FI56689C - PROCEDURE FOR THE FRAMEWORK OF ETHIN AMINOCYCLITOLANTIBIOTICS - Google Patents

Description

iry ^ .i Γβ1 advertisement publication ^ (11) UTLÄGGN I NGSSKRI FT 66689 C Patent granted 10 03 1930

Patent Meddelat ^ v ^ (51) K ».lk. * / Lnt.CI. * c 07 H 15/24 C 12 D 9/14 SUOMI — FINLAND (21) '« • "« n »·» "" » -p «« tt * n ^ iuiini 780198 (22) Haktmitpliva - Antäknlnpdif 23.01.78 ^ ^ (23) AlkupUvi — Glltlfh «tad« (17.02.76 (41) Tullut | external «k * l - Bllvlt offMitltf 23.01.78

National Board of Patents and Registration .... ...., ..., * (44) Nihtlväkilpanon μ kuLLPublished date. -

Patent and Administrative Cases Ansöktn utligd och utl. »Krlft« n published 30.11.79 (32) (33) (31) Privilege claimed — B «| ird prlorttet iQ.Q2.75 22.09.75 USA (US) 550273, 615593 ( 71) Sterling Drug Inc., 90 Park Avenue, New York, NY, USA (72) Soi Jacob Daum, Albany, New York, Robert La Grone Clarke, Delmar, New York, USA (7 * 0 Oy Kolster Ab (5 ^) Method for preparing an aminocyclitol antibiotic - Förfarande för framställning av ett aminocyclitolantibiotikum (62) Separated from application 760387 (patent 56026) 1.6689 CEL · I3

tA

r8-n (N \ ^

0 NH-R_ I

O / N-NH-R1

h ^ ~~ A

CH0NH I- 3 Jh wherein one of R1, R1 and R8 is a N-amino - N - hydroxy-lower alkanoyl group of the formula H2NCH2 (CH2) nCHOHCO- wherein n is 0 or 1, wherein the other groups R R 1, R 2 and R 8 are hydrogen atoms, or to prepare an acid addition salt thereof.

The process according to the invention is characterized in that the corresponding compound of the formula I in which R 1, R 8 and R 8 are hydrogen atoms is reacted with an N-hydroxysuccinimide ester of the formula 566e90.

IT

$ CH O-C-NHCH, (CH,) CHOHC-O-N \ IV

\ = / 2 ^ \ J

II

0 wherein n is as defined above, and the benzyloxycarbonyl group of the product formed is hydrogenated with hydrogen in the presence of a catalyst, and, if desired, the resulting free base is converted into its acid addition salt.

The starting compounds of formula I in which and R 9 are hydrogen atoms are prepared analogously to the method described in U.S. Patent No. 3,669,838 to Shier et al., As described in Finnish Patent Application No. 760387 (Publication No. 56,026). The improved method is described in Finnish Announcement 56,027 (separated by application 760387).

The reaction according to the invention gives a mixture of three possible isomeric monoamides in which one of the Rg or Rg-aminohydrogen atoms has been replaced by a w- (N-benzyloxycarbonyl) -amino-ed-hydroxy-lower alkanoyl group.

If these products are to be identified and studied as individuals, they must first be isolated from each other. The acylation reaction is carried out by reacting equimolar amounts of a compound of formula I and the N-hydroxysuccinimide ester at a temperature of -10 ° to about + 10 °, in the presence of water and an inert organic solvent, e.g.

Cleavage of the benzyloxycarbonyl group is performed in the presence of a palladium-on-carbon catalyst and an inert, water-miscible organic solvent, e.g. methanol, ethanol, dioxane, tetrahydrofuran, ethylene glycol dimethyl ether or propylene glycol dimethyl ether.

The N-hydroxysuccinimide ester of formula IV is well known.

The compounds of formula I have been tested in a standard serial dilution test for antibacterial activity and have been found to have antibacterial activity, in particular against gentamicin-resistant organisms. Thus, these compounds are useful antibacterial agents.

Preferably, the compounds of formula I are intended for oral, topical and parenteral administration and may be formulated as suspensions with a pharmaceutical carrier either as free bases or as pharmaceutically acceptable acid addition salts in an inert carrier, e.g. polyethylene glycol or as or for topical administration in the form of conventional creams or jellies.

The molecular structure of the compounds prepared according to the invention was determined in the same manner as in parent patent application 760387 (Publication No. 56,026).

Results of the antibacterial test:

Stock solutions containing 200 .mu.g / ml of each compound in distilled water were prepared and the filtration was sterilized. Cultures of test organisms were grown for 2 hours at 37 ° C in 10 ml tubes with tryptose phosphate or Miiller-Hinton medium. Each culture was adjusted to an optical density of 0.1 g in nutrient medium with 0.1 Spectronic 20 (approximately 10 cells / ml). Regulated cultures were diluted 1: 500 in nutrient medium for use as an inoculum (final cell concentration approximately 2 x 10 6 cells / ml). The antibacterial activity of the test compounds was studied by a single-row tube dilution method. Two-fold serial dilutions were made in drug stock medium and 0.2 ml of each drug concentration was added to 17 tubes 13 x 100 mm. 0.2 ml of appropriately diluted culture was transferred to all tubes (final cell concentration of tubes 10 μl / ml). Minimum inhibitory concentrations (lowest drug concentration without visible growth) were determined after 16 hours of incubation at 37 ° C.

The components of component 1 (0-3-deoxy-UC-methyl-3-methylamino-β-L-arabinopyranosyl- (1-> 6) -O- [2-amino-6-methylamino-6-C-methyl ”described in the example) are described. -SiS, β-tetradeoxy-ε-LD-erythroglucopyranosyl-C 1-7 +) 7-D-streptamine, 1-, 3- and 2 "- (S) -β-amino-β-hydroxybutyryl-7-amides ( labeled as following comp.1 (1-HABA) Comp. 1 (3-HABA) and comp. 1 (2 "-HABA) were tested compared to the corresponding 1-, 3- and 2" - / S (-) - - - amino - (/ - hydroxybutyryl amides (all described by Konishi et al., U.S. Patent No. 3,780,018; - denoted below by (1-HABA), C1 (3-HABA) and C1 (2 "-ΗΑΒΑ).

The results are shown in Table I below.

Table I _

Compound Experimental Georgejism B. subtilis S. aureus E. coli Ent.cloa- K.pneu- Ps.aerugi-ATCC 6633 Smith JR 76.2 cae many nosa A-20960 A-20636 A-20897 C1 (1-HABA) 0.39 3.13 12.5 3.13 6.25> 100

Comp. 1 (1-HABA) 0.78 6.25 12.5 6.25 12.5> 100 C1 (3-HABA) 3.13 25> 100 100> 100> 100

Comp. 1 (3-HABA) 6.25 50 100 50 50> 100 C1 (2 "-HABA) 6.25 50 100 25 50> 100

Comp. 1 (2 ^ -HABA) 1.56 25 50 25 50> 100

Cultivated in Miiller-Hinton nutrient medium.

5,0689

The following example illustrates the method of the invention.

Biosynthesis of starting material by M. purpurea ATCC 31 119 Starting material, O-3-deoxy-4-C-methyl-3-methylamino - [* - L-arabinopyranosyl- (1-> 6) -O- [2-amino-6- methylamino-6-C-methyl-2,3,4,6-tetadexoxy-β-β-erythro-glucopyranosyl- (1-> 4) -D-streptamine was prepared as described in Example 1 of Application 760387.

Synthesis of aminohydroxy-lower alkanyl derivatives

A solution of 270 mg (0.54) mmol of O-3-deoxy-4-C-methyl-3-methylamino-N-L-arabinopyranosyl- (1- * 6) -O- [2-amino-6 -methylamino-6-C-methyl-2,3,4,6-tetradeoxy-N-D-erythroglucopyranosyl- (1-4) 7-D-streptamine dissolved in 5 ml of an aqueous solution of 50- $ tetrahydrofuran, cooled to 5 ° Overnight in an ice bath and treated with 208 mg (0.59 mmol) of Sti-γ-N-benzyloxycarbonyl) -amino-N-hydroxybutyric acid N-hydroxysuccinimide ester (Konishi et al., U.S. Patent No. 3,780,018). and the mixture was stirred for 20 hours at 5 ° C.

The mixture was then concentrated in vacuo to 10 mL, 25 mL of n-butanol and 10 mL of water were added, and the layers were separated. The aqueous layer was washed with 10 ml of n-butanol. The combined organic phases were evaporated to dryness to give 513 mg of crude product which was set aside.

The aqueous phase was evaporated to dryness to give 304 mg of a residue which was dissolved in 25 ml of a 50- [. An additional 435 mg of crude product was obtained from the same reaction mixture, which was combined with 513 mg previously obtained and chromatographed on seven 40 x 20 cm silica gel plates, 1 mm thick, The system was developed with chloroform: methanol: 28- $ ammonium hydroxide (3: 1: 1 lower phase). In this solvent system, seven runs were required, and when the product zone visible under UV light was eluted, 229.5 mg of an impure mixture of three monoacylated products was obtained.

The mixture of acylated products was placed on three 40 x 20 cm silica gel plates, 1 mm thick, and the plates were developed five times with chloroform: isopropanol: concentrated (28- $) ammonium hydroxide (3: 1: 1, lower phase) and nine times with chloroform: methanol: concentrated (28- $) with ammonium hydroxide (h: 1, lower phase). Three clear bands visible under UV light were obtained. The zones were sectioned apart, eluted from silica gel with chloroform-methanol: 28- $ ammonium hydroxide (1: 1: 1, lower phase) to give three components: A 90.9 mg, B 59.1 mg and C 48.5 mg, which are 0-3 -deoxy-4-C-methyl-3-methylamino-N-L-arabinopyranosyl- (1-> 6) -O- [2-amino-6-methylamino-6-C-methyl-2,3 , 4,6-Tetradeoxy-N-D-erythroglucopyranosyl- (1-> U) -7-D-streptamine S - (-) - Y- (N-benzyloxycarbonyl) -amino-N-hydroxybutyramide derivatives - and $ 6,668 in 3rd position. The R 1 values of components A, B and C on silica gel with the chloroform: methanol: 28- # ammonium hydroxide system (U; 1: 1, lower phase) were: A = 0, U 8, B = 0.62 and C = 0.70.

Component B (1-amide, 56.1 mg) was dissolved in 25 ml of a 50-JC aqueous ethanol solution, and 20 mg of 10-palladium-carbon catalyst was added, and shaken in a Parr pcan reactor at a pressure of 3.85 kp / cm 5 1 / 2 hours, and then the catalyst was removed by filtration in the presence of a filter aid. Evaporation of the solvent gave 3 * +, 3 mg of a white glassy substance which was dissolved in 2.5 ml of water and treated with 11.0 mg of sulfuric acid in 0.1 ml of water. Upon addition of 10 ml of ethanol, 32 mg of 1- (S - (-) - N-amino-o-4-hydroxy-butyryl) -O-3-deoxy-1 + -C-methyl-3-methylamino- [5-L- arabinopyranosyl- (1- * 6) - [2-amino-6-methylamino-6-C-methyl-2,3,3 *, 6-tetradeoxy-N-D-erythroglucopyranosyl- (1- > U) -D-streptamine as pentasulfate salt; mp. 230-235 ° C (decomposed), thin layer chromatographic analysis: R f = 0.18 (silica gel, chloroform: methanol: concentrated (28- #) ammonium hydroxide: water (1: U: 2: 1), gentamicin C used as reference Rf = 0.73.

Analysis: C ^ H ^ O ^ Ng.5 HgSO4

Calculated: C 27.67 H 5.57 N 7.75

Found: C 27.50 H 5.58 H 7, k2

Components A and C were treated in the same manner. 1 + 7 mg of 2 "- [- (-) -] -? -? -? - hydroxybutyryl-O-3-deoxy-UC-methyl-3-methylamino- [2-L-arabinopyranosyl- (1- (6) -O- [2-amino-6-methylamino-6-C-methyl-2,3,3 *, 6-tetradeoxy- [D-erythroglucopyranosyl- (1-> U) 7-D- streptamine as dibasic tetrasulfate heptahydrate, m.p. 1), the reference substance gentamicin R 2 = 0.73 · -

Analysis: (C 2 H 50 N 6 O 10 ^ 2 * ^ H 2 SO 2 T 2 O 2

Calculated: C 35.16 H 7.20 N 9.8 * +

Found: C 35.38 H 7.08 N 9, * + 9

From component C, 26 mg of 3- (S - (-) -) -? - emino - t-hydroxybutyryl-O-3-deoxy-LC-methyl-3-methylamino- [®-L-arabinopyranosyl- (1 -> - 6) -O-2-amino-6-methylamino-6-C-methyl-2,3> * +> 6-tetradoxy-^ -De ry t roglucopyran o-silyl- ( 1-> U) -D-streptamine as dibasic pentasulfate trihydrate; mp. 220-230 ° C (decomposed), thin layer chromatographic analysis Rf = 0.30 (silica gel, chloroform'.methanol: concentrated (28- #) ammonium hydroxide: water (1: U: 2: 1), gentamycin C n Rf * 0.73.

Analysis: (C25H50N6 ° 10 ^ 2 * 5 H2SO3 '3 H2 °

Calculated: C 3 * + »36 H 6.7 * + N 9.76

Found: C 3 * +, 35 H 6.38 N 8.60

Claims (1)

A process for the preparation of an aminocyclitol antibiotic of the formula ch3 CH-NH-CH0 '^ Rq-NH xq NH-R3 - OH H0 ~~ ^ -7 \\ 0 ^ -HH-R CH3NH - ^ 3 OH wherein one of R 1, R 3 and R 8 is a u> -amino - / = - hydroxy-lower alkanoyl group of the formula HNNCHO (CH) CHOHCO-d ddn wherein n is 0 or 1, wherein the other groups R 1, R 8 and Rg are hydrogen atoms. or an acid addition salt thereof, characterized in that the corresponding compound of formula I in which R 1, R 9 and R 8 are hydrogen atoms is reacted with an N-hydroxysuccinimide ester of formula 0 / "Λ SS / l 1 V> CH 0 -C- NHCHo (CHO) CHOHC-ON IV W / 2 2 2 n \ _ ti 0 where n is as defined above, and the benzyloxycarbonyl group of the product formed is cleaved by hydrogenation with hydrogen in the presence of a catalyst and, if desired, the free base obtained is converted into its acid addition salt. framställning av ett aminocycitolantibiotikum med formeIn CH3 CH-NH-CH- - R8 1111 NH-R / \ nZ - ^ - OH HO 1 - ^ -— rs. \ Q N-NH-R1 \ ^ 0 - \ ΗΟ-Λ --- \ / ^ A ^ CH3 CH3NH ηρ -3 OH w color viscondera of R1, Rg and Rg betecnar en uu-amino-ot-hydroxy-lasgre-alkanoyl group with the formula HgNCHg (CH2) nCHOHCO- vari n är 0 eller 1 , varvid de andra av R ^, Rg och Rg är väte, eller ett syraadditionssalt därav, kännete cknat därav, att man omsätter en motsvarande Fören with the formula I »for R ^, Rg and Rg with the atomic atom, with the N-hydroxysuccinimide ester with the formula