SU667142A3 - Method of producing 1-/l (-)-gamma-amino-alphaoxybutyryl/-kanamycin a - Google Patents

Description

3 nov1nIffs are carried out with G, 0-1.3 molar amounts of L-(-) - y -benzyloxycarbonylamino-e-hydroxybutyric acid L-(-) - y -benzyloxycarbonylamino acid in dimethylformamide (DMF). Example 1. Preparation of L - (-) - J -beneyloxycarbonylamino-o-hydroxybutyric acid. 7.4 g (0.062 mol) of L - (-) - y-amino-oC-x-butyric acid was added to a solution of 5.2 g (0.13 mol) of sodium hydroxide in 50 ml of water, with stirring and 0-5 ° C for 0.5 hours p Uyk1Sh- 11.7 g (0.068 mol) of carbobenzoxychloride and stirred for 1 hour at the same temperature. The reaction mixture is washed with 50 ml of ether, acidified to pH 2 with dilute hydrochloric acid and extracted with 4 X 80 ml of ether. The extracts are combined, washed with a small volume, saturated with sodium chloride solution, dried over anhydrous sodium sulfate, and filtered. The filtrate is evaporated in vacuo, the residue is crystallized from benzene and 11.6 g (74%) of colorless lamellar crystals are obtained, m.p. 78.5-79 ,. Calculated,%: C 56.91; H 5.97; N 5.53. C, 2 H, 5 N05 Found; C, 56.66; H 5.97; N 5.47. measure 2. Preparation of M-oxysuccinimide ester L - (-) - -benzyloxycarbonylamino. -oxime and Noah. Acids. A solution of 10.6 g (0.042 mol) of the compound obtained in Example 1 and 4.8 g (0.042 mol) of N-hydroxysuc1nimide in 200 ml of ethyl acetate is cooled to 8.6 g (0.042 mol) of dicyclohexylcarbodiimide is added; overnight in the fridge at dicyclohexylurea is filtered, the filtrate is evaporated to a volume of ml in vacuo, colorless crystals are filtered off and 6.4 g of product are obtained, T.Sh1. 121-122, The filtrate is evaporated to dryness in vacuo, the residue is punched with 20 ml of a mixture of benzene-n-hexane and an additional amount of the desired compound is isolated. Total yield 13.4 g (92%); оС J- 1.5 (, chloroform), and VacncLeno,% 8 С 54.85; H 5.18; N 8.00. Found,%: C 54.79; 54.70; H 5.2 5.28; N 8.14; 8.12. And measure 3. Preparation of N- (benzyl oxide to arbonyl x)) succium to them yes. 23 g (0.2 mol) of N-oxysuccinimide are dissolved. In picTBOpe 9 g (0.22 mol of sodium hydroxide in 200 ml of water, 34 g (0.2 mol) of benzoxychloride carbr667142 are added dropwise with stirring and cooling. ay at room temperature overnight. The precipitate is filtered off, washed with silica gel and dried in air. Yield 41 g (82%). After recrystallization of the mixture from benzene-n-hexane (10: 1), colorless prisms are obtained, mp., Example 4. Preparation of 6-carbobenzoxycanamycin A. A solution of 42.5 g (90 mmol) of kanamycin A (in the form of the free base) in 450 ml of water and 500 ml of DMF is cooled to a temperature below 0 ° C and move vigorously, a solution of 2.4 g (90 mmol) of N- (benzyloxycarbonyloxy) -succinimide in 500 ml of DMF is added dropwise in an hour, and stirred at (-10) overnight and at room temperature for days and evaporated in vacuo at a temperature below 5 ° C. The oily residue is dissolved in a mixture of 500 ml of water and 500 ml of butanol, filtered, the butanol and water layer is treated, and water is saturated with butanol (2 x 500 ml) and butanol saturated with water (2 X 500 ml), respectively, using a method similar to countercurrent extraction y. (The three aqueous layers are combined and evaporated to dryness in vacuo. To an oily residue, a part of which crystallizes on standing at room temperature / 1 pO ml of methanol is added, which dissolves the oil and thus separates epr from the crystals. After adding v3OO ml the ethanol mixtures are kept at room temperature overnight and the crystalline mass is filtered off. The product (44 g) contains a small amount of kanamycin A according to thin layer chromatography (TLC) in the n-propanol-pyridine-acetic KieSlta system - time yes (15: 10: 3: 12) when developing by ninhydrin. The crude product is dissolved in 300 ml of water and chromatographed on a column (diameter 30 mm) with CG-50 resin in NHt form (500 ml) with elution with 0.1N solution ammonium hydroxide and collecting fractions of 10 ml each. Fractions 10-110 are combined and evaporated to dryness in vacuo to give 24.6 g (45%) of colorless white arbobenzoxycanamycin A, which begins to Shayytbbya and interrogated at 204s and decomposed at 212 ° C with gas, tl06 (, water). With those on silica gel F 254 and manifestations of ninhydrin in the system n-propanl-pyridine-acetic acid water (10: 10: 3: 12); acetone-acetic acid - water (20: 6: 74); Chloroform-methanol-concentrated ammonium hydroxide solution. - water (1: 4: 2: 1) and methyl acetate-n-propanol-ointhene solution of yelmonium oxide hydrate (45: 105; 60) for 6-car1

5 buoenzoksikanamitsin A and kanamycin A magnitude R | is 0.42 (main), 0.33 (minor) and 0.04; 0.24 and 0.1.4; 0.76 and 0.50 and 0.22 (detection by reaction with anthron-sulfuric acid) and 0.04 (detection by reaction with anthron-sulfuric acid), respectively / The final product contains small amounts of two impurities (according to those in one of these solvent systems). Example 5. Preparation of L - (-) - - amino - with - oxybenzoic acid. 5.0 g of ambutirosine A is boiled for 1 h under reflux with 160 ml / 0.5 N sodium hydroxide solution. The hydrolyzate is neutralized with 6 and hydrochloric acid and chromatographed on a column with CG-50 in the NH.-form% The target acid is eluted with water and dried by freezing. A crystalline product is obtained, mp. 212.5-214 ,, Example 6. Preparation of L - (-) - j4 -amino-oL-hydroxybutyric acid. A solution of 29 g (0.1 kg) of dehydroabiethylamine in 130 ml of ethanol was added to paCTBiopy 25 g (0.1 mol) of 2-hydroxy / - PhtaLimid-HCl acid in 200 ml of ethanol, vigorously with BfiTpSxff for 1 minute and kept for 5 hours at room temperature. Thin needles 6filter filter, washed with 50 ml of this; nola and dried in air. 30.1 g (56%) of dihydroabiethylamine salt are obtained, dihydroabiethylamine, tg. 93-94 C; 15 (, 5, meta; node). .. After recrystallization, 23.2 g (43%) of pure 1, -cC-hydroxy-phthalimidobutyrate dehydroabiethylammonium, m.p. 94-9 +10.8 (, 5, methanol). Further crystallization does not lead to a change in the melting point and o {.e. Calculated,%: C 69.64; H 8.02; N 5.07. C32H42N205-H20 Found: C, 69.58; H 8.08; N 5.07. : To a solution of 1.5 g (0.014 mol) of sodium carbonate. In 40 ml of water was added 5.3 g (0.01 mol) of L-oC-hydroxy-y-ammonium dehydroabiethyl ammonium and 60-ml ether., vigorously mix until dissolved. The ether layer is separated. The aqueous layer was washed with 2 x 20 ml of ether and evaporated to a volume of 15 ml in vacuo, 10 ml of concentrated hydrochloric acid was added and the mixture was heated under reflux for 10 h. After cooling, the phthalic acid was filtered, the filtrate was evaporated in vacuo, the residue dissolved in 10 ml of water and evaporated to dryness. This operation is repeated two times to remove excess hydrochloric acid. The remaining syrup is dissolved in 10 ml of water and filtered to remove a small amount of phthalic acid. The filtrate is passed through a column of IR-120 resin in the H-form (1 cm x X 35 cm). The column was washed with 300 ml of water and eluted with 1N ammonium hydroxide solution, collecting 15 ml fractions. The fractions giving a positive reaction to ninhydrin (10-16) are combined and evaporated in vacuo to give a syrup, which gradually crystallizes. The crystals are triturated with ethanol, filtered and dried in a vacuum desiccator. Obtain 0.78 g (66%) of the desired product, so pl. 206-20700; (оС -29 "(, 5, water). The IR spectrum of the product is identical. Spectrum of a sample obtained from amber, butyrina, 1 Example 7. Preparation of 6-carbobenzoxy-1, 3,3-tri-p-nitrobenzalkanamycin A. 9.0 g (14.5 mmol) of 6-carbobenzoxyanamic A are suspended in 500 ml; absolute e Sholapri and 8.7945 g (58.2 mmol) of p-nitrobenzaldehyde are added, heated to boiling point with a reflux condenser, the observed formation a clear yellow solution, followed by a quick release of a white solid. - The mixture is boiled for 3 hours under reflux, cooled, and KT is filtered off and washed with a small volume of absolute ethanol. The yield of a white crystalline substance is 14.0 g (94.5%), mp 233234 ° C. Calculated,%: C 55.46; H 5.05; N 9.63 C4- / K51CHO e Found,%: C 55.39; H 5.08; N9., 60. Example 8. Preparation of (-) - 3 -amino-eC-Oxybutyryl-canamycin A. 5.0 g ( . mmol) 6-carbobenzoxy-1, 3,3-tri-p-nitrobenzalkanamycin A sweat solution in 50 ml of DMF at 2.064 g (5.895 mmol) of N-oxysuccinimide ester L - (-) - -benzyloxycarbonylamino-oit-- hydroxybutyric acid is dissolved in 20 ml of DMF with and slowly added with vigorous stirring to prepare pacTlBOpy Schiff bases for 75 min, stirred overnight at 24 ° C. In the vacuum created by the steam ejector, the solution is instantly evaporated at, then instantly

evaporated by addition of 100 ml of methanol.

 The irradiated viscous oil is dissolved in 10Q MU mixtures of dirxane-water (Isl), 10 ml of glacial acetic acid is added, placed in a Parr volume of 500 ml, 2.5 g of 5% palladium on carbon (Engelhardt ) and hydrogenated for 4 hours at a hydrogen pressure of 3105 N / m and a temperature. The total pressure drop over this period of time (with the bottle closed) is 6003 N / m2, including a periodic increase in pressure.

The mixture is filtered through a layer of diatomaceous earth, washed with 3 x 50 ml of 50% aqueous dioxane.

The combined filtrates are instantly evaporated to transfer the product to a solid state and are subjected to azerotropic distillation with 100 ml of n-buta-Nol, and then immediately evaporated with methanol to obtain a viscous oil, which is dissolved in 30 ml of methanol and slowly, but is drunk in 1000 ml of ether () ether with vigorous stirring ...--.

After stirring the resulting suspension for 0.5 hours in ice buy, the precipitate is filtered off and immediately dried in a vacuum cleaner over phosphorus.

4.9620 g of product are obtained, which, according to the data of those, consists in osnova of cellei compound, kanamycin A, 4-amino-2-hydroxybutyric acid, and dosages of di- and tri-substituted kanamycin A.

A glass sponge 100 i with amberlite 1RC-40 in NHf-form {type 1, particle size 100/200 mesh, height of resin layer 960 mm) is charged with 4,600 g of product dissolved in 15 ml of water, eluting with a gradient method first with water, then 2 N, hydroxide solution. Collect 295 fractions. Wash the column with 1.5 L of 3N ammonium hydroxide solution. Fractions homogeneous in size with p are pooled and lyophilized.

M.p. (decomp.) {€ Le +85 (, WEDA).

Calculated,%: C 40.62; H 6.68f N 9.87.

С2гН4з% о з2НгСО

Found,% 1 C 40.21 39.79; H 6.96; 6, 9.37; 9.49.

Example 9. Obtaining monosulfate 1- L- (-) - -acmo-oC-hydroxybutyryl-KanMitsina A.

1 mol of (-) - f -amino-c-hydroxybutyryl-kanamycin A is dissolved in 1-3 l of water, filtered, cooled with stirring, 1 mol of sulfuric acid dissolved in 500 ml of water is added, stirred for 30 minutes, added cold ethanol Prior to the deposition of crystals that are filtered out ..

Example 10. Obtaining disulfate (-) - -amic-oC-hydroxybutyryl) -canamycin A.

35 g of (-) --j - amino-oC-hydroxybutyryl) -canamycin A (in the form of monobicarbonate trihydrate) are dissolved

in 125 ml of deionized water (pH "9.0). The pH is adjusted to 7-7.5 by the addition of 50% (volume / volume) sulfuric acid.

Add 8.5 grams of activated

charcoal and stirred for 0.5 h at, the coal is filtered off and washed with 40 ml of water. The washings are added to the filtrate, acidified to a pH of 2-2.6 with 50% (volume / volume) sulfuric acid, a large amount of carbon dioxide is observed. The solution is degassed under vacuum for 20 minutes, added 8.5 g of charcoal, stirred for 0.5 h at, the carbon is filtered off and washed with 35 ml of deionized water. Wash water is added to the filtrate, acidified to a pH of 1-1.3 with the aid of 50% (volume / volume) sulfuric acid, added with rapid stirring in 10 minutes to 600-800 ml of methanol (3-4 volumes of methanol ), stirred for 5 minutes at a pH of 1-1.3, passed through a sieve (100 mesh), stirred for 2 minutes and the solid was allowed to settle for 5 minutes. Most of the upper liquid layer is decanted. The remaining sludge is filtered off, proElvigot 200 ml of methanol and

5 is dried under vacuum for 24 h. The yield of g1 morphic disulfate is 32-34 g for monohydrate ot J2j) +74.75. The product decomposes at 22 p-223 ° C.

Q Elemental analysis is given for dry matter.

Found,%: C 32.7; ; 33.52 32.3; N 8.78; 8.7; 8.2; 8.8; S 8.75; 8.9; 7.8; 8.85,

Calculated; C 33.5; N8.97;

S 8,2.

Claims (3)

1. The method of obtaining (-) - -amino-L-hydroxybutyryl) -canamycin A of the formula CH -CHN-JANs are distinguished by the fact that S benenloxycarbonylcanamycin A is reacted with at least a 3-fold molar excess of aldehyde of the general formula R-CHO, where R is phenyl, p-nitrophenyl, o-oxyphenyl, p-methoxyphenyl or tert-butyl, in an environment of a lower alcohol at a temperature from to the boiling point of the solvent, forming the base Miff of the general formula H, "HjUHdOOCKj leHs HO-v tojOtt but fl:; has: it means that the abovementioned ones are acylated with L-hydroxy: 210 L - (-) - -benzyloxycarbonylamino-ot-hydroxybutyric acid, followed by hydrogenation of the reaction mixture. 2. A process according to claim 1, characterized in that N-benzyloxycarbonylcanamycin A is reacted with 3.5-4.5 mol r: m amount of p-nitrobenzaldehyde, salicylic aldehyde or benzaldehyde in absolute methanol, ethanol, n-propanol or isopropanol at the boiling point of the reaction mixture 3. A process according to, characterized in that the acidification of the Schiff base is carried out with a 1.0-1.3 molar amount of the N-hydroxysuccinimide ester of L - (-) -f-benzyloxycarbonylamino-oL-hydroxybutyric acid in dimethylformamide media. Sources of information taken in. attention during examination 1. McOmi JS. Protective groups, Sat. Successes of organic chemistry, M., world, vol. 3, p. 203, 1966.