 mg of (S) ileucinu kiselinou with p-toluenesulfonic acid, 130 mg of N-Уldroзyrloeozlгiazolu, and 0.14 ml triehliiaminu 16 mg of N, N * dicycloylcarboxylic acid was treated similarly to that described in Comparative Example 2.

28 mg of benzyl benzyl ylacartonyl- (2RS, ZN) -T-piperazin- (8 J-fluorin) benzyl ester is obtained.

The substance has a TT ^and 93 ° C, from 9- [alfalHQ - 34.7 ° ^(c = 0.49, k ^ ^ Eli acid).

Treatment of 200 mg of bennyloxycarbonyl- (2RS, 3R) -phinyliolseyl- (S) -leucine benzyl isomer according to the procedure of Example 2 gives 4- mg of (2RS, 3R) -3-amino-2-yldooyl-phenyl-phenyl-thiooyl (S) -leucine.

[al ^fa ]. ¹ . -. 2.4 ° ^(c = ⁰ 3 ³ tyoelioa acid).

Pro. C ^ 22 ^ 2 vlcounted: 61.20% C, 7, δ3% H 9 ^ 2% N; o-Iiz-oo: 60.92% C, 7.67% H, 9.46% N. Comparative Example 23

A mixture of 1.81 g 0enzyluyУcarbonyl- (2RS, 3R)) -3-amino-2-yidroxy -p-cyiorfenylmásel.né kiselini ^{t. t.} 14 ⁷ C, prepared enzylly ^{b l l} carboxylic ^b nn OC ^and it tyselin ^{1 (R} 2) 3 ^-R 5) -3iamino-2i droyy ^{yl-4-p-cУlc)} feonylmáoelyé, OOH 2.37 g of benzyl ( The s-ileucine with p-trifluoromethyltin, 810 mg of N-hydroxysyrloeozlriazole, 0.84 ml of triethylamine and 1.03 g of N, N-dicyclinylcarodilimide were treated similarly to that described in Comparative Example 2.

There was thus obtained 2.0 g of benzyl-1-cyanocarbolyl- (2RS, 3) -3-amino-12-hydroxy-4-picyclo [eta] &lt; 6 &gt; -18-eucine.

Substances ^and AA TT 122 ^and from 124 DEG C. [alpha] | 7 ^g - 12.3 ° ^(c = 3 ^ kyselica acid).

1.00 g of the above-mentioned compound was obtained as described in Comparative Example 2 to give 6-0 mg of (2RS, 3R) -3-aminophenyl-4-p-1-chlorophenylbutyl chloride. - (S) 1 -leucine.

^{The substance} ka has a specific rotation [alpha] ^{| γ8} - 11.3 °. ^(C = ^1.33, lctová acid) ^Rf = ^0, 42 = 0, -2.

For C 16 H 23 N 2 O 4 Cl calculated: C 56.05%, H 6.76%, N 8.17%; Found: C 55.88, H 7.29, N 8.11.

Comparative example

A mixture of 726 mg of benzyloxycarbonyl-IBRS, 3R)) · 3-lamnoo2 "HY-Cl'ooyc - c) _{chlorobenzimidoyl} ftnylmáselné mp 136.5 ° C, prepared benzyloxykarbonylováním (2RS, 3R)" 3-amino- 2-Chloro-4-O-chlorophenylbutyric acid, 944 mg of benzyl ester salt (S-leucine with p-toluenesulfonic acid, 324 mg of N-chlorobenzoic acid), 0.34 ml of thiamine and 412 mg of dicyclohexylcarbodiimide as it was treated similarly. was described in Comparative Example 2.

There was thus obtained 747 mg of benzyl ester (1X, 3R) -3-amino-1H-Croxy-4-c-chlorophenylcarbonyl (S). -ltucine.

Substance ^and has mp 129 HZ 132 ° C - 29.1 ° (c = ^1.02, ₁ SOUR acid).

Go away! with 640 mg of benzyl ester of benzo [eta] [eta] &lt; 5 &gt; -ethoxy] -1- (1-BS, 3RS) -amino-S-hydroxy-4-oxo-C-(S) -ltucine similarly as described in Comparative Example 2, 107 mg of (2RS, 3K) -l-2-hydroxy-2-hydroxy-4-yl-benzoyl-(S) -ltucine was obtained.

^L atka has speci ^f istic rotation [βΐ γθ ^^ - .20, ⁸ ° ^(c = 0.4, ^8, acidulous oc ^{t OVI) _(R} = 0.4 ^{on the 6th}

H, 6.76; N, 8.17.

Found:% C, 56.21;% H, 6.56;% N, 7.90.

Comparative example

A mixture of 1.57 g of bensyloxycarbonyl-2RS, 3R, 3'-amino-2-hydroxy-4-methyl-4-methylphenylmethyl, mp 166.5-168 ° C, prepared by benzyloxycarbonylation of the acid. Sodium-4-p-methycllenylbutyrate, 1.42 g of benzyeserium salt (S-leucine with p-toluenesulfonic acid, 486 mg of N-hydoxylbenzaziazole, 0.50 ml of 1-hyamine and 618 mg of dicyclo-carbodiimide) were added. is processed analogously according to Comparative Example 2.

Benzyloxycarbonyl- (2RS, -hydroxy-4-phenyl-butane-C- (S) -1-eucine) is thus obtained in oily form.

When the oily product was treated similarly to that described in Comparative Example 2, 497 mg of (2RS, 3RS) -1-1H-amino-2-hydroxy-C-p-methylbutanoyl- (S) -leuciline was obtained.

^L and let ^K has ^R = ^0, and 0 5 ^{8, 53,} [Al ^f a] ^{573-7, 2} ° (c = ^{0, 9, 6, k,} acetic acid).

^{For C} 17 ^H 26 ^N 2 ^O 4 calculated: 63.33% C, 8.03% H, 83.99% Ni;

found: 63.08 53 C, 8.33% H, β, 93% 3 N.

103 ml of bestethics are dissolved in 5 ml of acetic acid with the addition of 5 ml of ethanol, and the reaction mixture is hydrogenated catalytically over platinum oxide for 14 hours. After filtration, the filtrate was concentrated to dryness under reduced pressure, and tritroating the residue with ethyl acetate (61% acetic acid) gave 61 mg of (2S, 3R) -3-loin (1- 2-) after filtration. hydroza-14-cycloheyl-l-a-ll-l (S)-l-alcinl.

The substance having Rf = 0.47 [furo ^f-g - 30.5 ° (c = 0.67, Acetic acid).

Comparative Example 26 ^Pro ^ 6 ^ 0 ^ ⁰⁴

calculated: 61.12% C, 9.62% H, 8.91 «N; found: 62.12% C, 9.52% H, 8.81% N. Comparative Example 27 300 mg suspends in 10 l of concentrated sulfuric acid and 0.3 l. fuming

nitric acid at 0 ° C. The reaction is allowed to proceed for one hour at the indicated temperature; during this time, the suspended product is dissolved. The reaction mixture is diluted by pouring into 100 ml of water and adsorbed on a Dowex 50X4 resin column (hydrogen cycle). Ad8rn-noaoý ma ^ erá! The solution is concentrated to dryness, the residue is taken up in 10 ml of water, the solution is filtered and the filtrate is filtered to give 213 mg of (2S, 3R) -3-amino-12-amino-4- ppnZiгofbln ^r lbulanznl- (S) -lenine.

The substance has [.alpha.] D @ ₂₀ + 5.2 DEG (c = 0.5, acetic acid), Rf = 0.58.

^For ^ 23 ^ ⁵ ° Calculated: 54 ^ 8% C, 6.58% H, 11.88% N;

Found: C, 54.38; H, 6.56; 11.89

Comparative example

100 mg of (2S, 3R) -3-amino-2-hydroxy-4-p-nitrophosphonolbulta-1- (S) -buffer as obtained in Comparative Example 27 are dissolved in 10 ml of oethrool and the solution is hydrogenated for 5 hours on palladium black. . The catalyst is filtered, the filtrate is concentrated under reduced pressure, the residue is dissolved in 5 ml of water, and after filtration, the filtrate is taken up in a yellowish tower. 81 mg of (2S, 3R) -3-amino-2-hydroxy-1,4-p-10-amino-1-benzyl-1- (S) -benzyl is obtained.

^Substance a has [alpha] ^{8 - 23,22} ° ⁽ c = ⁰ , 5, the same) ^{, Rf} = ^0, 25.

^^ ^P ro 3 ° ^4: calculated: 59.42% C, 7.79% H, 13.00% N;

Found:% C, 59.01;% H, 7.91;% N, 12.81.

Comparative example

2.11 g of (2RS, 3R) -3-lincin-2-hydroxy-4-phthalen-2-yl] -phenyl] -butyl acid prepared in Example 10 are dissolved in 10 of 1 N sodium hydroxide solution. Under vigorous stirring and ice cooling, 4.5 ml of benzyl ^is added in three portions over 30 minutes ^{to the} benzylic acid (reagent).

the mixture was stirred vigorously with ice-cooling for 1 hour and at room temperature for 3 hours. During the reaction, the pH of the reaction was adjusted to 8-9 with 1 N sodium hydroxide solution.

After completion of the reaction, 6 N hydrochloric acid was added and the reaction mixture was adjusted to pH 2. The resulting oily product was then extracted twice with 100 ml of ethyl acetate. The ethyl acetate phase is washed with water and dehydrated to dryness using anhydrous magnesium sulfate.

After the magnesium sulfate was filtered off, the filtrate was concentrated under reduced pressure and the residue was crystallized in a mixture of ethyl acetate and petroleum ether. To give 3.64 g of benzyloxycarbonyl - '(2SS, 3RS) -3-Aninka) o2-4hydro2xr-4 "p benzyloxykBrbQnyío: xyťenylbutanové acid t. · T. 138-140 C. <j _e

479 mg of benzyloxycarbones of 1- (2SS 3SS) -3-9mino-2-hydroxy-4-p-benzyloxycarbonyloxyphenylbutonic acid and 162 mg of N-trifluoromethylene oxide are dissolved in 10 adolescent / drofurnau. After the addition of 472 mg of the (S) -leucine benzyl ester salt with p-toluenesuffonic acid, the mixture was added with 0.168 ml of triethylamine and cooled to -5 ° C.

206 mg of N, N-dicyclohexylcarbodiimide are then added and the reaction mixture is allowed to stand overnight. Tetrahydrofuran was distilled off under reduced pressure and 30 ml of ethyl acetate were added. After filtering off the insoluble matter, the filtrate was washed with 1 N sulfuric acid / water, 5% aqueous sodium hydrogencarbonate solution and water, respectively. After drying over anhydrous magnesium sulphate, the filtrate is concentrated under reduced pressure. The residue obtained solidified in a mixture of ethyl acetate and petroleum.

Acidification from the same solvent afforded 450 mg of benzyloxycarbonyl- (2RS, 3R) -phenyl-2-hydroxy-4-p-benzyl-oxycarbonynyl-phenyl-butannyl-4 (S) -leucine bennydester.

Mp. 98-99 ° C, [alpha] ⁸ ® = -14.0 ° ^(c = 0.58, Una · KYSE-acetic acid ^a).

400 mg of benzyloxycarbonyl (RS, 3S) -3-amine-4-benzyloxycarbonyl (S) -leucine (400 mg) are dissolved in 10 ml of methanol and hydrogenated. with 3 hours on 10 mg palladium black.

The catalyst was filtered off and the solvent was bent under reduced pressure. When recrystallization was carried out in a mixture of methanol and ethyl acetate, 219 mg of (2RS, 3R) -1-3-amino-2-hydroxy-4-p-pyrrolidinylbenzanyl (-) - leucine was obtained.

[.alpha.] D @ ²⁰ = -8.8 DEG (c = 0.90, acetic acid), Sf = 0.48 and 0.51.

For ^C 16 ^{H 24 N} 2 O ₅ Calculated: C 59.24, H 7.46, N 8.64.

found: 58.38% C, 7.23% Η, β, 95% N.

Comparative example

Benzylo: χ -carbonyl- (2S, 3R)) * 3-amino-2-] -pyrrole-4-p-hydroxyphenyl-butyric acid, prepared from 1.05 g of benzyl oxycarbonyl- (2S, 3S) -3-amino-2 salt -dlydroxy-4-piperidinylphenylbutanoic acid with dicyclohexylminine in a conventional manner using stlvestate and dilute sulfuric acid, 866 mg of (S) -leucine benzyl ester of p-toluenesulfonic acid, 405 mg of N-hydroxybenzooriazole, N.308 ml of tri-ethylbutylbutylide to react according to Comparative Example 29.

The yield was oily benzyloxycarbonyl- (2S _, 3R) -3-8mino-2-hydroxy-4β-hydroxybenzylbenzyl ester (SEleucine).

If the oily is pure! benzyloxycarbonyloxy (3S, 3R) -3-α-methyl-2-hydroxy-4- (4-hydroxy-4-hydroxy-benzyl) (S) -leucine was reacted as described in Comparative Example 29, yielding 630 mg of (2S, 3Ю-3-aminno32hy rooχ _34hphl-r-0 oχfennlbbUQnnol'o (S) -Ίeueinu.

[.alpha.] D = -19.9 DEG (c = 1.19, acetic acid, Rf = 0.48).

calculated: 60.55% C, 7.30% H,

10.08% N;

found:

% C, 59.98;% H, 7.42;

10.42% N.

Claims (3)

SUBJECT OF THE INVENTION A process for the preparation of an aminoclave generally having the formula I wherein R 1 -CH-CH-COOH II nh ₂ OH (I) ^R 1 is an alkyl of 1 to 4 carbon atoms, or a group of the formula cdclopentyllh1, cdklolexylh1-, cdkloleptdietlk1 where hydrogen, chlorine, methyl, is 0; or 1, ^R 2 n characterized in that the nitrile is generally wherein ^R 1 or as defined above the nitrile derivative 2. A process according to formula III wherein ^R8 is hydrogen, 3. The method according to 2сн ₂ ) _п nitro, lydroxdl or amino and formula III » with a deprotected amino group, hydrolyzed by acid (III) of item 1, characterized in that as the starting compound, where R 1 is a compound of the formula CH ₂ - chloro, methoxyl, nitro, lydroxyl or amino, as defined in claim 1, characterized in that it is a starting material. the compound is applied to the general formula III wherein R 1 is benzyl. merged4. 1. The process of claim 1 wherein the starting compound is a compound of formula III wherein R1 is p-lydroxybennyd.