CS261853B2 - Method of aminoglycoside antibiotic's selectively acylated n-protected derivative production - Google Patents
Method of aminoglycoside antibiotic's selectively acylated n-protected derivative production Download PDFInfo
- Publication number
- CS261853B2 CS261853B2 CS797711A CS771179A CS261853B2 CS 261853 B2 CS261853 B2 CS 261853B2 CS 797711 A CS797711 A CS 797711A CS 771179 A CS771179 A CS 771179A CS 261853 B2 CS261853 B2 CS 261853B2
- Authority
- CS
- Czechoslovakia
- Prior art keywords
- amino
- hydrogen
- formula
- zinc
- hydroxy
- Prior art date
Links
- 239000002647 aminoglycoside antibiotic agent Substances 0.000 title claims abstract description 112
- 229940126574 aminoglycoside antibiotic Drugs 0.000 title claims abstract description 94
- 238000000034 method Methods 0.000 title claims description 112
- 238000004519 manufacturing process Methods 0.000 title claims description 8
- 239000011701 zinc Substances 0.000 claims abstract description 97
- 229910052725 zinc Inorganic materials 0.000 claims abstract description 97
- -1 zinc cations Chemical class 0.000 claims abstract description 91
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 claims abstract description 72
- 229960000318 kanamycin Drugs 0.000 claims abstract description 69
- 150000001875 compounds Chemical class 0.000 claims abstract description 64
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims abstract description 62
- 125000003277 amino group Chemical group 0.000 claims abstract description 57
- 229930182823 kanamycin A Natural products 0.000 claims abstract description 57
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 claims abstract description 51
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 claims abstract description 43
- 125000002252 acyl group Chemical group 0.000 claims abstract description 28
- 230000003115 biocidal effect Effects 0.000 claims abstract description 28
- 150000001768 cations Chemical class 0.000 claims abstract description 16
- URWAJWIAIPFPJE-UHFFFAOYSA-N Rickamicin Natural products O1CC(O)(C)C(NC)C(O)C1OC1C(O)C(OC2C(CC=C(CN)O2)N)C(N)CC1N URWAJWIAIPFPJE-UHFFFAOYSA-N 0.000 claims abstract description 7
- URWAJWIAIPFPJE-YFMIWBNJSA-N sisomycin Chemical compound O1C[C@@](O)(C)[C@H](NC)[C@@H](O)[C@H]1O[C@@H]1[C@@H](O)[C@H](O[C@@H]2[C@@H](CC=C(CN)O2)N)[C@@H](N)C[C@H]1N URWAJWIAIPFPJE-YFMIWBNJSA-N 0.000 claims abstract description 7
- 229930182566 Gentamicin Natural products 0.000 claims abstract description 6
- 229930192786 Sisomicin Natural products 0.000 claims abstract description 6
- 229960005456 sisomicin Drugs 0.000 claims abstract description 6
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 claims abstract 3
- 229960002518 gentamicin Drugs 0.000 claims abstract 3
- 229910052739 hydrogen Inorganic materials 0.000 claims description 90
- 239000000203 mixture Substances 0.000 claims description 80
- 239000001257 hydrogen Substances 0.000 claims description 74
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 claims description 73
- 150000003751 zinc Chemical class 0.000 claims description 61
- PTFCDOFLOPIGGS-UHFFFAOYSA-N Zinc dication Chemical compound [Zn+2] PTFCDOFLOPIGGS-UHFFFAOYSA-N 0.000 claims description 60
- 238000005917 acylation reaction Methods 0.000 claims description 56
- 230000008569 process Effects 0.000 claims description 55
- 229940006486 zinc cation Drugs 0.000 claims description 55
- 229940126575 aminoglycoside Drugs 0.000 claims description 53
- 150000002431 hydrogen Chemical group 0.000 claims description 47
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 42
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 42
- 239000003795 chemical substances by application Substances 0.000 claims description 41
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 claims description 40
- 239000003960 organic solvent Substances 0.000 claims description 38
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 36
- 239000002253 acid Substances 0.000 claims description 27
- 239000011541 reaction mixture Substances 0.000 claims description 22
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 21
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 20
- 125000004435 hydrogen atom Chemical group [H]* 0.000 claims description 20
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 claims description 20
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 19
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 claims description 18
- 239000011347 resin Substances 0.000 claims description 18
- 229920005989 resin Polymers 0.000 claims description 18
- 239000003495 polar organic solvent Substances 0.000 claims description 17
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 claims description 16
- 125000002091 cationic group Chemical group 0.000 claims description 15
- 229960001192 bekanamycin Drugs 0.000 claims description 14
- 229910052751 metal Inorganic materials 0.000 claims description 14
- 239000002184 metal Substances 0.000 claims description 14
- 239000002904 solvent Substances 0.000 claims description 14
- 125000004432 carbon atom Chemical group C* 0.000 claims description 13
- 150000002148 esters Chemical class 0.000 claims description 13
- 239000003729 cation exchange resin Substances 0.000 claims description 12
- 125000000250 methylamino group Chemical group [H]N(*)C([H])([H])[H] 0.000 claims description 12
- SKKLOUVUUNMCJE-FQSMHNGLSA-N kanamycin B Chemical compound N[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SKKLOUVUUNMCJE-FQSMHNGLSA-N 0.000 claims description 10
- 229930182824 kanamycin B Natural products 0.000 claims description 10
- 229930182825 Kanamycin C Natural products 0.000 claims description 9
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 9
- 125000000217 alkyl group Chemical group 0.000 claims description 9
- WZDRWYJKESFZMB-FQSMHNGLSA-N kanamycin C Chemical compound O([C@H]1[C@H](N)C[C@@H]([C@H]([C@@H]1O)O[C@@H]1[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O1)N)N)[C@H]1O[C@H](CO)[C@@H](O)[C@H](N)[C@H]1O WZDRWYJKESFZMB-FQSMHNGLSA-N 0.000 claims description 9
- 150000007522 mineralic acids Chemical class 0.000 claims description 9
- 150000007524 organic acids Chemical class 0.000 claims description 9
- RWSOTUBLDIXVET-UHFFFAOYSA-N Dihydrogen sulfide Chemical compound S RWSOTUBLDIXVET-UHFFFAOYSA-N 0.000 claims description 8
- 230000006870 function Effects 0.000 claims description 8
- 125000000524 functional group Chemical group 0.000 claims description 8
- 229910000037 hydrogen sulfide Inorganic materials 0.000 claims description 8
- 150000003839 salts Chemical class 0.000 claims description 8
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 claims description 7
- 229920001661 Chitosan Polymers 0.000 claims description 7
- 239000002585 base Substances 0.000 claims description 7
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 claims description 7
- 150000001732 carboxylic acid derivatives Chemical class 0.000 claims description 7
- 229920000642 polymer Polymers 0.000 claims description 7
- QUSNBJAOOMFDIB-UHFFFAOYSA-N Ethylamine Chemical compound CCN QUSNBJAOOMFDIB-UHFFFAOYSA-N 0.000 claims description 6
- 239000005083 Zinc sulfide Substances 0.000 claims description 6
- 229910052977 alkali metal sulfide Inorganic materials 0.000 claims description 6
- 239000003957 anion exchange resin Substances 0.000 claims description 6
- 239000003153 chemical reaction reagent Substances 0.000 claims description 6
- 238000000605 extraction Methods 0.000 claims description 6
- ZBGPYVZLYBDXKO-HILBYHGXSA-N netilmycin Chemical compound O([C@@H]1[C@@H](N)C[C@H]([C@@H]([C@H]1O)O[C@@H]1[C@]([C@H](NC)[C@@H](O)CO1)(C)O)NCC)[C@H]1OC(CN)=CC[C@H]1N ZBGPYVZLYBDXKO-HILBYHGXSA-N 0.000 claims description 6
- DRDVZXDWVBGGMH-UHFFFAOYSA-N zinc;sulfide Chemical compound [S-2].[Zn+2] DRDVZXDWVBGGMH-UHFFFAOYSA-N 0.000 claims description 6
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 claims description 5
- ZOIORXHNWRGPMV-UHFFFAOYSA-N acetic acid;zinc Chemical compound [Zn].CC(O)=O.CC(O)=O ZOIORXHNWRGPMV-UHFFFAOYSA-N 0.000 claims description 5
- 239000003085 diluting agent Substances 0.000 claims description 5
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 claims description 5
- 229960000808 netilmicin Drugs 0.000 claims description 5
- 239000001632 sodium acetate Substances 0.000 claims description 5
- 235000017281 sodium acetate Nutrition 0.000 claims description 5
- 239000004246 zinc acetate Substances 0.000 claims description 5
- 229910052984 zinc sulfide Inorganic materials 0.000 claims description 5
- XUSXOPRDIDWMFO-CTMSJIKGSA-N (2r,3r,4r,5r)-2-[(1s,2s,3r,4s,6r)-4,6-diamino-3-[[(2s,3r)-3-amino-6-[(1s)-1-aminoethyl]-3,4-dihydro-2h-pyran-2-yl]oxy]-2-hydroxycyclohexyl]oxy-5-methyl-4-(methylamino)oxane-3,5-diol Chemical compound O1C[C@@](O)(C)[C@H](NC)[C@@H](O)[C@H]1O[C@@H]1[C@@H](O)[C@H](O[C@@H]2[C@@H](CC=C(O2)[C@H](C)N)N)[C@@H](N)C[C@H]1N XUSXOPRDIDWMFO-CTMSJIKGSA-N 0.000 claims description 4
- 229920002101 Chitin Polymers 0.000 claims description 4
- 125000003545 alkoxy group Chemical group 0.000 claims description 4
- 125000004453 alkoxycarbonyl group Chemical group 0.000 claims description 4
- 150000008064 anhydrides Chemical class 0.000 claims description 4
- 239000002738 chelating agent Substances 0.000 claims description 4
- 235000011194 food seasoning agent Nutrition 0.000 claims description 4
- 150000004820 halides Chemical class 0.000 claims description 4
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 claims description 4
- 229920003176 water-insoluble polymer Polymers 0.000 claims description 4
- XUSXOPRDIDWMFO-UHFFFAOYSA-N Verdamicin Natural products O1CC(O)(C)C(NC)C(O)C1OC1C(O)C(OC2C(CC=C(O2)C(C)N)N)C(N)CC1N XUSXOPRDIDWMFO-UHFFFAOYSA-N 0.000 claims description 3
- 229910052784 alkaline earth metal Inorganic materials 0.000 claims description 3
- 239000000908 ammonium hydroxide Substances 0.000 claims description 3
- 238000001704 evaporation Methods 0.000 claims description 3
- 125000004390 alkyl sulfonyl group Chemical group 0.000 claims description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-O ammonium group Chemical group [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 claims description 2
- 125000005140 aralkylsulfonyl group Chemical group 0.000 claims description 2
- 125000003435 aroyl group Chemical group 0.000 claims description 2
- 125000004391 aryl sulfonyl group Chemical group 0.000 claims description 2
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 claims description 2
- 239000003054 catalyst Substances 0.000 claims description 2
- 238000007865 diluting Methods 0.000 claims description 2
- 125000002485 formyl group Chemical group [H]C(*)=O 0.000 claims description 2
- 125000003884 phenylalkyl group Chemical group 0.000 claims description 2
- 238000001179 sorption measurement Methods 0.000 claims description 2
- 125000000542 sulfonic acid group Chemical group 0.000 claims description 2
- UGZADUVQMDAIAO-UHFFFAOYSA-L zinc hydroxide Chemical compound [OH-].[OH-].[Zn+2] UGZADUVQMDAIAO-UHFFFAOYSA-L 0.000 claims description 2
- 229910021511 zinc hydroxide Inorganic materials 0.000 claims description 2
- 229940007718 zinc hydroxide Drugs 0.000 claims description 2
- LKKVGKXCMYHKSL-LLZRLKDCSA-N gentamycin A Chemical compound O[C@H]1[C@H](NC)[C@@H](O)CO[C@H]1O[C@@H]1[C@@H](O)[C@H](O[C@@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)N)[C@@H](N)C[C@H]1N LKKVGKXCMYHKSL-LLZRLKDCSA-N 0.000 claims 4
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims 2
- RHRAMPXHWHSKQB-GGEUKFTFSA-N betamicin Chemical compound O1C[C@@](O)(C)[C@H](NC)[C@@H](O)[C@H]1O[C@@H]1[C@@H](O)[C@H](O[C@@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CN)O2)O)[C@@H](N)C[C@H]1N RHRAMPXHWHSKQB-GGEUKFTFSA-N 0.000 claims 2
- RHRAMPXHWHSKQB-UHFFFAOYSA-N gentamicin B Natural products O1CC(O)(C)C(NC)C(O)C1OC1C(O)C(OC2C(C(O)C(O)C(CN)O2)O)C(N)CC1N RHRAMPXHWHSKQB-UHFFFAOYSA-N 0.000 claims 2
- VEGXETMJINRLTH-BOZYPMBZSA-N gentamycin C1a Chemical compound O1C[C@@](O)(C)[C@H](NC)[C@@H](O)[C@H]1O[C@@H]1[C@@H](O)[C@H](O[C@@H]2[C@@H](CC[C@@H](CN)O2)N)[C@@H](N)C[C@H]1N VEGXETMJINRLTH-BOZYPMBZSA-N 0.000 claims 2
- 125000004178 (C1-C4) alkyl group Chemical group 0.000 claims 1
- OTMSDBZUPAUEDD-UHFFFAOYSA-N Ethane Chemical compound CC OTMSDBZUPAUEDD-UHFFFAOYSA-N 0.000 claims 1
- 125000005098 aryl alkoxy carbonyl group Chemical group 0.000 claims 1
- 229940023913 cation exchange resins Drugs 0.000 claims 1
- CCAFPWNGIUBUSD-UHFFFAOYSA-N diethyl sulfoxide Chemical compound CCS(=O)CC CCAFPWNGIUBUSD-UHFFFAOYSA-N 0.000 claims 1
- 229920005588 metal-containing polymer Polymers 0.000 claims 1
- 210000000056 organ Anatomy 0.000 claims 1
- 238000007363 ring formation reaction Methods 0.000 claims 1
- SQQWBSBBCSFQGC-JLHYYAGUSA-N ubiquinone-2 Chemical compound COC1=C(OC)C(=O)C(C\C=C(/C)CCC=C(C)C)=C(C)C1=O SQQWBSBBCSFQGC-JLHYYAGUSA-N 0.000 claims 1
- 239000000126 substance Substances 0.000 abstract description 23
- 229930027917 kanamycin Natural products 0.000 abstract description 18
- 239000003242 anti bacterial agent Substances 0.000 abstract description 16
- 229940088710 antibiotic agent Drugs 0.000 abstract description 15
- 229910052759 nickel Inorganic materials 0.000 abstract description 9
- DTFAJAKTSMLKAT-JDCCYXBGSA-N 2-deoxystreptamine Chemical compound N[C@H]1C[C@@H](N)[C@H](O)[C@@H](O)[C@@H]1O DTFAJAKTSMLKAT-JDCCYXBGSA-N 0.000 abstract description 8
- 125000003282 alkyl amino group Chemical group 0.000 abstract description 8
- 229910052802 copper Inorganic materials 0.000 abstract description 8
- 229960004821 amikacin Drugs 0.000 abstract description 2
- LKCWBDHBTVXHDL-RMDFUYIESA-N amikacin Chemical compound O([C@@H]1[C@@H](N)C[C@H]([C@@H]([C@H]1O)O[C@@H]1[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O1)O)NC(=O)[C@@H](O)CCN)[C@H]1O[C@H](CN)[C@@H](O)[C@H](O)[C@H]1O LKCWBDHBTVXHDL-RMDFUYIESA-N 0.000 abstract description 2
- 239000000243 solution Substances 0.000 description 64
- 239000007787 solid Substances 0.000 description 50
- 238000002360 preparation method Methods 0.000 description 35
- 229960001760 dimethyl sulfoxide Drugs 0.000 description 32
- RNHDAKUGFHSZEV-UHFFFAOYSA-N 1,4-dioxane;hydrate Chemical compound O.C1COCCO1 RNHDAKUGFHSZEV-UHFFFAOYSA-N 0.000 description 30
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 30
- 239000000047 product Substances 0.000 description 30
- 238000006243 chemical reaction Methods 0.000 description 28
- 239000012458 free base Substances 0.000 description 26
- 230000010933 acylation Effects 0.000 description 20
- 229910052799 carbon Inorganic materials 0.000 description 20
- 229910052757 nitrogen Inorganic materials 0.000 description 20
- WHQSYGRFZMUQGQ-UHFFFAOYSA-N n,n-dimethylformamide;hydrate Chemical compound O.CN(C)C=O WHQSYGRFZMUQGQ-UHFFFAOYSA-N 0.000 description 18
- PXHVJJICTQNCMI-UHFFFAOYSA-N Nickel Chemical compound [Ni] PXHVJJICTQNCMI-UHFFFAOYSA-N 0.000 description 16
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 14
- MJSHDCCLFGOEIK-UHFFFAOYSA-N benzyl (2,5-dioxopyrrolidin-1-yl) carbonate Chemical compound O=C1CCC(=O)N1OC(=O)OCC1=CC=CC=C1 MJSHDCCLFGOEIK-UHFFFAOYSA-N 0.000 description 13
- 125000006239 protecting group Chemical group 0.000 description 13
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- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 description 10
- 229910052723 transition metal Inorganic materials 0.000 description 10
- YZYKBQUWMPUVEN-UHFFFAOYSA-N zafuleptine Chemical compound OC(=O)CCCCCC(C(C)C)NCC1=CC=C(F)C=C1 YZYKBQUWMPUVEN-UHFFFAOYSA-N 0.000 description 10
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 9
- 238000005349 anion exchange Methods 0.000 description 9
- 238000005755 formation reaction Methods 0.000 description 9
- BMYNFMYTOJXKLE-UHFFFAOYSA-N 3-azaniumyl-2-hydroxypropanoate Chemical compound NCC(O)C(O)=O BMYNFMYTOJXKLE-UHFFFAOYSA-N 0.000 description 8
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- 125000001584 benzyloxycarbonyl group Chemical group C(=O)(OCC1=CC=CC=C1)* 0.000 description 7
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- 239000010941 cobalt Substances 0.000 description 7
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- 229920003303 ion-exchange polymer Polymers 0.000 description 7
- YYROPELSRYBVMQ-UHFFFAOYSA-N 4-toluenesulfonyl chloride Chemical compound CC1=CC=C(S(Cl)(=O)=O)C=C1 YYROPELSRYBVMQ-UHFFFAOYSA-N 0.000 description 6
- WFDIJRYMOXRFFG-UHFFFAOYSA-N Acetic anhydride Chemical compound CC(=O)OC(C)=O WFDIJRYMOXRFFG-UHFFFAOYSA-N 0.000 description 6
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- 229910052793 cadmium Inorganic materials 0.000 description 6
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- 229910021529 ammonia Inorganic materials 0.000 description 5
- 239000007788 liquid Substances 0.000 description 5
- 230000001376 precipitating effect Effects 0.000 description 5
- DTQVDTLACAAQTR-UHFFFAOYSA-N trifluoroacetic acid Substances OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 5
- RYHBNJHYFVUHQT-UHFFFAOYSA-N 1,4-Dioxane Chemical compound C1COCCO1 RYHBNJHYFVUHQT-UHFFFAOYSA-N 0.000 description 4
- 230000006181 N-acylation Effects 0.000 description 4
- 230000000903 blocking effect Effects 0.000 description 4
- 238000010668 complexation reaction Methods 0.000 description 4
- 238000010511 deprotection reaction Methods 0.000 description 4
- CEAZRRDELHUEMR-UHFFFAOYSA-N gentamicin Chemical class O1C(C(C)NC)CCC(N)C1OC1C(O)C(OC2C(C(NC)C(C)(O)CO2)O)C(N)CC1N CEAZRRDELHUEMR-UHFFFAOYSA-N 0.000 description 4
- 230000007246 mechanism Effects 0.000 description 4
- 229920006395 saturated elastomer Polymers 0.000 description 4
- 239000000741 silica gel Substances 0.000 description 4
- 229910002027 silica gel Inorganic materials 0.000 description 4
- 239000007858 starting material Substances 0.000 description 4
- 229960000707 tobramycin Drugs 0.000 description 4
- JIAARYAFYJHUJI-UHFFFAOYSA-L zinc dichloride Chemical compound [Cl-].[Cl-].[Zn+2] JIAARYAFYJHUJI-UHFFFAOYSA-L 0.000 description 4
- ODIGIKRIUKFKHP-UHFFFAOYSA-N (n-propan-2-yloxycarbonylanilino) acetate Chemical compound CC(C)OC(=O)N(OC(C)=O)C1=CC=CC=C1 ODIGIKRIUKFKHP-UHFFFAOYSA-N 0.000 description 3
- 241000894006 Bacteria Species 0.000 description 3
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 3
- 229920005654 Sephadex Polymers 0.000 description 3
- 230000000844 anti-bacterial effect Effects 0.000 description 3
- 150000001721 carbon Chemical group 0.000 description 3
- 239000013522 chelant Substances 0.000 description 3
- 238000004090 dissolution Methods 0.000 description 3
- 239000000706 filtrate Substances 0.000 description 3
- 238000007327 hydrogenolysis reaction Methods 0.000 description 3
- 230000007062 hydrolysis Effects 0.000 description 3
- 238000006460 hydrolysis reaction Methods 0.000 description 3
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- 101100134922 Gallus gallus COR5 gene Proteins 0.000 description 1
- WHXSMMKQMYFTQS-UHFFFAOYSA-N Lithium Chemical compound [Li] WHXSMMKQMYFTQS-UHFFFAOYSA-N 0.000 description 1
- 239000012359 Methanesulfonyl chloride Substances 0.000 description 1
- OKIZCWYLBDKLSU-UHFFFAOYSA-M N,N,N-Trimethylmethanaminium chloride Chemical compound [Cl-].C[N+](C)(C)C OKIZCWYLBDKLSU-UHFFFAOYSA-M 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
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- 150000001342 alkaline earth metals Chemical class 0.000 description 1
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- 125000004103 aminoalkyl group Chemical group 0.000 description 1
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- 229940124350 antibacterial drug Drugs 0.000 description 1
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- 125000005099 aryl alkyl carbonyl group Chemical group 0.000 description 1
- 125000005161 aryl oxy carbonyl group Chemical group 0.000 description 1
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- OOMGRWNPCPICJN-UHFFFAOYSA-N benzyl (2,5-dioxopyrrolidin-3-yl) carbonate Chemical compound C1C(=O)NC(=O)C1OC(=O)OCC1=CC=CC=C1 OOMGRWNPCPICJN-UHFFFAOYSA-N 0.000 description 1
- HSDAJNMJOMSNEV-UHFFFAOYSA-N benzyl chloroformate Chemical compound ClC(=O)OCC1=CC=CC=C1 HSDAJNMJOMSNEV-UHFFFAOYSA-N 0.000 description 1
- 125000000051 benzyloxy group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])O* 0.000 description 1
- 238000005976 benzyloxycarbonylation reaction Methods 0.000 description 1
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- NCMHKCKGHRPLCM-UHFFFAOYSA-N caesium(1+) Chemical compound [Cs+] NCMHKCKGHRPLCM-UHFFFAOYSA-N 0.000 description 1
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- 239000001569 carbon dioxide Substances 0.000 description 1
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- 239000003518 caustics Substances 0.000 description 1
- 229940006165 cesium cation Drugs 0.000 description 1
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- FLASNYPZGWUPSU-SICDJOISSA-N chitosan Chemical compound O([C@@H]1[C@@H](CO)O[C@H]([C@@H]([C@H]1O)N)O[C@@H]1[C@@H](CO)O[C@H]([C@@H]([C@H]1O)N)O[C@@H]1[C@@H](CO)O[C@H]([C@@H]([C@H]1O)N)O[C@@H]1[C@@H](CO)O[C@H]([C@@H]([C@H]1O)N)O[C@@H]1[C@@H](CO)O[C@H]([C@@H]([C@H]1O)N)O[C@H]1[C@H](O)[C@H]([C@@H](O[C@@H]1CO)O[C@@H]1[C@H](O[C@@H](O[C@@H]2[C@H](O[C@@H](O)[C@H](N)[C@H]2O)CO)[C@H](N)[C@H]1O)CO)NC(=O)OC)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1N FLASNYPZGWUPSU-SICDJOISSA-N 0.000 description 1
- FZFAMSAMCHXGEF-UHFFFAOYSA-N chloro formate Chemical compound ClOC=O FZFAMSAMCHXGEF-UHFFFAOYSA-N 0.000 description 1
- AOGYCOYQMAVAFD-UHFFFAOYSA-N chlorocarbonic acid Chemical compound OC(Cl)=O AOGYCOYQMAVAFD-UHFFFAOYSA-N 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
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- 230000008878 coupling Effects 0.000 description 1
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- 238000002523 gelfiltration Methods 0.000 description 1
- XBECWGJPSXHFCS-UHFFFAOYSA-N imidazole-1-carbaldehyde Chemical compound O=CN1C=CN=C1 XBECWGJPSXHFCS-UHFFFAOYSA-N 0.000 description 1
- 230000001771 impaired effect Effects 0.000 description 1
- 238000011065 in-situ storage Methods 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 229910001853 inorganic hydroxide Inorganic materials 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 229910052744 lithium Inorganic materials 0.000 description 1
- HBUKSGRCRAUHHN-UHFFFAOYSA-L magnesium diacetate dihydrate Chemical compound O.O.[Mg++].CC([O-])=O.CC([O-])=O HBUKSGRCRAUHHN-UHFFFAOYSA-L 0.000 description 1
- 238000003328 mesylation reaction Methods 0.000 description 1
- QARBMVPHQWIHKH-UHFFFAOYSA-N methanesulfonyl chloride Chemical compound CS(Cl)(=O)=O QARBMVPHQWIHKH-UHFFFAOYSA-N 0.000 description 1
- GSYSFVSGPABNNL-UHFFFAOYSA-N methyl 2-dimethoxyphosphoryl-2-(phenylmethoxycarbonylamino)acetate Chemical group COC(=O)C(P(=O)(OC)OC)NC(=O)OCC1=CC=CC=C1 GSYSFVSGPABNNL-UHFFFAOYSA-N 0.000 description 1
- 125000000325 methylidene group Chemical group [H]C([H])=* 0.000 description 1
- PGXWDLGWMQIXDT-UHFFFAOYSA-N methylsulfinylmethane;hydrate Chemical compound O.CS(C)=O PGXWDLGWMQIXDT-UHFFFAOYSA-N 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 150000002815 nickel Chemical class 0.000 description 1
- FEMOMIGRRWSMCU-UHFFFAOYSA-N ninhydrin Chemical compound C1=CC=C2C(=O)C(O)(O)C(=O)C2=C1 FEMOMIGRRWSMCU-UHFFFAOYSA-N 0.000 description 1
- BSCHIACBONPEOB-UHFFFAOYSA-N oxolane;hydrate Chemical compound O.C1CCOC1 BSCHIACBONPEOB-UHFFFAOYSA-N 0.000 description 1
- MUMZUERVLWJKNR-UHFFFAOYSA-N oxoplatinum Chemical compound [Pt]=O MUMZUERVLWJKNR-UHFFFAOYSA-N 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 229910003445 palladium oxide Inorganic materials 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 125000000951 phenoxy group Chemical group [H]C1=C([H])C([H])=C(O*)C([H])=C1[H] 0.000 description 1
- 125000000612 phthaloyl group Chemical group C(C=1C(C(=O)*)=CC=CC1)(=O)* 0.000 description 1
- 229910003446 platinum oxide Inorganic materials 0.000 description 1
- WYVAMUWZEOHJOQ-UHFFFAOYSA-N propionic anhydride Chemical compound CCC(=O)OC(=O)CC WYVAMUWZEOHJOQ-UHFFFAOYSA-N 0.000 description 1
- 230000006289 propionylation Effects 0.000 description 1
- 238000010515 propionylation reaction Methods 0.000 description 1
- 239000003223 protective agent Substances 0.000 description 1
- 125000001453 quaternary ammonium group Chemical group 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- YGSDEFSMJLZEOE-UHFFFAOYSA-M salicylate Chemical compound OC1=CC=CC=C1C([O-])=O YGSDEFSMJLZEOE-UHFFFAOYSA-M 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 229910000029 sodium carbonate Inorganic materials 0.000 description 1
- 239000012312 sodium hydride Substances 0.000 description 1
- 229910000104 sodium hydride Inorganic materials 0.000 description 1
- ZAPXGKVOIFARAC-UHFFFAOYSA-M sodium;acetate;tetrahydrate Chemical compound O.O.O.O.[Na+].CC([O-])=O ZAPXGKVOIFARAC-UHFFFAOYSA-M 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 239000008247 solid mixture Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000005507 spraying Methods 0.000 description 1
- 230000000087 stabilizing effect Effects 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 125000001424 substituent group Chemical group 0.000 description 1
- 150000004763 sulfides Chemical class 0.000 description 1
- 229910052717 sulfur Inorganic materials 0.000 description 1
- 150000004685 tetrahydrates Chemical class 0.000 description 1
- 238000004809 thin layer chromatography Methods 0.000 description 1
- 238000007070 tosylation reaction Methods 0.000 description 1
- 238000005583 trifluoroacetylation reaction Methods 0.000 description 1
- 125000004950 trifluoroalkyl group Chemical group 0.000 description 1
- 125000004385 trihaloalkyl group Chemical group 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 238000012784 weak cation exchange Methods 0.000 description 1
- 238000010626 work up procedure Methods 0.000 description 1
- 239000011667 zinc carbonate Substances 0.000 description 1
- 235000004416 zinc carbonate Nutrition 0.000 description 1
- 229910000010 zinc carbonate Inorganic materials 0.000 description 1
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical class [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 description 1
- RCQZCHPRZSTYAX-UHFFFAOYSA-N zinc tetrahydrate Chemical compound O.O.O.O.[Zn] RCQZCHPRZSTYAX-UHFFFAOYSA-N 0.000 description 1
- JQBMTMBJMXRRCJ-UHFFFAOYSA-N zinc;dihydrate Chemical compound O.O.[Zn] JQBMTMBJMXRRCJ-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H23/00—Compounds containing boron, silicon or a metal, e.g. chelates or vitamin B12
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
- C07H15/20—Carbocyclic rings
- C07H15/22—Cyclohexane rings, substituted by nitrogen atoms
- C07H15/222—Cyclohexane rings substituted by at least two nitrogen atoms
- C07H15/226—Cyclohexane rings substituted by at least two nitrogen atoms with at least two saccharide radicals directly attached to the cyclohexane rings
- C07H15/234—Cyclohexane rings substituted by at least two nitrogen atoms with at least two saccharide radicals directly attached to the cyclohexane rings attached to non-adjacent ring carbon atoms of the cyclohexane rings, e.g. kanamycins, tobramycin, nebramycin, gentamicin A2
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Saccharide Compounds (AREA)
Abstract
Description
ÄESKOSLOVENSKA SOCIALISTICKĂ
REPUBLIKA (19)
POPIS VYNĂLEZU
K PATENTU 261853 (Î ) (B2)
ĂĆAD PRO VYNĂLEZYA OBJEVY (22) PĆihlĂĄĆĄeno 12 11 79 (21) (PV 7711-79) (32) (31) (33) PrĂĄvo pĆednosti od 11 11 78(138402) Japonsko (51) Int. Cl.4C 07 H 15/234 (40) ZveĆejnÄno 15 06 88(45) VydĂĄno 15 07 89 (72)
Autor vynĂĄlezu UMEZAWA HAMAO, UMEZAWA SUMIO, TOKIO, TSUCHIYA TSUTOMU,TAKAGI YASUSHI, JIKIHARA ΀ÎÎÎ, KANAGAWA (Japonsko) (73)
Majitel patentu ZAIDAN HOJIN BISEIBUTSU KAGAKU KENKYU KAI, TOKIO (Japonsko) (54) ZpĆŻsob vĂœroby selektivnÄ acylovanĂ©ho N-clirĂĄnÄnĂ©ho derivĂĄtuaminoglykosidovĂ©ho antibiotika 1
VynĂĄlez se tĂœkĂĄ zpĆŻsobu vĂœroby selek-tivnÄ acylovanĂ©ho N-chrĂĄnÄnĂ©ho derivĂĄtuaminoglykosidovĂ©ho antibiotika.
Tento vynĂĄlez se zvlĂĄĆĄtÄ tĂœkĂĄ novĂ©hozpĆŻsobu vĂœroby selektivnÄ chrĂĄnÄnĂ©ho N--acylcvanĂ©ho derivĂĄtu aminoglykosidovĂ©hoantibiotika, pĆi kterĂ©m nÄkterĂ© aminosku-piny nebo alkylaminoskupiny v urÄitĂœch po-lohĂĄch. aminoglykosidovĂ© molekuly jsou se-lektivnÄ chrĂĄnÄny neboli blokovĂĄny acylo-vou skupinou. VynĂĄlez se tudĂĆŸ tĂœkĂĄ novĂ©-ho zpĆŻsobu selektivnĂ ochrany nÄkterĂœchaminoskupin nebo alkylaminoskupin v ur-t^Ăœch polohĂĄch aminoglykosidavĂ©ho' anti-biotika a nalĂ©zĂĄ hlavnĂ pouĆŸitĂ pĆi vĂœrobÄselektivnÄ chrĂĄnÄnĂ©ho N-acylovanĂ©ho deri-vĂĄtu aminoglykosidovĂ©ho antibiotika, kterĂ©obsahuje deoxystreptaminovou strukturu ob-sahujĂcĂ 3â-aminoglykosyloTOu skupinu spo-jenou s 6-hydroxyskupinou deoxystreptami-novĂ© ÄĂĄsti v aminoglykosidovĂ© molekule.
AminoglykosidovĂ© antibiotikum, pouĆŸitel-nĂ© podle tohoto vynĂĄlezu, mĆŻĆŸe bĂœt pĆesnÄ-ji definovĂĄno jako aminoglykosidovĂ© anti-biotikum sestĂĄvajĂcĂ z 6-O-(3â-amino- nebo3â-alkylamino-3â-deoxyglykosyl) -2-deoxy-streptaininu, kterĂœ mĆŻĆŸe popĆĂpadÄ obsaho-vat 4-0-(6â-aminoglykosyl)ovĂœ substituent.TypickĂœmi pĆĂklady jsou kanamyciny, gent- 2 amiciny, sisomicin, netilmicin a verdami-cin.
Je znĂĄmo, ĆŸe aminoglykosidovĂĄ antibioti-ka, jako napĆĂklad kanamyciny, jsou lĂĄtkyobsahujĂcĂ nÄkolik aminoskupin a hydroxy-skupin, kterĂ© majĂ pomÄrnÄ vysokĂœ a roz-liÄnĂœ stupeĆ reaktivnosti. MnohĂ© druhy po-lysyntetickĂœch aminoglykosidovĂœch antibio-tik, kterĂ© jsou odvozenĂ© od pĆŻvodnĂch ami-noglykosidovĂœch antibiotik byly pĆipravenysynteticky. PĆi semisyntĂ©ze tÄchto derivĂĄtĆŻje Äasto nutnĂ© nebo vĂœhodnĂ© zajistit, abynÄkterĂ© aminoskupiny a/nebo nÄkterĂ© hyd-roxyskupiny ve vĂœchozĂm arainoglykosido-vĂ©m antibiotiku byly selektivnÄ chrĂĄnÄnyalespoĆ jednou vhodnou ochrannou skupi-nou.
Pro selektivnĂ ochranu aminoskupin a//nebo hydroxyskupin v aminoglykosidovĂ©mantibiotiku byly vyvinuty rĆŻznĂ© uĆŸiteÄnĂ©zpĆŻsoby, kterĂ© jsou jako takovĂ© pouĆŸitelnĂ©pro selektivnĂ ochranu hydroxyskupiny. ProselektivnĂ ochranu nÄkterĂœch vybranĂœch a-minoskupin z mnoĆŸstvĂ aminoskupin, kterĂ©jsou v aminoglykosidovĂ©m antibiotiku, jsouvĆĄak tyto· znĂĄmĂ© zpĆŻsoby buÄ tÄĆŸko prove-ditelnĂ©, nebo vyĆŸadujĂ nÄkterĂ© sloĆŸitĂ© ope-race. To je zpĆŻsobeno tĂm, ĆŸe vĆĄechny ami-noskupiny v aminoglykosidovĂ©m antibiotiku 261853 281853 '' Î nemajĂ vÄtĆĄĂ rozdĂl ve svĂ© reaktivnosti. Cha-rakteristickĂœm pĆĂkladem Je 6â-aminosku-pina kanamycinu A, avĆĄak takovĂĄ amino-nebo methylaminoskupina, kterĂĄ je vĂĄzĂĄnana urÄitĂœ atom uhlĂku, kterĂœ je naopak vĂĄ-zĂĄn na pouze jeden atom uhlĂku v amino-glykosidovĂ© molekule, vytvĂĄĆĂ vyĆĄĆĄĂ reaktiv-nost neĆŸ takovĂĄ amino- nebo methylamino-skupina, kterĂĄ je vĂĄzĂĄna na atom uhlĂku,kterĂœ je vĂĄzĂĄn na dva nebo vĂce atomĆŻ uh-lĂku v tĂ©to aminogrykosidovĂ© molekule. Ztohoto dĆŻvodu prvnĂ zmĂnÄnĂœ typ amino-nebo methylaminoskupiny je mnohem vĂceschopen reagovat s acylaÄnĂm Äinidlem, kte-rĂ© obsahuje acylovou skupinu urÄenou prozavedenĂ jako amino-ochrannou skupinu,neĆŸ druhĂœ zmĂnÄnĂœ typ amino- nebo methyl-aminoskupiny, takĆŸe N-chrĂĄnÄnĂœ derivĂĄt,kterĂœ mĂĄ prvnĂ typ amino- nebo methyla-minoskupiny pĆednostnÄ chrĂĄnÄnĂœ acylovouskupinou, mĆŻĆŸe bĂœt vyrĂĄbÄn s vyĆĄĆĄĂm vĂœ-tÄĆŸkem neĆŸ jinak N-chrĂĄnÄnĂ© derivĂĄty. PĆed nÄkolika lety nÄkteĆĂ z autorĆŻ toho-to vynĂĄlezu zjistili, ĆŸe kdyĆŸ aminoskupinaa hydroxyskupina jsou navzĂĄjem v soused-nĂch polohĂĄch v pĂĄru ve sterickĂ© konfigu-raci molekuly aminoglykosidovĂ©ho antibio-tika, mĆŻĆŸe bĂœt tato aminoskupina a hydro-xyskupina navzĂĄjem selektivnÄ slouÄena dotvaru cyklickĂ©ho karbamĂĄtu reakcĂ s hyd-ridem sodnĂœm, takĆŸe pĂĄr sestĂĄvajĂcĂ z ami-noskupiny a hydroxyskupiny mĆŻĆŸe bĂœt vcyklickĂ©m karbamĂĄtu chrĂĄnÄn souÄasnÄ bezblokovĂĄnĂ jinĂœch aminoskupin pĆĂtomnĂœchv tĂ©ĆŸe molekule [viz âJournal of Antibioticsâ25, 12, 741â742 [1972], a dĂĄle US paten-ty Ä. 3 925 354 a 3 965 0B9],
NedĂĄvno Nagabhushan a kol. zjistili, ĆŸekdyĆŸ se sĆŻl dvojmocnĂ©ho pĆechodnĂ©ho ko-vu (M+ + J zvolenĂ©ho ze souboru zahrnujĂ-cĂho dvojmocnou mÄÄ, nikl, kobalt a kad-mium nechĂĄ v inertnĂm organickĂ©m roz-pouĆĄtÄdle reagovat s aminoglykosidovĂœmantibiotikem ze souboru zahrnujĂcĂho 4-0-- (aminoglykosyl) -6-0- [ aminoglykosyl )-2-de-oxystreptamin, pĆedstavovanĂœ kanamyciny,gentamiciny a sisomicinem, tento dvojmoc-nĂœ kation pĆechodnĂ©ho kovu je komplexo-vĂĄn s dvojicĂ sestĂĄvajĂcĂ z aminoskupiny ahydroxyskupiny, kterĂ© jsou zvlĂĄĆĄtÄ ve vici-nĂĄlnĂm uspoĆĂĄdĂĄnĂ v molekule aminoglyko-sidu, ÄĂmĆŸ je vytvoĆen kationtovĂœ komplexaminoglykosidovĂ©ho antibiotika s pĆechod-nĂœm kovem (japonskĂœ zveĆejnÄnĂœ spis Ä.Sho-52-153 944 a US patent Ä. 4 136 254, u-dÄlenĂœ dne 23. ledna 1979 J. V tomto kationtovĂ©m komplexu amino-glykosidovĂ©ho antibiotika s transitnĂm ko-vem je komplexovanĂĄ aminoskupina chrĂĄ-nÄna kationtem dvojmocnĂ©ho pĆechodnĂ©hokovu. KdyĆŸ se potom tento komplex nechĂĄreagovat s nÄkterĂœm acylaÄnĂm Äinidlem ob-sahujĂcĂm acylovou skupinu, mohou bĂœt vkovovĂ©m komplexu acylovĂĄny pouze ne-skomplexovanĂ© aminoskupiny, kterĂ© nejsouchrĂĄnÄny kationtem dvojmocnĂ©ho kovu, tak-ĆŸe se dosĂĄhne selektivnĂ N-ochrany acylo- vou skupinou. To je ukĂĄzĂĄno dĂĄle s odvolĂĄ-nĂm na kanamycin A jako pĆĂklad. Je-li te-dy kation dvojmocnĂ©ho pĆechodnĂ©ho kovu(M+ + ) zvolenĂœ ze souboru zahrnujĂcĂhodvojmocnou mÄÄ, nikl, kobalt a kadmiumuveden do reakce s kanamycinem A, kom-plexaÄnĂ reakce kationtu dvojmocnĂ©ho ko-vu (M+ + J nastane mezi 1-aminoskupinou a2â-hydroxyskupinou a mezi 3â-aminoskupi-nou a 4â-hydroxyskupinou molekuly kana-mycinu A, jak znĂĄzorĆuje vzorec Î:
Ve vĂœĆĄe uvedenĂ© komplexaÄnĂ reakci jetudĂĆŸ zĆejmĂ©, ĆŸe je tĆeba alespoĆ 2 molĆŻ so-li pĆechodnĂ©ho kovu na 1 mol kanamycinuA. Ve vĂœslednĂ©m kovovĂ©m komplexu jsou1-amino- a 3â-aminoskupiny blokovĂĄny sou-ÄasnÄ. KdyĆŸ se tento komplex vzorce I*zpracuje s acylaÄnĂm Äinidlem obsahujĂcĂmacylovou skupinu, kterĂĄ je vhodnĂĄ jakoskupina pro ochranu aminoskupiny, znĂĄmĂĄpĆi obvyklĂ© syntĂ©ze polypeptidĆŻ, jsou acy-lovĂĄny pouze nezkomplexovanĂ© 3-amino- a6â-aminoskupiny, ÄĂmĆŸ se dostane 3,6â-di-N--acylovanĂœ derivĂĄt [âJournal of AmericanChemical Societyâ, 100, 5 253â5254 [1978J],
AutoĆi tohoto vynĂĄlezu vzali v Ășvahu vĂœ-ĆĄe uvedenou skuteÄnost, avĆĄak provedli dal-ĆĄĂ vlastnĂ vĂœzkumy interakce jinĂœch rozliÄ-nĂœch kationtĆŻ kovĆŻ s aminoglykosidovĂœmiantibiotiky jako je kanamycin A a kanamy-cin B, jakoĆŸ i s polosyntetickĂœmi derivĂĄtyaminoglykosidovĂœch antibiotik. Jako vĂœsle-dek zjistili, ĆŸe aÄkoliv kation dvojmocnĂ©hozinku vykazuje chovĂĄnĂ podstatnÄ odliĆĄnĂ©od kationtĆŻ z vĂœĆĄe uvedenĂ©ho souboru za-hrnujĂcĂho dvojmocnĂ© kationty niklu, kobal-tu, mÄdi a kadmia, je kation zinku schopnĂœsilnÄ vytvĂĄĆet komplex a chrĂĄnit jak 1-ami-no- nebo 1-alkylaminoskupinu, tak 3â-ami-no- nebo 3â-alkylaminoskupinu aminogly-kosidovĂ© slouÄeniny, napĆĂklad kanamycinuA, B nebo C, kterĂĄ obsahuje deoxystrepta- 261853 minovou ÄĂĄst zahrnujĂcĂ 3â-aminoglykosy-lovou skupinu nebo 3â-alkylaminoglykosy-lovou skupinu pĆipojenou k 6-hydroxysku-pinÄ zmĂnÄnĂ© deoxystreptaminovĂ© ÄĂĄsti.
Podle Nagabhushana a kol. by bylo moĆŸ-no oÄekĂĄvat, ĆŸe kdyĆŸ se nechĂĄ reagovat ka-tion dvojmocnĂ©ho niklu, dvojmocnĂ©ho ko-baltu, dvojmccnĂ© mÄdi nebo dvojmocnĂ©hokadmia napĆĂklad s kanamycinem. B, mÄlaby se vytvoĆit kovovĂĄ koplexnĂ sĆŻl kana-mycinu B vzorce II
Tento pĆedpoklad je moĆŸno podepĆĂt po-jednĂĄnĂm Nagabhushana a kol. ve vĂœĆĄe u-vedenĂ©m âJournal cf American ChemicalSocietyâ, podle kterĂ©ho pĂĄry vicinĂĄlnĂch a-mincihydroxyskupin by mohly vytvĂĄĆet re-versibilnĂ komplexy s kationty dvojmocnĂœchpĆechodnĂœch kovĆŻ vzhledem ke skuteÄnos-ti, ĆŸe kauamyciu B obsahuje tĆi pĂĄry vici-nĂĄlnĂch aminohydroxyskupin mezi poloha-mi 1 a 2â, mezi polohami 2â a 3â a mezi po-lohami 2â a 3â molekuly kanamycinu B.
NicmĂ©nÄ bylo nynĂ zjiĆĄtÄno, ĆŸe kdyĆŸ sekanamycin B nechĂĄ reagovat se zineÄnatĂœmkationtem, potom vytvoĆenĂœ komplex kana-mycinu B se zineÄnatou solĂ obsahuje volnĂ©2â-amino- a 3â-hydroxyskupiny, kterĂ© nejsoublok' vĂĄny kationtem zinku, coĆŸ je protikla-dem k pĆedpokladu Nagabhushana a kol. IkdyĆŸ nastane komplexctvornĂĄ reakce zineÄ-natĂ©ho kationtu s 2â-amino- a 3â-hydroxy-skupinou, sĂla vytvĂĄĆenĂ komplexu je velminĂzkĂĄ, takĆŸe v praxi nenĂ 2â-amino- a 3â-hyd-roxyskupina blokovĂĄna. KdyĆŸ se tedy kom-plex kanamycinu B se zineÄnatĂœm kationtemacyluje napĆĂklad reakcĂ s N-benzyloxykar-bonyloxysukcinimidem k zavedenĂ benzylo-xykarbonylovĂ© skupiny jako acylovĂ© skupi-ny chrĂĄnĂcĂ aminoskupinu, vytvoĆĂ se tri--3,2â,6â-N-acylovanĂœ derivĂĄt, ve kterĂ©m tĆiaminoskupiny, 3-, 2â- a 6â-aminoskupina by-ly acylovĂĄny, a to ve skuteÄnosti pĆi vyĆĄ-ĆĄĂm vĂœtÄĆŸku neĆŸ u jinak N-acylovanĂœch de-rivĂĄtĆŻ, avĆĄak potom nemĆŻĆŸe bĂœt ve skuteÄ-nosti zĂskĂĄn 3,6â-di-N-acylovanĂœ derivĂĄt (vizdĂĄle uvedenĂœ pĆĂklad 19 J. Tato experimen- tĂĄlnĂ skuteÄnost ukazuje, ĆŸe zineÄnatĂœ ka-tion mĂĄ jinĂ© chovĂĄnĂ neĆŸ vĂœĆĄe uvedenĂ© ka-tionty ÄtyĆ pĆechodnĂœch kovĆŻ, zejmĂ©na vtom smÄru, ĆŸe zineÄnatĂœ kation nevytvĂĄĆĂkomplexy s vicinĂĄlnĂm pĂĄrem 2â-aminc- a3â-hydroxyskupiny.
Jako dalĆĄĂ pĆĂklad se uvĂĄdĂ, ĆŸe se kdyĆŸkanamycin A nechĂĄ reagovat s zineÄnatĂœmkationtem s nĂĄsledujĂcĂ acylacĂ benzyloxy-karbonylovou skupinou (vztahuje se k vĂœĆĄeuvedenĂ©mu vzorci T], pozoruje se, ĆŸe jakohlavnĂ produkt acylace se vytvoĆĂ 3,6â-di-N--benzyloxykarbonylkanamycin A v pĆĂpadÄ,ĆŸe zineÄnatĂœ kation je pĆĂtomen v mnoĆŸstvĂnepatrnÄ vyĆĄĆĄĂm neĆŸ 1 mol na 1 mol kana-mycinu A. V tomto pĆĂpadÄ je tĆeba pozna-menat, ĆŸe tato acylaÄnĂ reakce vytvĂĄĆĂ 1,3,-6â,3â-tetra-N-benzyloxakarbonylovĂœ derivĂĄtkanamycinu A a dĂĄle souÄasnÄ v jistĂ©m roz-sahu neacylovanĂœ, poÄĂĄteÄnĂ kanamycin A,avĆĄak vytvĂĄĆĂ tri-N-benzyloxykarbnnylovĂœderivĂĄt kanamycinu A pouze s nĂzkĂœm vĂœ-tÄĆŸkem, aÄkoliv podle vysvÄtlenĂ mechanis-mu reakce podle Nagabhushana a kol. byse oÄekĂĄvalo, ĆŸe tri-N-benzyloxykarbonylo-vĂœ derivĂĄt se vytvoĆĂ s vyĆĄĆĄĂm vĂœtÄĆŸkem neĆŸjinĂ© N-acylovanĂ© derivĂĄty (viz pĆĂklad 7 u-vedenĂœ dĂĄle). V popise a zejmĂ©na v bodÄ 4nĂĄrokĆŻ US patentu Ä. 4 136 254 Nagabhu-sahan a kol. uvedli k tomuto jevu, ĆŸe sĆŻldvojmocnĂ©ho pĆechodnĂ©ho kovu, napĆĂkladmÄdi, niklu, kobaltu atd. je nutno pouĆŸĂt vcelkovĂ©m mnoĆŸstvĂ alespoĆ 2 molĆŻ na 1mol kanamycinu A pro vytvoĆenĂ komplex-nĂ soli kanamycinu A s pĆechodnĂœm kovem,jak je zĆejmĂ© z vĂœĆĄe uvedenĂ©ho vzorce I*.
Pokus autorĆŻ tohoto vynĂĄlezu ukĂĄzal, ĆŸena rozdĂl od katioatĆŻ ÄtyĆ uvedenĂœch pĆe-chodnĂœch kovĆŻ je zineÄnatĂœ kation scho-pen dosĂĄhnout blokovĂĄnĂ 1-amino- a 3â-ami-noskupiny kanamycinu A, kdyĆŸ se kationpouĆŸije v celkovĂ©m mnoĆŸstvĂ alespoĆ 1 mo-lu na 1 mol kanamycinu A. Podle zkouĆĄkydĂĄle bylo zjiĆĄtÄno, ĆŸe kdyĆŸ se k reakci po-uĆŸije nikelnatĂ© soli v mnoĆŸstvĂ nepatrnÄvyĆĄĆĄĂm neĆŸ 1 mol na 1 mol kanamycinu A,s nĂĄsledujĂcĂ acylacĂ vĂœslednĂ© komplexnĂsoli kanamycinu A s niklem benzyloxykar-bonylovou skupinou, zĂskĂĄ se pouze velminĂzkĂœ vĂœtÄĆŸek 3,6â-di-N-benzyloxykarbonyl-kanamycinu A, kterouĆŸto· slouÄeninu by by-lo moĆŸnĂ© zĂskat s vĂœznamnĂœm vĂœtÄĆŸkem a-cylacĂ komplexnĂ soli kanamycinu A se zin-kem (viz pĆĂklad 7 uvedenĂœ dĂĄle). Z vĂœĆĄeuvedenĂœch skuteÄnostĂ bylo vyvozeno, ĆŸezineÄnatĂœ kation vytvĂĄĆĂ mechanismus vznikukrmplexu s nÄkterĂœm aminoglykosidem, kte-rĂœ je odliĆĄnĂœ cd mechanismu vytvĂĄĆenĂ kom-plexu s kationtem dvojmocnĂ©ho niklu, ko-baltu, mÄdi a kadmia a ĆŸe kationtovĂœ kom-plex aminoglykosidu se zinkem mĂĄ stabilitukomplexu, kterĂĄ je odliĆĄnĂĄ od stability ka-tlontovĂ©ho komplexu aminoglykosidu sdvojmocnĂœm niklem, kobaltem, mÄdĂ nebokadmiem. Pro komplexaci zineÄnatĂ©ho ka-tiontu s aminoglykosidovĂœm antibiotikem 261853 7 mĆŻĆŸe bĂœt zineÄnatĂœ kation pouĆŸit ve formÄzineÄnatĂ© soli, kterĂĄ mĂĄ vĂœhodu, ĆŸe je lev-nĂĄ a pravdÄpodobnÄ nenĂ zdrojem zneÄiĆĄ-tÄnĂ okolnĂho prostĆedĂ.
AutoĆi tohoto vynĂĄlezu tudĂĆŸ zjistili, ĆŸekdyĆŸ se zineÄnatĂœ kationt nechĂĄ reagovatv inertnĂm organickĂ©m rozpouĆĄtÄdle s ami-noglykosidovĂœm antibiotikem, kterĂ© obsa-huje deoxystreptaminovou ÄĂĄst zahrnujĂcĂ 3--aminoglykosylovou nebo 3-alkylaminogly-kosylovou skupinu spojenou s 6-hydroxy-skupinou deoxystreptaminovĂ© ÄĂĄsti a pĆĂ-padnÄ zahrnujĂcĂ aminoglykosylovou skupi-nu spojenou se 4-hydroxyskupinou deoxy-streptaminovĂ© ÄĂĄsti, zineÄnatĂœ kation vytvĂĄ-ĆĂ komplex s pĂĄry aminohydroxylovĂœch sku-pin umĂstÄnĂœch v urÄitĂœch polohĂĄch, kterĂ©mohou bĂœt rozliÄnĂ© v zĂĄvislosti na vlast-nostech aminoglykosidovĂ©ho antibiotika, aĆŸe kdyĆŸ takto vytvoĆenĂœ kationtovĂœ kom-plex aminoglykosidovĂ©ho antibiotika sezinkem se nechĂĄ reagovat s acylaÄnĂm Äi-nidlem obsahujĂcĂm acylovou skupinu po-uĆŸĂvanou obvykle pro zavedenĂ ochrannĂ©aminoskupiny pĆi syntĂ©ze polypeptidĆŻ, totoacylaÄnĂ Äinidlo acyluje alespoĆ jednu ztakovĂœch aminoskupin v aminoglykosidovĂ©mantibiotiku, kterĂ© nevytvĂĄĆejĂ komplex atedy nejsou blokovĂĄny zineÄnatĂœm kation-tem, takĆŸe takto acylovanĂĄ aminoskupinaje chrĂĄnÄna, a dĂĄle, ĆŸe kdyĆŸ vĂœslednĂœ pro-dukt acylace, tj. kationtovĂœ komplex ami-noglykosidovĂ©ho antibiotika se zinkem ob-sahujĂcĂ acylovanou aminoskupinu nebo acy-lovanĂ© aminoskupiny se nechĂĄ reagovat svhodnĂœm Äinidlem, kterĂ© odstranĂ zineÄna-tĂœ kation z tohoto produktu acylace, kom-plex zinku se rozruĆĄĂ, coĆŸ poskytne selek-tivnÄ chrĂĄnÄnĂœ N-acylovanĂœ derivĂĄt amino-glykosidovĂ©ho antibiotika, u kterĂ©ho pĆŻvod-nÄ se zinkem nekomplexovanĂĄ aminoskupi-na nebo aminoskupiny byly selektivnÄ chrĂĄ-nÄny acylskupinou.
VynĂĄlez tedy ĆeĆĄĂ zpĆŻsob vĂœroby selektiv-nÄ acylovanĂ©ho N-chrĂĄnÄnĂ©ho derivĂĄtu ami-noglykosidovĂ©ho antibiotika.
PĆedmÄtem vynĂĄlezu je zpĆŻsob vĂœroby se-lektivnÄ acylovanĂ©ho N-chrĂĄnÄnĂ©ho derivĂĄ-tu aminoglykosidovĂ©ho antibiotika obsahu-jĂcĂho· 4-0- (aminoglykosyl j -6-0- (3â-amino-nebo 3â-methylamino-3â-deoxyglykosyl) -2--deoxystreptamin, ve kterĂ©m 1-amino- a 3â--aminoskupiny jsou nechrĂĄnÄny, avĆĄakvĆĄechny ostatnĂ aminoskupiny jsou chrĂĄnÄ-ny amino-ochrannou acylskupinou, obecnĂ©-ho vzorce I kde Râ znamenĂĄ atom vodĂku nebo ethylovouskupinu, G znamenĂĄ formyl, alkanoyl se 2 aĆŸ 5atomy uhlĂku, trifluoroalkanoyl se 2 aĆŸ 5atomy uhlĂku, alkoxykarbonyl s 1 aĆŸ 4 ato-my uhlĂku v alkoxylovĂ© ÄĂĄsti, fenoxykarbo-nyl, fenylalkyloxykarbonyl s 1 aĆŸ 4 atomyuhlĂku v alkylovĂ© ÄĂĄsti nebo p-methoxyfe-nylalkoxykarbonyl s 1 aĆŸ 4 atomy uhlĂku valkoxylovĂ© ÄĂĄsti, Q1 znamenĂĄ N-chrĂĄnÄnou aminoglykosy-lovou skupinu obecnĂ©ho vzorce Ila
W znamenĂĄ hydroxyskupinu nebo N-chrĂĄ-nÄnou aminoskupinu vzorce âNHG, kde G mĂĄ vĂœznam definovanĂœ vĂœĆĄe, X znamenĂĄ atom vodĂku nebo hydroxy-skupinu, Y znamenĂĄ atom vodĂku nebo hydroxy-skupinu, Zâ znamenĂĄ atom vodĂku, hydroxyskupi-nu, N-chrĂĄnÄnou aminoskupinu vzorceâNHG nebo N-chrĂĄnÄnou alkylaminosku-pinu vzorce Râ /
âN \
G ve kterĂœch G mĂĄ vĂœznam definovanĂœ vĂœĆĄe a Râ znamenĂĄ methyl, Zâ znamenĂĄ atom vodĂku nebo methylo-vou skupinu, nebo QL znamenĂĄ N-chrĂĄnÄnou 3â,4'-dideoxy-3â--eno-aminoglykosylovou skupinu obecnĂ©hovzorce lila
261833 10 ve kterĂ©m G mĂĄ vĂœznam definovanĂœ vĂœĆĄe, neboQ1 znamenĂĄ N-chrĂĄnÄnou 3â,4â-dideoxy-4â- -eno-aminoglykosylovou skupinu obecnĂ©hovzorce IVa
R"-CH-NHG
ve kterĂ©m Râ â znamenĂĄ atom vodĂku nebo methylo-vou skupinu a G mĂĄ vĂœznam definovanĂœ vĂœĆĄe, znamenĂĄ 3â-amino-3â-deoxy dykosylo- vou skupinu obecnĂ©ho vzorce Va
ve kterĂ©m M znamenĂĄ hydroxyskupinu nebo atomvodĂku a M; znamenĂĄ hydroxyskupinu nebo atomvodĂku, nebo· Q2 znamenĂĄ 3â-methylamino-3â-deoxygly-kosyiovou skupinu obecnĂ©ho vzorce Via
ve kterém
Rââ znamenĂĄ atom vodĂku nebo methylo-vou skupinu, kterĂœ spoÄĂvĂĄ v tom, ĆŸe sesĆŻl zineÄnatĂ©ho katiĂłntu s anorganickou ne-bo organickou kyselinou nechĂĄ reagovat saminoglykotsidovĂœm antiblotikem obecnĂ©hovzorce VII kde Râ znamenĂĄ atom vodĂku nebo ethylovouskupinu, Q3 znamenĂĄ aminoglykosylovou skupinuobecnĂ©ho vzorce lib
kde Wâ znamenĂĄ hydroxyskupinu nebo amino-skupinu, X znamenĂĄ atom vodĂku nebo hydroxy-skupinu, Y znamenĂĄ atom vodĂku nebo hydroxy-skupinu, Zââ znamenĂĄ atom vodĂku, hydroxyskupi-nu, aminoskupinu nebo methylaminoskupi-nu obecnĂ©ho vzorce âNHRâ, kde Râ znamenĂĄ methylovou skupinu, Zâ znamenĂĄ atom vodĂku nebo· methylo-vou skupinu nebo Q3 znamenĂĄ 3â,4â-dideoxy-3â-eno-aminogly-kosylovou skupinu vzorce Illb
nebo' Q3 znamenĂĄ 3â,4â-dideoxy-4â-eno-aminogly- kosylovou skupinu obecnĂ©ho vzorce IVb 261853 11
kde Rââ znamenĂĄ atom vodĂku nebo methylo-vou skupinu a Q4 znamenĂĄ 3â-amino-3â-deoxyglykosylo-vou skupinu nebo 3â-methylamino-3â-deoxy-glykosylovou skupinu shodnou s vĂœĆĄe uve-denou skupinou Q2 obecnĂ©ho vzorce Va ne-bo Via, v molĂĄrnĂm pomÄru alespoĆ 1 dĂ-lu molĂĄrnĂho, s vĂœhodou 2 aĆŸ 6 dĂlĆŻ molĂĄr-nĂch soli zineÄnatĂ©ho kationtu s anorganic-kou nebo organickou kyselinou na 1 dĂlmolĂĄrnĂ aminoglykosidovĂ©ho antibiotika o-becnĂ©ho vzorce VII za teploty mezi â10 a100 °C v inertnĂm organickĂ©m rozpouĆĄtÄdlezvolenĂ©m ze souboru zahrnujĂcĂho dime-thylsulfoxid, vodnĂœ dimethylsulfoxid, dime-thylformamid, vodnĂœ dimethylformamid,smÄs dimethylsulfoxidu a dimethylformami-du, tetrahydrofuranu, vodnĂœ tetrahydrofu-ran, methynol, vodnĂœ methancl, ethanol avodnĂœ ethanol, popĆĂpadÄ v pĆĂtomnosti oc-tanu sodnĂ©ho, za vzniku kationtovĂ©ho kom-plexu aminoglykosidovĂ©ho antibiotika sezinkem, potĂ© se tento kationtovĂœ komplexaminoglykosidovĂ©ho antibiotika se zinkemnechĂĄ reagovat s acylaÄnĂm Äinidlem zvo-lenĂœm ze souboru zahrnujĂcĂho karboxylo-vou kyselinu obecnĂ©ho vzorce IVa RRCOOH (IVa) kde R5 znamenĂĄ atom vodĂku, alkylovou sku-pinu s 1 aĆŸ 4 atomy uhlĂku, trifluoralkylo-vou skupinu s 1 aĆŸ 4 atomy uhlĂku, nebos halogenidem, anhydridem nebo aktivnĂmesterem vĂœĆĄe uvedenĂ© karboxylovĂ© kyseli-ny obecnĂ©ho vzorce IVa, chloroformiĂĄt o-becnĂ©hOâ vzorce IVb R6OâCOâCl (IVb) p-nitrofenylkarbonĂĄt obecnĂ©ho vzorce IVcRfiOâCOâOâC6H5âpâNO·, (IVc) aktivnĂ N-hydroxysukcinimidester obecnĂ©ho vzorce IVd 12 0 0 (IVd) a azidoformiĂĄt obecnĂ©ho vzorce IVeReOâCOâN3 (IVe) v kterĂœchĆŸto vzorcĂch
Rr> mĂĄ vĂœĆĄe uvedenĂœ vĂœznam a R6 znamenĂĄ alkylovou skupinu s 1 aĆŸ 4atomy uhlĂku, fenylovou skupinu, fenylal-kylovou skupinu s 1 aĆŸ 4 atomy v alkylovĂ©ÄĂĄsti uhlĂku nebo p-methoxyfenylalkylovouskupinu s 1 aĆŸ 4 atomy uhlĂku, za teplotyod â20 do 100 °C, pro acylaci nezkomple-xovanĂœch aminoskupin pĆĂtomnĂœch v ka-tiontovĂ©m komplexu aminoglykosidovĂ©ho·antibiotika se zinkem a tedy pro vytvoĆenĂkationtovĂ©ho· komplexu N-acylovanĂ©ho ami-noglykosidovĂ©ho antibiotika se zinkem apotom se kationtovĂœ komplex N-acylovanĂ©-ho aminoglykosidovĂ©ho antibiotika se zin-kem nechĂĄ reagovat s vodou nebo s vod-nĂœm nebo bezvodĂœm polĂĄrnĂm organickĂœmrozpouĆĄtÄdlem zvolenĂœm ze souboru zahr-nujĂcĂho methanol, ethanol, kapalnĂœ amo-niak, ethylamin a triethylamin, nebo· se si-rovodĂkem, sirnĂkem alkalickĂ©ho kovu nebosirnĂkem kovu alkalickĂ© zeminy nebo shydroxidem amonnĂœm ve vodÄ nebo· ka-tiontomÄniÄovou pryskyĆicĂ obsahujĂcĂ funk-ce karboxylovĂ© nebo sulfonovĂ© kyseliny, ne-bo s aniontomÄniÄovou pryskyĆicĂ obsahujĂ-cĂ amoniovĂ© funkce, nebo chelatomÄniÄovoupryskyĆicĂ obsahujĂcĂ kovovĂ© chelatizaÄnĂfunkce nebo s chitinem nebo chitosanem ja-ko ve vodÄ nerozpustnĂœm vyĆĄĆĄĂm polyme-rem obsahujĂcĂm funkce schopnĂ© slouÄenĂ skovem, za teploty mezi â10 a 100 °C, proodstranÄnĂ zineÄnatĂœch kationtĆŻ z komple-xu a pro vytvoĆenĂ N-acylovanĂ©ho amino-glykosidovĂ©ho antibiotika obecnĂ©ho vzorceI.
ZpĆŻsob podle vynĂĄlezu je vhodnĂœ pro vĂœ-robu selektivnÄ acylovanĂ©ho N-chrĂĄnÄnĂ©hoderivĂĄtu aminoglykosidovĂ©ho antibiotika a-cylacĂ nÄkterĂœch aminoskupin, jinĂœch neĆŸjsou 1- a 3â-aminoskupiny vĂœchozĂho· ami-noglykosidovĂ©ho antibiotika, a takovĂœ se-lektivnÄ N-chrĂĄnÄnĂœ derivĂĄt je vhodnĂœ prochemickou syntĂ©zu 1-N-aminoacylovanĂœchderivĂĄtĆŻ aminoglykosidovĂœch antibiotik,napĆĂklad kanamycinC^, vÄetnÄ amikacinu(âJournal of Antibioticsâ 25, 695â708 (1972)], o kterĂ©m bylo v minulĂœch letech 261853 13 11 dokĂĄzĂĄno, ĆŸe je ĂșÄinnĂœm antibakteriĂĄlnĂmlĂ©Äivem. Tyto 1-N-aminoacylovanĂ© derivĂĄtyaminoglykosidovĂœch antibiotik zahrnujĂ iderivĂĄty odvozenĂ© od poÄetnĂ©ho druhu aroi-noglykosidĂș, jako jsou kanamycin A, kana-mycin B, kanamycin C, gentamiciny, siso-micin a jinĂ©, jakoĆŸ i jejich rozliÄnĂ© deoxy-derivĂĄty, avĆĄak vĆĄechny se shodujĂ v tem,ĆŸe jejich J-aminoskupina je acylovĂĄna nÄ-kterou «-hydroxy-w-aminoalkanoylovou sku-pinou (viz US patenty Ä. 3 781 268, 3 939 143,3 940 382 a 4 001 208). Vlivem tĂ©to 1-N-ami-noacylace dostĂĄvajĂ aminoglykosidovĂĄ an-tibiotika antibakteriĂĄlnĂ aktivitu vĆŻÄi odol-nĂœm bakteriĂm, proti kterĂœm souÄasnĂĄ an-tibiotika nejsou ĂșÄinnĂĄ, a tato aminoglyko-sidovĂĄ antibiotika takĂ© zĂskĂĄvajĂ zlepĆĄenouantibakteriĂĄlnĂ aktivitu proti ĆĄirĆĄĂ oblastikmenĆŻ bakteriĂ ve srovnĂĄnĂ s dosavadnĂmiaminoglykosidovĂœmi antibiotiky.
NynĂ bude podrobnÄji popsĂĄno provĂĄdÄnĂzpĆŻsobu podle pĆedloĆŸenĂ©ho vynĂĄlezu.
AminoglykosidovĂ© antibiotikum, kterĂ© mĂĄbĂœt uvedeno do reakce s kationtem zinku kvytvoĆenĂ komplexu zinku, kterĂœ mĆŻĆŸe bĂœtrovnÄĆŸ oznaÄen jako komplexnĂ sĆŻl zinku,podle pĆedloĆŸenĂ©ho vynĂĄlezu, zahrnuje ta-kovĂĄ aminoglykosidovĂĄ antibiotika obsahu-jĂcĂ deoxy-streptaminovou strukturou, jejĂĆŸ6-liydroxyskupina je substituovĂĄna 3-amino-glykosylovou nebo 3-alkylaminoglykosy-lovou skupinou, a jejĂĆŸ 4-hydroxy-skupinamĆŻĆŸe bĂœt pĆĂpadnÄ substituovĂĄna nÄkterouaminoglykosylovou skupinou. PĆesnÄji ĆeÄeno, aminoglykosidovĂ© anti-biotikum pouĆŸitĂ© v pĆedloĆŸenĂ©m vynĂĄlezupro vytvoĆenĂ komplexu zinkovĂ©ho kation-tu mĆŻĆŸe bĂœt definovĂĄno jako takovĂ©, kterĂ©obsahuje 6-O-(3â-amino- nebo 3â-alkylami-no-3â-deoxyglykosyl j -2-dcoxystreptamin ma-jĂcĂ podle volby 4-0-(amino-glykosyl)-ovouskupinu. KromÄ toho· aminoglykosidovĂ© an-tibiotikum mĆŻĆŸe bĂœt nÄkterĂœ 1-N-al.kylamĂ-noglykosid, napĆĂklad netilmicin. Jako pĆĂ-klady aminoglykosidovĂœch antibiotik tĆĂdypouĆŸitelnĂ© v pĆedloĆŸenĂ©m vynĂĄlezu mohoubĂœt uvedeny skupina antibiotik kanamycinuA, jako samotnĂœ kanamycin A, 6*-N-alkyl-kanamycin A, zvlĂĄĆĄtÄ 6â-N-methylkanamy-crn A, 3â-deoxykanamycin A, 6â-N-methyl-3â-deoxykanamycin A, 4â-deoxykanamycin A,6â-N-methyl-4â-deoxykanamycin A, 3â,4â-dide-oxykanamyciu A (viz japonskou pat. pĆihl.Ä. 11 402/79), a 6â-deoxy- nebo 4â,6â-dide-oxykanamycin A (viz japonskou pat. pĆihl.Ä. 34 733/79), skupina antibiotik kanamyci-nu B, to je kanamycin B samotnĂœ, 3â-deoxy-kanamycin B (to je tobramycin), 4â-deoxy-kanamycin B, 3â,4â-dideoxykanamycin B (toje dibekacin), 3â,4â-dideoxy-3â-eno-kanamy-cin B. 6â-N-mcthyl-3â,4â-dideoxykanamycin B,skupina antibiotik kanamycinu C, to je sa-motnĂœ kanamycin C, 3â-deoxykanaraycin C,3â,4â-dideoxykanamycin C, gentamiciny A,B a C, verdamicin, sisomicin a netilmicin(to je 1-N-ethylsisomycin), jakoĆŸ i jinĂ© znĂĄ-mĂ© aminoglykosidy.
ZpĆŻsob podle prvnĂ myĆĄlenky pĆedloĆŸenĂ©-ho· vynĂĄlezu je pouĆŸitelnĂœ nejen na takovĂĄnovĂĄ aminoglykosidovĂĄ antibiotika, kterĂĄdnes jeĆĄtÄ nejsou znĂĄma a budou objevenav budoucnosti, nĂœbrĆŸ i na novĂ© semisynte-tickĂ© derivĂĄty aminoglykosidovĂœch antibio-tik, kterĂ© budou v budoucnosti vyrĂĄbÄnychemickou transformacĂ znĂĄmĂœch amino-glykosidovĂœch antibiotik.
TypickĂ© pĆĂklady aminoglykosidovĂœch an-tibiotik, na kterĂ© je pouĆŸitelnĂœ pĆedloĆŸenĂœvynĂĄlez, jsou kanamycin A, kanamycin B,kanamycin C, deoxy-derivĂĄty tÄchto kana-mycinĆŻ jakoĆŸ i jejich 6â-N-alkyl-derivĂĄty,kterĂ© jsou definovĂĄny tĂmto obecnĂœm vzor-
kde R1 je hydroxyskupina nebo aminoskupina,R2 a R3 jsou kaĆŸdĂœ buÄ atom vodĂku, ne-bo hydroxyskupina, a R4 je hydroxyskupinanebo aminoskupina nebo alkylaminoskupi-na obsahujĂcĂ alkyl s 1 aĆŸ 4 atomy uhlĂku,zejmĂ©na methylaminoskupina.
Pro vytvoĆenĂ komplexu aminoglykosido-vĂ©ho antibiotika se zinkovĂœm kationtem re-akcĂ aminoglykosidovĂ©ho antibiotika se zin-kovĂœm kationtem podle pĆedloĆŸenĂ©ho· vy-nĂĄlezu se pĆĂsluĆĄnĂ© aminoglykosidovĂ© an-tibiotikum, buÄ ve formÄ volnĂ© bĂĄze, nebove formÄ jeho aditivnĂ soli s kyselinou roz-pustĂ nebo suspenduje ve vhodnĂ©m orga-nickĂ©m rozpouĆĄtÄdle a vĂœslednĂ©mu roztokunebo suspenzi se pĆidĂĄ vhodnĂĄ sĆŻl zinkuv mnoĆŸstvĂ alespoĆ 1 molu na 1 mol pou-ĆŸitĂ©ho aminoglykosidovĂ©ho antibiotika. Protento ĂșÄel mĆŻĆŸe bĂœt pouĆŸito kterĂ©koli obvyk-lĂ© organickĂ© rozpouĆĄtÄdlo, pokud zinkovĂœkomplex vytvoĆenĂœ po pĆidĂĄnĂ soli zinku jealespoĆ ÄĂĄsteÄnÄ v nÄm rozpustnĂœ. PĆed-nostnÄ by vĆĄak mÄlo bĂœt vylouÄeno· pouĆŸitĂvelkĂ©ho objemu polĂĄrnĂho organickĂ©ho roz-pouĆĄtÄdla a zejmĂ©na vÄtĆĄĂho objemu vody,neboĆ„ pĆĂtomnost polĂĄrnĂho organickĂ©horozpouĆĄtÄdla a vody by sniĆŸovala stabilituvytvoĆenĂ©ho vĂœslednĂ©ho komplexu amino-glykosidovĂ©ho antibiotika a kationtu zinku,takĆŸe nĂĄsledujĂcĂ acylaÄnĂ reakce k zavede- 261853 13 16 nĂ ochrannĂ© aminoskupiny by mohla dĂĄtneuspokojivĂœ vĂœsledek.
Je tedy ĆŸĂĄdoucĂ pouĆŸĂt organickĂ© roz-pouĆĄtÄdlo s velkou schopnostĂ rozpouĆĄtÄcĂ,napĆĂklad dimethyl-sulfoxid jako rozpouĆĄ-tÄdlo, ve kterĂ©m mĂĄ bĂœt vytvoĆen zinkovĂœkomplex, je vĆĄak moĆŸnĂ© pouĆŸĂt i vodnĂœ di-methlsulfoxid, dimethylformamid, vodnĂœ di-methylformamid, smÄs dimethylsiĂlfoxidu adimethylformamidu, tetrahydrofuran, vodnĂœtetrahydrofuran, a takĂ© niĆŸĆĄĂ alkanol jakomethanol, ethanol a vodnĂœ methanol.
Kationt zinku mĆŻĆŸe bĂœt pĆidĂĄn ve formÄsoli zinku do systĂ©mu, ve kterĂ©m se vytvĂĄĆĂzinkovĂœ komplex. Pro tento ĂșÄel v pĆedlo-ĆŸenĂ©m vynĂĄlezu mĆŻĆŸe bĂœt pouĆŸita nÄkterĂĄsĆŻl zinku vytvoĆenĂĄ reakcĂ zinkovĂ©ho ka-tiontu s obvyklou organickou nebo· anorga-nickou kyselinou. ObecnÄ je vĆĄak vĂœhodnĂ©pouĆŸĂt zinkovou sĆŻl nÄkterĂ© slabĂ© kyseliny,napĆĂklad octan zineÄnatĂœ, neboĆ„ je obvyk-lĂ©, ĆŸe mezi komplexy kovĆŻ obsahujĂcĂmi a-minoskupinu je komplex nekvartĂ©rnĂ ami-noskupiny se solĂ kovu stabilnÄjĆĄĂ neĆŸ kom-plex aminu amoniovĂ©ho typu se solĂ kovu,a ĆŸe pouĆŸitĂ soli zinku a slabĂ© kyseliny nor-mĂĄlnÄ nevede k vytvoĆenĂ pomÄrnÄ nesta-bilnĂho komplexu kovu obsahujĂcĂho aminamoniovĂ©ho typu. PouĆŸije-li se sĆŻl zinku asolnĂ© kyseliny, napĆĂklad chlorid zineÄnatĂœ,mĆŻĆŸe bĂœt takĂ© vytvoĆen ĆŸĂĄdanĂœ komplexzinku, je vĆĄak vĂœhodnĂ© pĆidat slabÄ alka-lickou sĆŻl, napĆĂklad octan sodnĂœ k solizinku pro neutralizaci prostĆedĂ reakce. Po-dobnÄ je ĆŸĂĄdoucĂ pĆidat urÄitĂ© mnoĆŸstvĂ oc-tanu sodnĂ©ho nebo hydroxidu sodnĂ©ho jakoneutralizaÄnĂho Äinidla, kdyĆŸ vĂœchozĂ ami-noglykosidovĂ© antibiotikum je pouĆŸito veformÄ jeho aditivnĂ soli se silnou kyselinou,napĆĂklad kyselinou chlorovodĂkovou. V tom-to pĆĂpadÄ je tĆeba zamezit pouĆŸitĂ neuĆŸi-teÄnĂ©ho pĆebytku neutralizaÄnĂho Äinidla,neboĆ„ jinak by doĆĄlo ke sraĆŸenĂ hydroxiduzineÄnatĂ©ho a tĂm naruĆĄenĂ vytvoĆenĂ kom-plexu. NapĆĂklad kdyĆŸ se pouĆŸije tetrahyd-rochlorid aminoglykoeidovĂ©ho antibiotikapro vytvoĆenĂ komplexu, pĆidajĂ se vĂœhod-nÄ 4 moly hydroxidu sodnĂ©ho pro neutrali-zaci reakÄnĂ smÄsi.
Pokud je celkovĂ© molĂĄrnĂ mnoĆŸstvĂ uĆŸitĂ©soli zinku alespoĆ rovno molĂĄrnĂmu mnoĆŸ-stvĂ aminoglykosidovĂ©ho antibiotika, reak-ce k vytvoĆenĂ komplexu mĆŻĆŸe probĂhat.NicmĂ©nÄ je vĂœhodnĂ© pouĆŸĂt soli zinku vmnoĆŸstvĂ podstatnÄ vyĆĄĆĄĂm neĆŸ 1 mol na 1mol aminoglykosidovĂ©ho antibiotika, takĆŸerovnovĂĄha reakce k vytvoĆenĂ komplexu jeposunuta ve prospÄch vytvĂĄĆenĂ komplexu. PĆĂznivĂœ vĂœtÄĆŸek komplexu zinku mĆŻĆŸe bĂœtzĂskĂĄn pĆi uĆŸitĂ soli zinku v mnoĆŸstvĂ 2,3aĆŸ 6 molĆŻ na 1 mol aminoglykosidu, v pra-xi je vĆĄak nejvhodnÄjĆĄĂ pouĆŸĂt soli zinku vmnoĆŸstvĂ 4 aĆŸ 5 molĆŻ na 1 mol aminoglyko-sidu. Äas potĆebnĂœ pro Ășplnou reakci provytvoĆenĂ komplexu po pĆidĂĄnĂ soli zinkuse mĆŻĆŸe mÄnit v zĂĄvislosti na pouĆŸitĂ©m or-ganickĂ©m rozpouĆĄtÄdle a mĆŻĆŸe bĂœt v roz- mezĂ âokamĆŸitĂœâ (pĆi pouĆŸitĂ vodnĂ©ho orga-nickĂ©ho rozpouĆĄtÄdla) aĆŸ do 20 hodin. Re-akce pro vytvoĆenĂ komplexu mĆŻĆŸe normĂĄl-nÄ probĂhat pĆi teplotÄ mĂstnosti, mĆŻĆŸe vĆĄakbĂœt provĂĄdÄno zahĆĂvĂĄnĂ nebo ochlazovĂĄnĂ.
Touto cestou se pĆipravĂ roztok nebo sus-penze obsahujĂcĂ komplex zinku a amino-glykosidovĂ©ho antibiotika, ke kterĂ©mu sepotom pĆidĂĄ acylaÄnĂ Äinidlo majĂcĂ acyl-skupinu pro zavedenĂ do komplexu jako·ochrannĂĄ aminoskupina.
AcylaÄnĂ Äinidlo pouĆŸitĂ© pĆi zpĆŻsobu po-dle pĆedloĆŸenĂ©ho vynĂĄlezu mĆŻĆŸe bĂœt ob-vyklĂ© Äinidlo pro ochranu aminoskupiny,a pouĆŸije se proto, aby volnĂ©, komplex ne-tvoĆĂcĂ aminoskupiny ve vĂœslednĂ©m kom-plexu aminoglykosidovĂ©ho antibiotika a ka-tiontu zinku byly acylovĂĄny a blokovĂĄny a-cylskupinou acylaÄnĂho Äinidla.
AcylaÄnĂ skupina mĆŻĆŸe bĂœt alka-noylovĂĄ skupina, aroylovĂĄ skupina, alko-xykarbonylovĂĄ skupina, aralalkoxykarbony-lovĂĄ skupina, aryloxykarbonylovĂĄ skupina,alkylsulfonylovĂĄ skupina, aralalkylsulfony-lovĂĄ skupina nebo arylsulfonyovĂĄ skupina,coĆŸ jsou vĆĄechno obvykĂ© skupiny pro och-ranu aminoskupiny.
AcylaÄnĂ Äinidlo pouĆŸitelnĂ© pro tento Ći-Äel mĆŻĆŸe takĂ© bĂœt karbonovĂĄ kyselina toho-to obecnĂ©ho vzorce (IVa): R5COOH (IVa) kde R5 je atom vodĂku, alkylskupina, zejmĂ©naalkylskupina s 1 aĆŸ 6 atomy uhlĂku, arylsku-pina, zejmĂ©na fenyl, nebo aralkylskupina,zejmĂ©na benzyl, a tyto skupiny jsou pĆĂpad-nÄ dĂĄle substituovĂĄny, nebo nÄkterĂœ halid,anbydrid nebo aktivnĂ ester zmĂnÄnĂ© karbo-novĂ© kyseliny vzorce (IVa), nebo chloro-formĂĄt tohoto obecnĂ©ho vzorce (IVb): R5O âCOâ-Cl (IVb) nebo p-nitrcfenylkarbonĂĄt tohoto obecnĂ©hovzorce (IVc): R5O_COâOâCGHv-p--NO., (IVc) nebo aktivnĂ N-hydroxysukcinimidester to-hoto vzorce (IVd):
O il o (IVd) nebo azidomravenÄan tohoto vzorce (IVe):R5OâCOâN9 (IVe) 261853 17 18 kde R' mĂĄ vĂœznam definovanĂœ vĂœĆĄe, nebo sul-fonovĂĄ. kyselina tohoto obecnĂ©ho vzorce(IVf):
Rhsn-ii (ivf) kde R,; jo a hnu vodĂku, alkylsknpina. zejmĂ©naalkyl skupina s i aĆŸ 6 atomy uhlĂku, aryl-skupion. zejmĂ©na fenyl, nebo aralkylskupi-na, zejmĂ©na fenylalkylskupina, napĆĂkladbenzyi, a tyto skupiny mohou bĂœt pĆĂpadnÄdĂĄle substituovĂĄny, nebo halogenid, anhyd-rid nebo aktivnĂ ester tĂ©to sulfonovĂ© kyse-liny. je tudĂĆŸ zĆejmĂ©, ĆŸe acylaÄnĂ reakcepro ochranu aminoskupin podle pĆedloĆŸe-nĂ©ho vynĂĄlezu je acylace v ĆĄirokĂ©m smys-lu, zahrnujĂcĂ napĆĂklad formylaci, acetyla-ci, propionylaci, trifluoracetylaci, benzyl-oxykarbonylaci, p-methoxybenzyloxykarbci-nylrci, t-butoxykarbonylaci, fenexykarbony-laci, tosylaci, mesylaci a jinĂ© ekvivalentnĂreakce.
ZvlĂĄĆĄtnĂ pĆĂklady pouĆŸitelnĂ©ho acylaÄnĂ-bo Äinidla jsou acetoxyformyl, p-nitrofenyl-formiĂĄt, anhydrid octovĂœ, acetylchlorid, an-hydrid propionovĂœ, p-nitrofenolester kyseli-ny trifluoroctovĂ©, ester kyseliny trifluoroc-tevĂ©, N-benzyloxykarbnnybmkcinjmid (vĂœ-znamnĂœ aktivnĂ ester j, N-benzyloxykarbo-nyloxyftalimid, benzyloxykarbonylchlorid,p-methoxybenzyloxykarbonyloxy-p-nitrofe-nyl, t-butoxykarbonylazid, fenoxykarbonyl-chlorid, tosylchlorid, mesylchlorid a jinĂ©,
AcylaÄnĂ Äinidlo, buÄ jako takovĂ©, nebojako roztok v rozpouĆĄtÄdle, napĆĂklad te-trahydrofuranu nebo dimothylsulfoxidu ne-bo jejich smÄsi, mĆŻĆŸe bĂœt pĆidĂĄno do roz-toku nebo suspenze, kterĂ© obsahujĂ kom-plex aminodykosidovĂ©ho antibiotika a zin-ku. M dĂĄrnĂ mnoĆŸstvĂ acylaÄnĂbo ÄinidlamĆŻĆŸe bĂœt obvykle rovno nebo s mĂrnĂœmpĆebytkem vyĆĄĆĄĂ neĆŸ jo poÄet nezkranplexo-vanĂœch aminoskupin, se kterĂœmi mĂĄ acylaÄ-nĂ Äinidlo reagovat. NicmĂ©nÄ v nÄkterĂœchpĆĂpadech mnoĆŸstvĂ pĆidanĂ©ho acylaÄnĂboÄinidla mĆŻĆŸe bĂœt od molĂĄrnĂho mnoĆŸstvĂ asitĆikrĂĄt vyĆĄĆĄĂ, neĆŸ je p ?Äet nezkcmplexova-nĂœch aminoskupin.
AcylaÄnĂ Äinidlo mĆŻĆŸe bĂœt pĆidĂĄno, buÄnajednou, nebo v dĂĄvkĂĄch bÄhem asi 2 aĆŸ3 hodin, aÄkoliv obvykle mĆŻĆŸe bĂœt pĆidĂĄnobÄhem 30 minut aĆŸ jednĂ© hodiny. AcylacemĂĄ bĂœt provĂĄdÄna pĆi teplotÄ od â20 °Cdo 100 °C, mĆŻĆŸe vĆĄak bĂœt normĂĄlnÄ provĂĄ-dÄna pĆi teplotÄ v rozmezĂ od 0 °C do tep-loty mĂstnosti. V nÄkterĂœch pĆĂpadech tep-lota reakce mĆŻĆŸe bĂœt udrĆŸovĂĄna nĂzkĂĄ vdobÄ pĆidĂĄnĂ acylaÄnĂho Äinidla a potom po-stupnÄ zvyĆĄovĂĄna jak probĂhĂĄ acylace. Nor-mĂĄlnÄ mĆŻĆŸe bĂœt acylaÄnĂ rekce provĂĄdÄnain sĂtu v organickĂ©m rozpouĆĄtÄdle, ve kte-rĂ©m byl vytvoĆen komplex aminoglykosido-vĂ©ho antibiotika se zinkovĂœm kationtem.
Tato acylace zinkovĂ©ho komplexu vytvoĆĂN-acylcvanĂœ zinkovĂœ komplex, to je kom-plex zinkovĂœch kationtĆŻ se selektivnÄ N-acy-lovanĂœm derivĂĄtem cminoglykosidovĂ©ho an-tibiotika.
Podle zpĆŻsobu podle prvnĂ myĆĄlenky pĆed-loĆŸenĂ©ho vynĂĄlezu acylace komplexu ami-noglykosidovĂ©ho antibiotika se zinkovĂœmkationtem je nĂĄsledovĂĄna krokem, pĆi kte-rĂ©m se odstranĂ zinkovĂœ kationt z N-acylo-vanĂ©ho zinkovĂ©ho komplexu, jmenovitÄ zin-kovĂœ komplex se rozloĆŸĂ, aby se zĂskal se-lektivnÄ chrĂĄnÄnĂœ N-acylovanĂœ derivĂĄt ami-noglykcsidovĂ©ho antibiotika, kterĂœ je pros-tĂœ zinkovĂœch kationtĆŻ.
Pro odstranÄnĂ zinkovĂ©ho kationtĆŻ z N--acylovanĂ©ho zinkovĂ©ho komplexu je nutnĂ©nechat reagovat N-acylovanĂœ zinkovĂœ kom-plex s vhodnĂœm Äinidlem, kterĂ© odstranĂ zin-kovĂœ kationt ze zmĂnÄnĂ©ho ' N-acvlovanĂ©hozinkovĂ©ho komplexu. Pro tento ĂșÄel jsou kpouĆŸitĂ mnohĂ© zpĆŻsoby. PrvnĂ zpĆŻsob spo-ÄĂvĂĄ v reakci Äinidla srĂĄĆŸejĂcĂho zinek, kte-rĂ© je schopnĂ© pĆemÄnit zinkovĂœ kationt naslouÄeninu zinku ve vodÄ rozpustnou, na-pĆĂklad na sirnĂk zineÄnatĂœ, hydroxid zineÄ-natĂœ nebo uhliÄitan zineÄnatĂœ, zatĂmco N--acvlovanĂœ zinkovĂœ komplex stĂĄle zĆŻstĂĄvĂĄrozpuĆĄtÄn ve smÄsi pro acylaÄnĂ reakce,kde byl acylovĂĄn komplex aminoglykosido-vĂ©ho antibiotika se zinkovĂœm kationtem,nebo po jeho pĆenesenĂ do novĂ©ho roztokuv ÄerstvĂ©m objemu organickĂ©ho, rozpouĆĄtÄd-la ze zmĂnÄnĂ© smÄsi pro acylaÄnĂ reakci. Äinidlo pro srĂĄĆŸenĂ zinku pouĆŸitelnĂ© vprvnĂm zpĆŻsobu mĆŻĆŸe bĂœt sirovodĂk, nÄkte-rĂœ sirnĂk alkalickĂ©ho kovu, sirnĂk amonnĂœ,sirnĂk nÄkterĂ© alkalickĂ© zeminy, napĆĂkladsirnĂk vĂĄpenatĂœ a uhliÄitan alkalickĂ©ho ko-vu, napĆĂklad uhliÄitan sodnĂœ, nebo takĂ©hydroxid amonnĂœ. V nÄkterĂœch pĆĂpadechmĆŻĆŸe bĂœt odstranÄnĂ zinkovĂœch kationtĆŻ zN-acylovanĂ©ho zinkovĂ©ho komplexu prove-deno pouze pĆidĂĄnĂm vody. Podle tohotoprvnĂho zpĆŻsobu pĆidĂĄnĂ Äinidla pro srĂĄĆŸe-nĂ zinku k roztoku N-acylovanĂ©ho zinkovĂ©-ho komplexu zpĆŻsobĂ pomÄrnÄ rychlĂ© sra-ĆŸenĂ nerozpustnĂ© slouÄeniny zinku vytvo-ĆenĂ© ze zinkovĂœch kationtĆŻ a sraĆŸenina mĆŻ-ĆŸe bĂœt odstranÄna filtracĂ. N-acylovanĂœ de-rivĂĄt aminoiglykosidovĂ©ho antibiotika, kte-rĂœ potom zĆŻstane v roztoku filtrĂĄtu mĆŻĆŸebĂœt zĂskĂĄn koncentracĂ roztoku nebo ex-trakcĂ z roztoku, a je-li tĆeba, mĆŻĆŸe bĂœt po-tom vyÄiĆĄtÄn. Pro vyÄistÄnĂ je napĆĂkladmoĆŸno pouĆŸĂt chromatografii ve sloupci sesilikagelem. DruhĂœ zpĆŻsob spoÄĂvĂĄ v tom,ĆŸe vĂœĆĄe zmĂnÄnĂĄ smÄs pro acylaÄnĂ reakcise (i) zkonceutruje nebo zkoncentruje dosucha odpaĆenĂm rozpouĆĄtÄdla nebo (ii) serozĆedĂ kapalnĂœm Ćedidlem, coĆŸ je takĂ© moĆŸ-no provĂ©st s novĂœm roztokem N-acylovanĂ©-ho zinkovĂ©ho komplexu pĆenesenĂ©ho doÄerstvĂ©ho objemu organickĂ©ho rozpouĆĄtÄdla,takĆŸe se zĂskĂĄ olejovitĂĄ nebo pevnĂĄ usaze-nina, koncentrĂĄt nebo zbytek, naÄeĆŸ se z 261853 19 tohoto nÄjakĂœm zpĆŻsobem zĂskĂĄ ĆŸĂĄdanĂœ N--acylovanĂœ derivĂĄt aminoglykosidovĂ©ho an-tibiotika. KapalnĂ© Ćedidlo pouĆŸitelnĂ© pĆitomto druhĂ©m zpĆŻsobu je voda nebo tako-vĂĄ organickĂĄ kapalina, ve kterĂ© buÄ N--acylovanĂœ zinkovĂœ komplex jako celek, ne-bo struktura N-acylovanĂ©ho derivĂĄtu ami-noglykosidovĂ©ho antibiotika zmĂnÄnĂ©ho N--acylovanĂ©ho zinkovĂ©ho komplexu nejevĂĆŸĂĄdnou nebo jevĂ jen malou rozpustnost.
Podle vĂœĆĄe uvedenĂ©ho druhĂ©ho zpĆŻsobuse pĆednÄ smÄs pro acylaÄnĂ reakci obsa-hujĂcĂ N-acylovanĂœ zinkovĂœ komplex (nebonovĂœ roztok N-acylovanĂ©ho zinkovĂ©ho kom-plexu pĆevedenĂœ do nÄjakĂ©ho organickĂ©horozpouĆĄtÄdla) zkoncentruje nebo zkoncen-truje do sucha k zĂskĂĄnĂ olejovitĂ© nebo pev-nĂ© usazeniny nebo zbytku. PouĆŸije-li se ja-ko prostĆedĂ pro reakci obtĂĆŸnÄ odpaĆitelnĂ©organickĂ© rozpouĆĄtÄdlo, napĆĂklad dime-thylsulfoxid atd. pro N-acylaci zinkovĂ©hokomplexu, je moĆŸnĂ©, ĆŸe reakÄnĂ smÄs proacylaci, obsahujĂcĂ N-acylovanĂœ zinkovĂœkomplex, se smĂchĂĄ s organickĂœm kapal-nĂœm Ćedidlem, napĆĂklad ethylĂ©terem, tak-ĆŸe tÄĆŸko odpaĆitelnĂ© organickĂ© rozpouĆĄtÄd-lo je rozpuĆĄtÄno v nebo zĆedÄno tĂmto Će-didlem, ÄĂmĆŸ se z nÄho usadĂ pevnĂĄ lĂĄtkanebo olej obsahujĂcĂ N-acylovanĂœ zinkovĂœkomplex. TĂmto zpĆŻsobem se zĂskĂĄ olejovi-tĂĄ nebo pevnĂĄ usazenina, coĆŸ je normĂĄlnÄsmÄs sestĂĄvajĂcĂ z (i) N-acylovanĂ©ho zin-kovĂ©ho komplexu, to je komplexu zinko-vĂœch kationtĆŻ s N-acylovanĂœm derivĂĄtem a-minoglykosidovĂ©ho antibiotika, (ii) z N-acy-lovanĂ©ho derivĂĄtu aminoglykosidovĂ©ho an-tibiotika zbavenĂ©ho rozloĆŸenĂm komplexo-tvornĂ©ho spojenĂ v ÄĂĄsti N-acylovanĂ©ho zin-kovĂ©ho komplexu vlivem podstatnĂ© nepĆĂ-tomnosti prostĆedĂ organickĂ©ho rozpouĆĄtÄd-la, (iii) z urÄitĂ©ho mnoĆŸstvĂ anorganickĂ© so-li zinku vytvoĆenĂ© rozleĆŸenĂm komplexo-tvornĂ©ho spojenĂ v ÄĂĄsti N-acylovanĂ©ho zin-kovĂ©ho komplexu, (iv) z urÄitĂ©ho mnoĆŸstvĂsoli zinku, kterĂĄ byla pĆidĂĄna na zaÄĂĄtkujako pĆebytek a zĆŻstala nezreagovĂĄna pĆireakci vytvĂĄĆenĂ komplexu, a moĆŸnĂĄ (v) zezbytkovĂ©ho mnoĆŸstvĂ organickĂ©ho rozpouĆĄ-tÄdla pouĆŸitĂ©ho v pĆedchozĂch operacĂch. VĂœĆĄe uvedenĂĄ olejovitĂĄ nebo pevnĂĄ usa-zenina nebo zbytek (vĂœĆĄe zmĂnÄnĂĄ smÄs)mĆŻĆŸe bĂœt potom zpracovĂĄna nÄkterĂœm z po-stupĆŻ (a), (b) a (c) uvedenĂœch dĂĄle. (a) OlejovitĂĄ nebo pevnĂĄ usazenina nebozbytek (vĂœĆĄe zmĂnÄnĂĄ smÄs) se smĂchĂĄ svodou nebo takovĂœm druhem polĂĄrnĂho or-ganickĂ©ho rozpouĆĄtÄdla, vodnĂ©ho polĂĄrnĂ-ho organickĂ©ho rozpouĆĄtÄdla nebo smÄsĂpolĂĄrnĂch organickĂœch rozpouĆĄtÄdel, kterĂ© jepolĂĄrnĂ organickou kapalinou pĆŻsobĂcĂ roz-loĆŸenĂ komplexotvornĂ©ho spojenĂ zinkovĂœchkationtĆŻ v N-acylovanĂ©m zinkovĂ©m kom-plexu pĆĂtomnĂ©m ve zmĂnÄnĂ© usazeninÄ ne-bo zbytku, a ve kterĂ© podĂly soli zinku u-volnÄnĂ© a zezaÄĂĄtku nezreagovanĂ© jsourozpustnĂ©, avĆĄak ve kterĂ© je ĆŸĂĄdanĂœ N-acy-lovanĂœ derivĂĄt aminoglykosidovĂ©ho antibio- 2Î tikĂĄ nerozpustnĂœ. TĂmto zpĆŻsobem se roz-loĆŸĂ N-acylovanĂœ zinkovĂœ komplex k uvol-nÄnĂ zinkovĂœch kaientĆŻ z nÄho, k umoĆŸnÄnĂrozpuĆĄtÄnĂ zinkovĂœch kationtĆŻ a jejich ex-trakci jako zinkovĂ© soli vodou nebo vodnĂœmorganickĂœm rozpouĆĄtÄdlem a k ponechĂĄnĂĆŸĂĄdanĂ©ho N-acylovanĂ©ho derivĂĄtu amino-glykosidovĂ©ho antibiotika ve formÄ neroz-pustnĂ©ho zbytku, kterĂœ mĂĄ bĂœt zĂskĂĄn. Ten-to zbytek mĆŻĆŸe bĂœt podle volby vyÄiĆĄtÄn o-pÄtnĂœm rozpuĆĄtÄnĂm v nÄkterĂ©m organic-kĂ©m rozpouĆĄtÄdle. PolĂĄrnĂ organickĂ© roz-pouĆĄtÄdlo pouĆŸitelnĂ© v tomto postupu (a)je napĆĂklad methanol, ethanol, kapalnĂœÄpavek, ethylamin a triethylamin. Tato po-lĂĄrnĂ organickĂĄ rozpouĆĄtÄdla a voda slou-ĆŸĂ jako Äinidlo k odejmutĂ zinkovĂ©ho ka-tiontu. (b) AlternativnÄ se olejovitĂĄ nebo pevnĂĄusazenina (vĂœĆĄe zmĂnÄnĂĄ smÄs) smĂchĂĄ stakovĂœm jinĂœm druhem polĂĄrnĂho' organic-kĂ©ho rozpouĆĄtÄdla, buÄ bezvodĂ©ho, nebovodnĂ©ho, kterĂ© rozklĂĄdĂĄ komplexotvornĂ©spojenĂ zinkovĂœch kationtĆŻ v N-acylovanĂ©mzinkovĂ©m komplexu pĆĂtomnĂ©m ve zmĂnÄnĂ©usazeninÄ nebo zbytku, a ve kterĂ©m uvolnÄ-nĂĄ sĆŻl zinku nenĂ rozpustnĂĄ, ale ĆŸĂĄdanĂœ N--acytovanĂœ derivĂĄt aminoglykosidovĂ©ho an-tibiotika je rozpustnĂœ, takĆŸe N-acylovanĂœzinkovĂœ komplex je rozloĆŸen k uvolnÄnĂN-acylovanĂ©ho derivĂĄtu aminoglykosidovĂ©-ho antibiotika z nÄho a k umoĆŸnÄnĂ jehorozpuĆĄtÄnĂ a extrakci zmĂnÄnĂœm polĂĄrnĂmorganickĂœm rozpouĆĄtÄdlem a tedy oddÄlenĂod soli zinku, kterĂĄ je uvolnÄna avĆĄak zĆŻs-tĂĄvĂĄ nerozpuĆĄtÄna ve zmĂnÄnĂ©m polĂĄrnĂmorganickĂ©m rozpouĆĄtÄdle. TĂmto zpĆŻsobemse zĂskĂĄ roztok ĆŸĂĄdanĂ©ho N-acylovanĂ©hoderivĂĄtu aminoglykosidovĂ©ho antibiotika vpolĂĄrnĂm organickĂ©m rozpouĆĄtÄdle a je-liĆŸĂĄdĂĄno, mĆŻĆŸe bĂœt vyÄiĆĄtÄn napĆĂklad chro-matograficky s nĂĄsledujĂcĂ koncentracĂ vy-ÄiĆĄtÄnĂ©ho roztoku pro oddÄlenĂ ĆŸĂĄdanĂ©hoN-acylovanĂ©ho produktu. (c) Podle dalĆĄĂ alternativy olejovitĂĄ ne-bo pevnĂĄ usazenina nebo zbytek (vĂœĆĄe zmĂ-nÄnĂĄ smÄs) zĂskanĂĄ ve vĂœĆĄe uvedenĂ©m dru-hĂ©m zpĆŻsobu mĆŻĆŸe bĂœt opÄt rozpuĆĄtÄna ja-ko celek ve vhodnĂ©m organickĂ©m rozpouĆĄ-tÄdle obsahujĂcĂm podĂl vody, kdyĆŸ je celĂĄusazenina nebo zbytek rozpustnĂĄ nebo pod-statnÄ rozpustnĂĄ ve vodÄ. Takto zĂskanĂœ roz-tok mĆŻĆŸe bĂœt potom podroben chromatogra-fickĂ©mu procesu, pĆi kterĂ©m mohou bĂœt u-volnÄnĂĄ sĆŻl zinku a uvolnÄnĂœ N-acylovanĂœderivĂĄt aminoglykosidovĂ©ho antibiotika zĂs-kĂĄny oddÄlenÄ z roztoku. PĆŻvodci pĆedlo-ĆŸenĂ©ho vynĂĄlezu zjistili, ĆŸe pro tento chro-matografickĂœ proces jsou uĆŸiteÄnĂ© rozliÄnĂ©druhy iontomÄnnĂœch pryskyĆic pro vĂœmÄnukationtĆŻ, vĂœmÄnu aniontĆŻ a vĂœmÄnu chelĂĄ-tĆŻ a ve vodÄ nerozpustnĂ© vysokĂ© polymeryobsahujĂcĂ funkÄnĂ skupiny schopnĂ© kombi-nace s kovem, napĆĂklad chitin nebo chito-san. VhodnĂ© stupnÄ pryskyĆice pro vĂœmÄ-nu kationtĆŻ pro tento ĂșÄel jsou ty, kterĂ© ob- 261853 21 22 sĂĄhujĂ karboxyskupiny ( âCOOHj jako vĂœ-mÄnnĂ© funkce, a ty, kterĂ© obsahujĂ sulfo-nylovĂ© skupiny (âSO:iHj jako vĂœmÄnnĂ©funkce. PouĆŸijĂ-li se pryskyĆice pro vĂœmÄ-nu kationtĆŻ obsahujĂcĂ karkoxylovĂ© funkcepro vĂœĆĄe uvedenĂœ chromatografickĂœ proces,vĂœĆĄe uvedenĂĄ olejovitĂĄ nebo pevnĂĄ usazeni-na nebo zbytek (vĂœĆĄe zmĂnÄnĂĄ smÄs] serozpustĂ ve vhodnĂ©m vodnĂ©m organickĂ©mrozpouĆĄtÄdle, napĆĂklad ve smÄsi vody amethanolu obsahujĂcĂ podle volby 10 % aĆŸ90 % objemovĂœch vody, nebo smÄsi vody adioxann obsahujĂcĂ podle volby 10 % aĆŸ 90proÄ. objemovĂœch vody, a vĂœslednĂœ roztokse zavede do sloupce zmĂnÄnĂ© pryskyĆicepro vĂœmÄnu kationtĆŻ. Sloupec se potom pro-myje dalĆĄĂm mnoĆŸstvĂm vĂœĆĄe uvedenĂ©hovodnĂ©ho organickĂ©ho rozpouĆĄtÄdla, naÄeĆŸnĂĄsleduje vyvolĂĄnĂ, pĆi kterĂ©m se jako e-luent pouĆŸije mnoĆŸstvĂ vĂœĆĄe uvedenĂ©ho vod-nĂ©ho organickĂ©ho rozpouĆĄtÄdla obsahujĂcĂ-ho dĂĄle jistĂ© mnoĆŸstvĂ kyseliny nebo zĂĄsa-dy. Jako kyselina se mĆŻĆŸe pouĆŸĂt slabĂĄ or-ganickĂĄ kyselina, napĆĂklad kyselina octo-vĂĄ, nebo zĆedÄnĂĄ anorganickĂĄ kyselina, na-pĆĂklad zĆedÄnĂĄ kyselina chlorovodĂkovĂĄ.
Jako zĂĄsada se mĆŻĆŸe pouĆŸĂt hydroxid a-inonnĂœ pro vÄtĆĄinu pĆĂpadĆŻ. Koncentrace ky-seliny nebo zĂĄsady ve vyvĂjecĂm rozpouĆĄ-tÄdle (eluentuj mĆŻĆŸe bĂœt s vĂœhodou 0,01 aĆŸ5 % hmot. vyvĂjecĂho roztoku. ĆœĂĄdanĂœ N--acylovanĂœ derivĂĄt aminoglykosidovĂ©ho an-tibiotika mĆŻĆŸe bĂœt oddÄlen od komplexotvor-nĂœch kationtĆŻ zinku bÄhem procesu vyvĂ-jenĂ, protoĆŸe pouĆŸitĂĄ pryskyĆice pro vĂœmÄnukationtĆŻ mĂĄ rozliÄnĂ© adsorpÄnĂ afinityvzhledem k ĆŸĂĄdanĂ©mu N-acylovanĂ©mu ami-noglykosidu a k zinkovĂœm kationtĆŻm, takĆŸesila prvnĂbo pro spojenĂ s pryskyĆicĂ jeodliĆĄnĂĄ od sĂly druhĂ©ho pro spojenĂ s prys-kyĆicĂ. TĂmto- zpĆŻsobem mĆŻĆŸe bĂœt eluĂĄtshromĂĄĆŸdÄn ve frakcĂch obsahujĂcĂch ĆŸĂĄ-danĂœ N-acylovanĂœ aminoglykosid prostĂœ so-li zinku, kterĂœ mĆŻĆŸe bĂœt potom zkoncentro-vĂĄn k zĂskĂĄnĂ ĆŸĂĄdanĂ©ho N-acylovanĂ©ho de-rivĂĄtu aminoglykosidovĂ©ho antibiotika.
PouzĂ je-li se pryskyĆice pro vĂœmÄnu ka-tiontĆŻ obsahujĂcĂ sulfonylovĂ© funkce provĂœĆĄe uvedenĂœ chromatografickĂœ proces, od-dÄlenĂ a vytÄĆŸenĂ ĆŸĂĄdanĂ©ho N-acylovanĂ©hoderivĂĄtu aminoglykosidovĂ©ho antibiotikamĆŻĆŸe bĂœt provedeno stejnĂœm zpĆŻsobem jakove vĂœĆĄe uvedenĂ©m pĆĂpadÄ, protoĆŸe k oddÄ-lenĂ N-acylovanĂ©ho aminoglykosidu cd kom-plexotvornĂœch zinkovĂœch kationtĆŻ je prove-den stejnĂœ mechanismus. Na druhĂ© stranÄ,pouĆŸije-11 se slabÄ nebo silnÄ zĂĄsaditĂĄ prys-kyĆice pro vĂœmÄnu aniontĆŻ pro chromatc-grafickĂœ proces, podĂl N-acylovanĂ©ho ami-noglykosidu v N-acylovanĂ©m zinkovĂ©m kom-plexu, kterĂœ obsahuje jednu nebo nÄkolikneacylovanĂœch aminoskupin, nenĂ normĂĄl-nÄ adsorbovĂĄn slabÄ nebo silnÄ zĂĄsaditoupryskyĆicĂ pro vĂœmÄru aniontĆŻ vlivem io-nĂckĂ©ho odpuzovĂĄnĂ mezi nimi, takĆŸe vyvĂ-jenĂ sloupce pryskyĆice pro vĂœmÄnu anion-tĆŻ vhodnĂœm vodnĂœm organickĂœm rozpouĆĄ- tÄdlem umoĆŸĆuje eluci N-acylovanĂ©ho deri-vĂĄtu aminoglykosidovĂ©ho antibiotika zesloupce, zatĂmco zinkovĂ© kationty zĆŻstanouve sloupci.
ProvĂĄdĂ-ii se chromatografickĂœ proces spouĆŸitĂm pryskyĆice pro vĂœmÄnu chelĂĄtĆŻ,kterĂĄ je schopnĂĄ kombinace se zinkovĂœmikationty, zavede se roztok vĂœĆĄe uvedenĂ© o-lejevitĂ© nebo pevnĂ© usazeniny nebo zbytku(vĂœĆĄe zmĂnÄnĂ© smÄsi) ve vodnĂ©m organic-kĂ©m rozpouĆĄtÄdle do sloupce pryskyĆice provĂœmÄnu chelĂĄtĆŻ, kterĂĄ se potom vyvinevhodnĂœm vyvĂjecĂm rozpouĆĄtÄdlem, aby ĆŸĂĄ-danĂœ N-ecylovanĂœ amino Tykosid byl eluo-vĂĄn pĆednostnÄ ze sloupce, zatĂmco zinko-vĂ© katio-nty zĆŻstanou vĂĄzĂĄny v pryskyĆicipro vĂœmÄnu chelĂĄtĆŻ. Ve vodÄ rozpustnĂœ vy-sokĂœ polymer obsahujĂcĂ funkce schopnĂ©kombinace s kovy, napĆĂklad chitin a chito-san, mĆŻĆŸe bĂœt pouĆŸit stejnĂœm zpĆŻsobem, ja-ko kdyĆŸ se pouĆŸije pryskyĆice pro vĂœmÄnuchelĂĄtĆŻ. (dj DĂĄle je moĆŸnĂœ tĆeli postup, pĆi kte-rĂ©m vĂœĆĄe uvedenĂĄ acylaÄnĂ reakÄnĂ smÄs,ve kterĂ© se provĂĄdÄla acylace zinkovĂ©hokomplexu pro ochranu aminoskupin, sepĆĂmo zavede do sloupce pryskyĆice pro vĂœ-mÄnu kationtĆŻ nebo vĂœmÄnu aniontĆŻ, che-lĂĄtĆŻ, nebo ve vodÄ nerozpustnĂ©ho vysokĂ©-ho polymeru majĂcĂho funkce kombinace skovy, takĆŸe N-acylovanĂœ zinkovĂœ komplexje adsorbovĂĄn pryskyĆicĂ nebo· vysokĂœm po-lymerem. Sloupec mĆŻĆŸe bĂœt potom promytvodnĂœm organickĂœm rozpouĆĄtÄdlem obsa-hujĂcĂm nebo neobsahujĂcĂm kyselinu nebozĂĄsadu, jak uvedeno v postupu (cj, naÄeĆŸse provedou podobnĂ© operace jako v po-stupu (c), ÄĂmĆŸ se dosĂĄhne odstranÄnĂ zin-kovĂœch kationtĆŻ z N-acylovanĂ©ho zinko-vĂ©ho komplexu, jakoĆŸ i vytÄĆŸenĂ ĆŸĂĄdanĂ©hoN-acylovanĂ©ho derivĂĄtu aminoglykosidovĂ©-ho antibiotika. (ej DĂĄle je moĆŸnĂœ ÄtvrtĂœ postup pro vy-tÄĆŸenĂ ĆŸĂĄdanĂ©ho N-acylovanĂ©ho derivĂĄtu a-minoglykosidovĂ©ho antibiotika, pĆi kterĂ©mse vĂœĆĄe uvedenĂĄ smÄs pro acylaÄnĂ reakciobsahujĂcĂ N-acylovanĂœ zinkovĂœ komplexzpracuje bezprostĆednÄ vodou pĆidĂĄnĂm vo-dy, v pĆĂpadÄ, ĆŸe ĆŸĂĄdanĂœ N-acylovanĂœ deri-vĂĄt aminoglykosidovĂ©ho antibiotika je ne-rozpustnĂœ ve vodÄ.
Jako pĆĂklad N-acylovanĂ©ho derivĂĄtu a-minoglykcsidovĂ©ho antibiotika, kterĂœ je vpodstatÄ nerozpustnĂœ ve vodÄ, mĆŻĆŸe bĂœtuveden 3,2â,6â-tri-N-benzyloxykarboinyldibe-kacin. V tomto pĆĂpadÄ, kdyĆŸ smÄs pro acy-laÄnĂ reakci obsahujĂcĂ N-acylovanĂœ zinko-vĂœ komplex obsahujĂcĂ N-acylovanĂœ derivĂĄtaminoglykosidu v podstatÄ nerozpustnĂœ vevodÄ, se bezprostĆednÄ smĂchĂĄ s vodou,rozruĆĄĂ se komplexotvornĂ© spojenĂ v N-a-cylovanĂ©m zinkovĂ©m komplexu a N-acylo-vanĂœ derivĂĄt aminoglykosidu se srazĂ jakopevnĂĄ lĂĄtka, zatĂmco sĆŻl zinku vytvoĆenĂĄ zuvolnÄnĂœch zinkovĂœch kationtĆŻ zĆŻstĂĄvĂĄ vroztoku, takĆŸe ĆŸĂĄdanĂœ N-acylovanĂœ derivĂĄt 281853 24 23 aminoglykosidovĂ©ho antibiotika jako v pod-statÄ ÄistĂœ produkt mĆŻĆŸe bĂœt vytÄĆŸen oddÄ-lenÄ od soli zinku.
Jak bylo uvedeno vĂœĆĄe, N-acylace, jme-novitÄ reakce k ochranÄ aminoskupin, seprovĂĄdĂ se zinkovĂœm komplexem aminogly-kosidovĂ©ho antibiotika v souhlase se zpĆŻso-bem podle prvnĂ myĆĄlenky pĆedloĆŸenĂ©ho vy-nĂĄlezu a komplex kattontĆŻ zinku s mono-,di-, tri- nebo poly-N-acylovanĂœm derivĂĄtemaminoglykosidu takto vytvoĆenĂœm je tako-vĂœ, ve kterĂ©m pouĆŸitĂ© zinkovĂ© kationty jsoukomplexovÄ sdruĆŸeny se strukturou N-acy-lovanĂ©ho derivĂĄtu aminoglykosidu. Je-li te-dy ĆŸĂĄdanĂœ N-acylovanĂœ derivĂĄt aminogly-kosidu nerozpustnĂœ nebo mĂĄlo rozpustnĂœ vevodÄ, zpĆŻsobĂ prostĂ© pĆidĂĄnĂ vody ke smÄsipro acylaÄnĂ reakci obsahujĂcĂ N-acylovanĂœzinkovĂœ komplex, ĆŸe ve vodÄ nerozpustnĂœN-acylovanĂœ derivĂĄt aminoglykosidu se vy-srĂĄĆŸĂ jako pevnĂĄ lĂĄtka, zatĂmco uvolnÄnĂ©zinkovĂ© kationty se ze smÄsi odstranĂ roz-puĆĄtÄnĂm ve vodÄ jako v pĆĂpadÄ ÄtvrtĂ©hopostupu popsanĂ©ho v pĆedchozĂm odstavci(ej. Takto zĂskanĂĄ sraĆŸenina, kterĂĄ je ne-rozpustnĂĄ ve vodÄ, mĆŻĆŸe bĂœt bezprostĆed-nÄ pouĆŸita jako vĂœchozĂ lĂĄtka pro nĂĄsledu-jĂcĂ reakce pro semisyntetickou pĆĂpravuĆŸĂĄdanĂ©ho koneÄnĂ©ho produktu. ObecnÄji Će-Äeno, i kdyĆŸ N-acylovanĂœ derivĂĄt aminogly-kosidovĂ©ho antibiotika je nÄkdy rozpustnĂœnebo ÄĂĄsteÄnÄ rozpustnĂœ ve vodÄ, a tudĂĆŸN-acylovanĂœ derivĂĄt aminoglykosidu mĆŻĆŸebĂœt vytÄĆŸen pouze pĆi podstatnÄ snĂĆŸenĂ©mvĂœtÄĆŸku, pouĆŸije-li se jednoduchĂœ postup spĆidĂĄnĂm vody bezprostĆednÄ do smÄsi proacylaÄnĂ reakci. Z tohoto dĆŻvodu mĆŻĆŸe bĂœtzĂskĂĄn lepĆĄĂ vĂœsledek, pouĆŸije-li se buÄ nÄ-kterĂœ z vĂœĆĄe uvedenĂœch postupĆŻ (bj a (cj,pĆi kterĂœch se N-acylovanĂœ komplex zinku,to je komplex zinkovĂœch kationtĆŻ s N-acy-lovanĂœm derivĂĄtem aminoglykosidovĂ©ho an-tibiotika vytvoĆenĂœ v N-acylaÄnĂ reakci, nej-prve oddÄlĂ od smÄsi pro acylaÄnĂ reakci,N-acylovanĂœ zinkovĂœ komplex takto oddÄle-nĂœ se potom rozpustĂ ve vodÄ nebo v nÄ-kterĂ©m vodnĂ©m organickĂ©m rozpouĆĄtÄdle avĂœslednĂœ roztok se dĂĄle zpracuje pro od-stranÄnĂ zinkovĂœch kationtĆŻ z nÄho. Jeden zjednoduchĂœch zpĆŻsobĆŻ odstranÄnĂ zinkovĂœchkationtĆŻ, kterĂœ je obecnÄ pouĆŸitelnĂœ, je ten,pĆi kterĂ©m se sirovodĂk nebo sirnĂk nÄkte-rĂ©ho alkalickĂ©ho kovu nechĂĄ reagovat ja-koĆŸto srĂĄĆŸecĂ Äinidlo se zinkovĂœmi kation-ty, aby se tyto vysrĂĄĆŸely jako sirnĂk zineÄ-natĂœ, coĆŸ je jeden zpĆŻsob prvnĂho postupupopsanĂ©ho vĂœĆĄe v odstavci (a). NicmĂ©nÄsirnĂk zineÄnatĂœ se nÄkdy vysrĂĄĆŸĂ jako ko-loidnĂ usazenina, kterĂĄ se velmi nesnadnofiltruje a kromÄ toho sirovodĂk i sirnĂkyalkalickĂœch kovĆŻ majĂ nepĆĂjemnĂœ zĂĄpacha nejsou vhodnĂ© pro pouĆŸitĂ pĆi komerÄnĂmvyuĆŸitĂ tohoto postupu. PĆŻvodci pĆedloĆŸe-nĂ©ho vynĂĄlezu tudĂĆŸ provedli rozsĂĄhlĂœ vĂœ-zkum za ĂșÄelem vytvoĆenĂ praktickĂ©ho· zpĆŻ-sobu k odstranÄnĂ zinkovĂœch kationtĆŻ zezinkovĂ©ho komplexu bez pouĆŸitĂ sirnĂkĆŻ, a podaĆilo se jim vyvinout ĂșÄinnĂœ a snadnĂœzpĆŻsob odstranÄnĂ zinkovĂœch kationtĆŻ pou-ĆŸitĂm vĂœĆĄe uvedenĂœch pryskyĆic pro vĂœmÄ-nu iontĆŻ nebo jinĂ©ho polymerovĂ©ho mate-riĂĄlu, jako v postupech (c) a (d) popsanĂœchvĂœĆĄe. Tyto postupy (cj a (dj jsou komerÄnÄvelmi vĂœhodnĂ© a hodnotnĂ©, neboĆ„ jsou snad-no proveditelnĂ©, dĂĄvajĂ vysokou ĂșÄinnostodstranÄnĂ zinkovĂœch kationtĆŻ a dĂĄvajĂ vy-sokĂœ vĂœtÄĆŸek ĆŸĂĄdanĂ©ho N-acylovanĂ©ho de-rivĂĄtu aminoglykosidovĂ©ho antibiotika.
Na zĂĄvÄr mohou bĂœt vĂœĆĄe popsanĂ© zpĆŻ-soby a postupy pro zpracovĂĄnĂ N-acylova-nĂ©ho zinkovĂ©ho· komplexu Äinidlem pro od-stranÄnĂ zinkovĂœch kationtĆŻ shrnuty takto: (i) Komplex zinkovĂœch kationtĆŻ se selek-tivnÄ N-acylovanĂœm derivĂĄtem aminoglyko-sidovĂ©ho antibiotika se oddÄlĂ ze smÄsi proacylaÄnĂ reakci pĆed tĂm, neĆŸ se nechĂĄ rea-govat s nÄkterĂœm Äinidlem pro odstranÄnĂzinkovĂœch kationtĆŻ z tohoto komplexu. (ii) Komplex zinkovĂœch kattontĆŻ se selek-tivnÄ N-acylovanĂœm derivĂĄtem aminoglyko-sidovĂ©ho· antibiotika se oddÄlĂ ze smÄsi pro·acylaÄnĂ reakci extrakcĂ nÄkterĂœm organic-kĂœm rozpouĆĄtÄdlem, odpaĆenĂm organickĂ©-ho rozpouĆĄtÄdla ze smÄsi pro acylaÄnĂ re-akci nebo zĆedÄnĂm smÄsi pro acylpÄnĂ re-akci Ćedidlem organickĂ©ho rozpouĆĄtÄdlapĆed tĂm, neĆŸ se nechĂĄ reagovat s Äinidlempro odstranÄnĂ zinkovĂœch kationtĆŻ.· i(iii) Komplex zinkovĂœch kationtĆŻ se se-lektivnÄ N-acylovanĂœm derivĂĄtem amino-glykosidovĂ©ho antibiotika jednou oddÄlenĂœse smĂchĂĄ s vodotu nebo· nÄkterĂœm polĂĄrnĂmorganickĂœm rozpouĆĄtÄdlem, buÄ bĂ©zvodĂœm,nebo vodnĂœm, kterĂ© slouĆŸĂ jako Äinidlo proodstranÄnĂ zinkovĂœch kationtĆŻ. Toto polĂĄr-nĂ organickĂ© rozpouĆĄtÄdlo je bud takovĂ©, vekterĂ©m je sĆŻl zinku rozpustnĂĄ, ve kterĂ©m jevĆĄak N-acylovanĂœ derivĂĄt aminoglykosido-vĂ©ho antibiotika nerozpustnĂœ, nebo takovĂ©,ve kterĂ©m je sĆŻl zinku nerozpustnĂĄ, ve kte-rĂ©m je vĆĄak N-acylovanĂœ derivĂĄt aminogly-kosidovĂ©ho antibiotika rozpustnĂœ. âąJĂv) Komplex zinkovĂœch kationtĆŻ s N-acy-lovanĂœm derivĂĄtem aminoglykosidovĂ©ho an-tibiotika jedrvou oddÄlenĂœ se opÄt rozpustĂĂșplnÄ v nÄkterĂ©m organickĂ©m rozpouĆĄtÄdleobsahujĂcĂm urÄitĂœ podĂl vody a vĂœslednĂœroztok se podrobĂ chromatografickĂ©mu pro-cesu s pouĆŸitĂm pryskyĆice pro vĂœmÄnu ka-tiontĆŻ, vĂœmÄnu aniontĆŻ, vĂœmÄnu chalĂĄtĆŻnebo se pouĆŸije ve vodÄ nerozpustnĂ©ho po-lymeru obsahujĂcĂho funkÄnĂ skupiny schop-nĂ© kombinace s kovem, slouĆŸĂcĂho jako Äi-nidlo· k odstranÄnĂ zinkovĂœch kationtĆŻ. (v) SmÄs pro acylaÄnĂ reakci se pĆĂmonechĂĄ projĂt sloupcem pryskyĆice pro vĂœ-mÄnu kationtĆŻ, vĂœmÄnu aniontĆŻ, vĂœmÄnuchelĂĄtĆŻ nebo ve vodÄ nerozpustnĂ©ho poly-meru obsahujĂcĂho funkce schopnĂ© kombi-nace s kovem pro adsorpci komplexu zinko-vĂœch kationtĆŻ s N-acylovanĂœm derivĂĄtem a-minoglykosidovĂ©ho antibiotika, a sloupec sepotom vyvine nÄkterĂœm vodnĂœm organickĂœm 261853 25 28 rozpouĆĄtÄdlem obsahujĂcĂm nebo neobsahu-jĂcĂm podĂl kyseliny nebo zĂĄsady, a eluĂĄtse shromĂĄĆŸdĂ ve frakcĂch, naÄeĆŸ se vytÄĆŸĂfrakce obsahujĂcĂ ĆŸĂĄdanĂœ selektivnÄ N-acy-lovanĂœ derivĂĄt amino dykosid aĂłIio antibio-tika avĆĄak neobsahujĂcĂ ĆŸĂĄdnĂ© zinkovĂ© ka-tionty. (vij Je-li ĆŸĂĄdanĂœ N-acylovanĂœ derivĂĄt a-mlnoglykosidovĂ©ho antibiotika nerozpustnĂœnebo v podstatÄ nerozpustnĂœ ve vodÄ, smÄspro acylaÄnĂ reakci se pĆĂmo smĂchĂĄ s vo-dou, takĆŸ;: zmĂnÄnĂœ derivĂĄt se srazĂ oddÄ-lenÄ cd soli. zinku, kterĂĄ zĆŻstane rozpuĆĄ-tÄna ve vodÄ. (vii) SmÄs pro acylaÄnĂ reakci se bez-prostĆednÄ nechĂĄ reagovat se sirovodĂkem,sirnĂkem alkalickĂ©ho kovu nebo sirnĂkemkovu alkalickĂ© zeminy, ÄĂmĆŸ se srazĂ zin-kovĂ© kalionly jako sirnĂk zineÄnatĂœ, nebos hydroxidem amonnĂœm, ÄĂmĆŸ se srazĂ zin-kovĂ© kationty jako· hydroxid zineÄnatĂœ. V zinkovĂ©m komplexu uvedenĂ©m ve zpĆŻ-sobu podle prvnĂ myĆĄlenky vynĂĄlezu jsouzinkovĂ© kaĂicnty principiĂĄlnÄ spojeny dokomplexu s 1-amino a 3â-aminoskupinamiaminoglykosidovĂ©ho antibiotika a tudĂĆŸ N--acylace komplexu aminoglykosidovĂ©ho an-tibiotika se zinkovĂœm kationtem nĂĄsledova-nĂĄ odstranÄnĂm zinkovĂœch kationtĆŻ z nÄhodĂĄvĂĄ normĂĄlnÄ N-acylovanĂœ derivĂĄt amino-glykosidovĂ©ho antibiotika, ve kterĂ©m ami-no u/nebo alkylaminoskupiny ji Ă© neĆŸ 1--aniino a 3â-aminoskupiuy jsou chrĂĄnÄnyacyl skupinou.
KdyĆŸ se takto zĂskanĂœ N-acylovanĂœ deri-vĂĄt aminoglykosidovĂ©ho antibiotika zpĆŻso-bem podle prvnĂ myĆĄlenky pĆedloĆŸenĂ©hovynĂĄlezu potem Î.-Î acyluje nÄkterou «-hyd-roxy-w-aminoalkanovcu kyselinou znĂĄmĂœmzpĆŻsobem popsanĂœm v patentovĂœch spisechSpojenĂœch stĂĄtĆŻ americkĂœch Ä. 3 781 268 aÄ. 3 939 143, naÄeĆŸ nĂĄsleduje odstranÄnĂzbĂœvajĂcĂch ochrannĂœch skupin chrĂĄnĂcĂchamineskupiny z vĂœslednĂ©ho 1-N-acylovanĂ©-ho produktu a zĂskĂĄ se semlsyntetickĂ© 1-N--acylovanĂ© aminuglykosidovĂ© antibiotikum,kterĂ© je znĂĄmĂ© jako uĆŸiteÄnĂ© antibakteriĂĄl-nĂ Äinidlo.
SyntĂ©za 1-N-acylov'anĂœch aminoglykosido-vĂœch antibiotik jo nynĂ popsĂĄna s odkazemna pĆĂkladnĂ© pouĆŸitĂ kanaraycinu A jakovĂœchozĂ lĂĄtky. PouĆŸije-li se kanamycin A ja-ko vĂœchozĂ lĂĄtka pĆi zpĆŻsobu podle prvnĂmyĆĄlenky vynĂĄlezu, zpoÄĂĄtku se blokujĂ 1--amino- a 3â-amlnoskuplny komplexacĂ sezinkovĂœmi kationty po vytvoĆenĂ jeho zin-kovĂ©ho komplexu. Je-li tedy komplex kanamycinu A se zinkovĂœm kationtem acylo-vĂĄn vhodnĂœm acylaÄnĂm Äinidlem podlepĆedloĆŸenĂ©ho vynĂĄlezu nebo· jinĂœm Äinid-lem pro blokovĂĄnĂ aminoskupin, nezkom-plexovanĂ© 3-amino- a 6â-aminoskupiny mo-lekuly kanamyemu A mohou bĂœt chrĂĄnÄnyacylskupinou pouĆŸitĂ©ho acylaÄnĂho Äinidlanebo jinĂœm druhem skupiny blokujĂcĂ ami-noskupltiu. Po nĂĄsledujĂcĂm odstranÄnĂ kom-plexotvornĂœch zinkovĂœch kationtĆŻ z komple- xu N-acylovanĂ©hO' kanamycinu A se zinko-vĂœm kationtem se vĂœslednĂœ N-acylovanĂœ de-rivĂĄt kanamycinu A nechĂĄ reagovat s acy-laÄnĂm Äinidlem majĂcĂm acylskupinu prozavedenĂ do 1-amin;skupiny molekuly ka-namycinu A. Tato· acylskuplna potom rea-guje pouze s neblokovanĂœmi 1-amino- a 3â--aminoskupinami kanamycinu A. V tĂ©to do-bÄ je 1-aminoskupina normĂĄlnÄ ponÄkud re-aktivnÄjĆĄĂ neĆŸ 3â-aminoskupina, takĆŸe po-ĆŸadovanĂœ 1-N-acylovanĂœ derivĂĄt kanamyci-nu A mĆŻĆŸe bĂœt zĂskĂĄn s ponÄkud vyĆĄĆĄĂmvĂœtÄĆŸkem neĆŸ 3â-N-acy] ovĂĄnĂœ derivĂĄt kana-mycinu A. NĂĄsledujĂcĂ zruĆĄenĂ N-ochrany1-N-acylovanĂ©ho derivĂĄtu kanamycinu Atakto zĂskanĂ©ho dĂĄvĂĄ 1-N-acylovanĂœ kana-mycin A jako ĆŸĂĄdanĂœ koneÄnĂœ produkt. Po-uĆŸije-li se tady zpĆŻsob podle prvnĂ myĆĄlen-ky pĆedloĆŸenĂ©ho vynĂĄlezu, je zĆejmĂ©, ĆŸe ĆŸĂĄ-danĂœ 1-N-acylkanamycin A mĆŻĆŸe bĂœt zĂs-kĂĄn pĆi vyĆĄĆĄĂm vĂœtÄĆŸku, ve srovnĂĄnĂ s pĆĂ-padem, kdy nechrĂĄnÄnĂœ kanamycin A nebo6â-N-chrĂĄnÄnĂœ kanamycin A je pĆĂmo uve-den do reakce s nÄkterĂœm acylaÄnĂm Äinid-lem za ĂșÄelem 1-N-acylace kanamycinu A.NechĂĄ-li se reagovat kanamycin bez jakĂ©-koli N-ochrany s nÄkterĂœm 1-N-acylaÄnĂmÄinidlem, zjistĂ se, ĆŸe se v tomto pĆĂpadÄvytvoĆĂ smĂĆĄenĂ© N-acylovanĂ© produkty ob-sahujĂcĂ velmi malĂœ podĂl, obvykle od 1 %do nÄkolika mĂĄlo % hmot. ĆŸĂĄdanĂ©ho 1-N-a-cyl .vanoho produktu.
PouĆŸije-li se zpĆŻsob podle prvnĂ myĆĄlenkypĆedloĆŸenĂ©ho vynĂĄlezu na kanamycin vĂœ-ĆĄe uvedenĂ©ho obecnĂ©ho vzorce (III), jsouchrĂĄnÄny nÄkterĂ© nebo vĆĄechny aminosku-piny jinĂ© neĆŸ 1-amino- a 3â-aminoskupinytohoto· pouĆŸitĂ©ho kanamycinu, coĆŸ dĂĄvĂĄ N--acylovanĂœ derivĂĄt kanamycinu odpovĂdajĂ-cĂ tomuto obecnĂ©mu vzorci (V):
kde
Rfa je hydroxyskupina, aminoskupina (âNHj), skupina âNHCOR5, nebo skupina âNHCO.OR5 nebo skupina âNHSO2R6, R< je hydroxyskupina, skupina âNHCOĆ5, sku- pina 261853 2? 28 R8 /
âN \ COR5 skupina â NHCO.OR5, skupinaR8 /
âN \ COâOR5 skupina âNHSO2Rfi nebo skupinaR8 /
âN \ SO2R° R3 a R:! majĂ vĂœznam definovanĂœ vĂœĆĄe vsouvislosti s obecnĂœm vzorcem (III), R7 jeskupina âCOR5, skupina â CO.OR5 neboskupina âSO2R6, R5 a RG majĂ vĂœznam defi-novanĂœ vĂœĆĄe v souvislosti se vzorci (IVa)aĆŸ (IVf), a R8 je alkylskupina, zejmĂ©na al-kylskupina s 1 aĆŸ 4 atomy uhlĂku. V pĆĂpadÄ, ĆŸe se zpĆŻsob podle prvnĂ myĆĄ-lenky pĆedloĆŸenĂ©ho vynĂĄlezu pouĆŸije na nÄ-kterĂœ kanamycin, zĂskĂĄ se obvykle N-chrĂĄ-nÄnĂœ derivĂĄt kanamycinu vzorce (V), vekterĂ©m vĆĄechny aminoskupiny jinĂ© neĆŸ ami-no- a/nebo alkylaminoekupiny pĆĂtomnĂ© vpolohĂĄch 1- a 3â-molekuly kanamycinu jsoublokovĂĄny.
KdyĆŸ vĆĄak acylskupina, kterĂĄ mĂĄ bĂœt za-vedena jako skupina blokujĂcĂ aminoskupl-nu, je pomÄrnÄ ĆĄirokĂĄ ve svĂ© sfĂ©rickĂ© veli-kosti, napĆĂklad t-butoxykarbonylskupina,nebo kdyĆŸ molĂĄrnĂ mnoĆŸstvĂ acylaÄnĂho Äi-nidla pouĆŸitĂ© v reakci je menĆĄĂ neĆŸ mnoĆŸ-stvĂ stechiometricky ĆŸĂĄdanĂ© pro acylacivĆĄech aminoskupin netvoĆĂcĂch komplex vmolekule kanamycinu, i kdyĆŸ acylskupinaacylaÄnĂho Äinidla je obvyklĂ© velikosti, ne-bo kdyĆŸ acylaÄnĂ reakce je zastavena na me-zistupni, zĂskĂĄ se takovĂœ N-chrĂĄnÄnĂœ deri-vĂĄt kanamycinu, ve kterĂ©m poÄet acylova-nĂœch aminoskupin v molekule kanamycinu je menĆĄĂ neĆŸ ve vĂœĆĄe uvedenĂ©m pĆĂpadÄ, apotom ve zvlĂĄĆĄtnĂch pĆĂpadech se zĂskĂĄ ta-kovĂœ omezenÄ N-acylovanĂœ derivĂĄt kanamy-cinu, ve kterĂ©m 6â-amino- nebo 6â-alkylami-noskupina je vĂœjimeÄnÄ acylovĂĄna, vlivemtoho, ĆŸe 6â-amino- nebo 6â-alkylaminosku-pina je reaktivnÄjĆĄĂ neĆŸ jinĂ© aminoskupinyv molekule kanamycinu. N-acylovanĂœ derivĂĄt kanamycinu obecnĂ©-ho vzorce (V) je vĂœznamnĂœ meziprodukt po-uĆŸitelnĂœ v semisyntetickĂ© pĆĂpravÄ rĆŻznĂœchdruhĆŻ derivĂĄtĆŻ kanamycinu. SlouÄeninavzorce (V) mĂĄ zvĂœĆĄenou hodnotu jako me-ziprodukt pro chemickou syntĂ©zu napĆĂkladkdyĆŸ se zavede do postupu k pĆĂpravÄ se-misyntetickĂĄ 1-N-acylovanĂĄ aminoglykosido-vĂĄ antibiotika aktivnĂ proti bakteriĂm odol-nĂœm vĆŻÄi kanamycinu ,acylaci 1-aminosku-piny slouÄeniny vzorce (V) nÄkterou a--hydroxy-w-aminoalkanoickou kyselinou anĂĄsledujĂcĂm odstranÄnĂm ochrannĂœch sku-pin z blokovanĂœch amino- a/nebo alkylami-noskupin vĂœslednĂ©ho 1-N-acylovanĂ©ho pro-duktu. MĂĄ-li napĆĂklad bĂœt pĆechodnĂĄ slouÄeni-na vzorce (V) acylovĂĄna nÄkterou acylsku-pinou, napĆĂklad (S)-4-benzyloxykarbonyl-amino-2-hydroxybutyrylskupinou, slouÄeni-na vzorce (5) mĆŻĆŸe bĂœt ve vhodnĂ©m roz-pouĆĄtÄdle, napĆĂklad vodnĂ©m tetrahydrofu-ranu, uvedena do reakce s pĆĂsluĆĄnÄ substi-tuovanou kyselinou mĂĄselnou nebo nÄkte-rĂœm jejĂm ekvivalentnĂm reaktivnĂm deri-vĂĄtem, napĆĂklad aktivnĂm esterem, napĆĂ-klad N-hydroxysukcinimidesterem. N-hydro-xyftalimidesterem nebo p-nitrofenolesterem,ÄĂmĆŸ se vytvoĆĂ produkt 1-N-acylace. NĂĄsle-dujĂcĂ odstranÄnĂ benzyloxykarbonylskupinya ochrannĂ© skupiny (R7) ve vzorci (V) zproduktu 1-N-acylace mĆŻĆŸe bĂœt provedenoobvyklou technikou zruĆĄenĂ N-ochrany, na-pĆĂklad buÄ hydrolĂœzou s kyselinou nebozĂĄsadou, nebo redukcĂ s redukÄnĂm kovem,nebo katalytickou hydrogenolĂœzou s vodĂ-kem, nebo redukcĂ radikĂĄlu sodĂkem v ka-palnĂ©m Äpavku, ÄĂmĆŸ se zĂskĂĄ semisyntetic-kĂœ derivĂĄt kanamycinu majĂcĂ (S)-4-amino--2-hydroxybutyrylskupinu vĂĄzanou na 1-a-minoskupinu kanamycinu a aktivnĂ proti o-dolnĂœm bakteriĂm, odpovĂdajĂcĂ tomuto o-becnĂ©mu vzorci (VI): " ·.
261853 29 30 kde R1, Râ, R:i a R'· majĂ vĂœznam definovanĂœvĂœĆĄe v souvislosti se vzorcem (III). Ve vĂœĆĄeuvedenĂ©m zpĆŻsobu mĆŻĆŸe bĂœt pouĆŸit obecnÄN-chrĂĄnÄnĂœ derivĂĄt nÄkterĂ© «-hydroxy-w-a-minoalkanoickĂ© kyseliny vzorce (VII): HOOCCH(CH2.)âNHa
OH (VII) kde n je rovno 1, 2 nebo 3, mĂsto (S) -4-ben zylnxykatâbonylainino-2-hydroxyinĂĄselnĂ© ky-seliny, ÄĂmĆŸ se zĂskĂĄ nÄkterĂœ derivĂĄt 1-N-··( (S)-w hydroxy-w-aminoalkynoylj-kana-mycinu.
ZpĆŻsob podle vynĂĄlezu umoĆŸĆuje pĆi vy-sokĂ©m vĂœtÄĆŸku pĆipravit 1-N-acylovanĂ© aini-nogiykosidovĂ© antibiotikum, kterĂ© je znĂĄmĂ©jako polosyntetlckĂœ antibakteriĂĄlnĂ prostĆe-dek. Tento vynĂĄlez tedy dĂĄle umoĆŸĆuje zpĆŻ-sob vĂœroby l-N-(a-hydroxy-w-aminoalka-noylj aminoglykosidovĂ©ho antibiotika, pĆikterĂ©m se vychĂĄzĂ ze znĂĄmĂ©ho aimnoglyko-sidovĂ©ho antibiotika, pĆiÄemĆŸ tento zpĆŻsobspoÄĂvĂĄ v tom, ĆŸe se nejprve vĂœĆĄe zmĂnÄnĂœmpostupem vyrobĂ kationtovĂœ komplex amino-glykosidovĂ©h-o antibiotika se zinkem, tedyÄĂĄsteÄnÄ chrĂĄnÄnĂœ N-acylovanĂœ derivĂĄt a-minoglykosidovĂ©ho antibiotika, ve kterĂ©m1-amino- a 3â-amino- nebo 3â-alkylamino-skupiny nejsou chrĂĄnÄny a vĆĄechny jinĂ© a-minoskupiny jsou chrĂĄnÄny, naÄeĆŸ se pĆi-pravĂ 1-N-nochrĂĄnÄnĂœ a jinak N-plnÄ chrĂĄ-nÄnĂœ derivĂĄt selektivnĂm 3â-acyIaÄnĂm po-stupem popsanĂœm v Äs. patentu Ä. 261 859a pohnu se 1-aminoskupina 1-N-nechrĂĄnÄ-nĂłbo a jinak N-plnÄ chrĂĄnÄnĂ©ho derivĂĄtuzĂskanĂ©ho v pĆedchozĂm stupni 3â-N-acy-lace, acyiuje a-hydroxy-fz>-aminoalkanovoukyselinou, zejmĂ©na 3-amino-2-hydroxypro-pionovou kyselinou (isoserin) nebo 4-ami-no-2-hydroxymĂĄsclnou kyselinou, a koneÄnÄse tak odstranĂ chrĂĄnĂcĂ skupiny z l-N-acy-levanĂ©ho produktu.
ZpĆŻsob vĂœroby l-N-(ty-hydroxy-<y-amino-alkynoyljovĂ©ho derivĂĄtu aminoglykosidovĂ©-ho antibiotika obsahujĂcĂho 6-0-(3â-amino-nebo 3â-alkylamino-3â-deoxyglykosyl)-2--deoxystreptaminovou ÄĂĄst, popĆĂpadÄ obsa-hujĂcĂ 4-O-aminoglykosylovou skupinu, seprovĂĄdĂ zpĆŻsobem, jehoĆŸ podstata je v tom,ĆŸe (aj zinkovĂ© kationty se nechajĂ reagovats aminoglykosidovĂœm antibiotikem v inert-nĂm organickĂ©m rozpouĆĄtÄdle k vytvoĆenĂkomplexu zinkovĂœch kationtĆŻ s aminoglyko-sidovĂœm antibiotikem, (bj s komplexem zinkovĂœch kationtĆŻ s aminoglykosidovĂœm antibiotikem, kterĂœ byl vytvoĆen ve vĂœĆĄe popsanĂ©m kroku (a), se nechĂĄ reagovat in ĆĄitu v inertnĂm organic- kĂ©m rozpouĆĄtÄdle acylaÄnĂ Äinidlo majĂcĂ acylskupinu k zavedenĂ jako skupinu proochranu amineskupin, k vytvoĆenĂ komple-xu zinkovĂœch kationtĆŻ se selektivnÄ N-acy-lovanĂœm derivĂĄtem aminoglykosidovĂ©ho an-tibiotika majĂcĂm pĆŻvodnÄ nezkomplexova-nĂ© aminoskupiny acylovĂĄny, (c) komplex selektivnÄ N-acylovanĂ©ho de-rivĂĄtu aminoglykosidovĂ©ho antibiotika sezinkovĂœm kaĂiontem zĂskanĂœ ve vĂœĆĄe popsa-nĂ©m kroku (bj se nechĂĄ reagovat s Äinid-lem, kterĂ© odstranĂ zinkovĂ© kationty z N--acylovanĂ©ho· zinkovĂ©ho komplexu, k vytvo-ĆenĂ ÄĂĄsteÄnÄ a selektivnÄ chrĂĄnÄnĂ©ho N--acylovanĂ©ho derivĂĄtu aminoglykosidovĂ©hoantibiotika, kterĂœ je prostĂœ zinkovĂœch ka-tiontĆŻ a ve kterĂ©m 1-amino- a 3â-amino- ne-bo 3â-alkylaminoskupiny jsou nechrĂĄnÄny,ale vĆĄechny jinĂ© aminoskupiny aminoglyko-sidovĂ© molekuly jsou chrĂĄnÄny acylskupi-nou, (dj ÄĂĄsteÄnÄ a selektivnÄ chrĂĄnÄnĂœ N-a-cylovanĂœ derivĂĄt zĂskanĂœ ve vĂœĆĄe popsanĂ©mkroku (cj se nechĂĄ reagovat s esterem nÄ-kterĂ© alkanoickĂ© kyseliny, kterĂœ odpovĂdĂĄvzorci (Vlil):
RaâCâRbII o (VIII) ve kterĂ©m
Ra je atom vodĂku nebo dihaloalkyl nebo·trihaloalkylskupina s 1 aĆŸ 6 atomy uhlĂku aRb je alkyloxyskupina s 1 aĆŸ 6 atomy uhlĂ-ku, zejmĂ©na benzyloxyskupina nebo nÄkterĂĄaryloxyskupina, zejmĂ©na fenoxyskupina, ne-bo N-formylimidazol, jakoĆŸto acylaÄnĂm Äi-nidlem, v inertnĂm organickĂ©m rozpouĆĄtÄd-le k selektivnĂ acylaci 3â-amino- nebo 3â--alkylaminoskupiny acylskupinou RaCOâzmĂnÄnĂ©ho acylaÄnĂho Äinidla k vytvoĆenĂ1-N-nechrĂĄnÄnĂ©ho a jinĂ©ho N-plnÄ chrĂĄnÄ-nĂ©ho derivĂĄtu aminoglykosidovĂ©ho' antibio-tika, ve kterĂ©m vĆĄechny aminoskupiny jinĂ©neĆŸ 1-aminoskupina jsou chrĂĄnÄny acylsku-pinou, (e) 1-N-nechrĂĄnÄnĂœ a jinĂœ N-chrĂĄnÄnĂœderivĂĄt zĂskanĂœ ve vĂœĆĄe popsanĂ©m kroku(dj se nechĂĄ reagovat s nÄkterou a-hydro-xy-w-aminoalkanovou kyselinou vzorce (IX j: HOOCâCH(CH2jinNH2
OH (IX) kde m je rovno 1 nebo 2, nebo s ekvivalent-nĂm reaktivnĂm derivĂĄtem tĂ©to kyseliny, je-hoĆŸ aminoskupina je bud1 nechrĂĄnÄna, nebochrĂĄnÄna, k acylaci 1-aniinoskupiny zmĂnÄ-nĂ©ho 1-N-nechrĂĄnÄnĂ©ho derivĂĄtu, (f) naÄeĆŸ se odstranĂ zbĂœvajĂcĂ skupiny 261853 31 32 chrĂĄnĂcĂ aminoskupiny z produktu 1-N-acy-lace zĂskanĂ©ho ve vĂœĆĄe popsanĂ©m kroku(ej obvyklĂœm zpĆŻsobem pro zruĆĄenĂ ochra-ny. DĂĄle bude podrobnÄji popsĂĄno provĂĄdÄ-nĂ zpĆŻsobu podle tĆetĂ myĆĄlenky pĆedloĆŸe-nĂ©ho vynĂĄlezu.
AminoglykosidovĂĄ antibiotika, kterĂĄ jsouvhodnĂĄ jako vĂœchozĂ lĂĄtka pro prvnĂ krok(a) tohoto zpĆŻsobu jsou stejnĂĄ jako anti-biotika popsanĂĄ vĂœĆĄe pĆi zpĆŻsobu podle prv-nĂ myĆĄlenky vynĂĄlezu. Reakce komplexo-tvornĂœch zinkovĂœch kationtĆŻ s aminoglyko-sidovĂœm antibiotikem se provĂĄdĂ stejnĂœmzpĆŻsobem, jak bylo popsĂĄno vĂœĆĄe. Acylacekomplexu aminoglykosidovĂ©ho antibiotikase zinkovĂœm kationtem zĂskanĂ©ho v prvnĂmkroku (aj mĆŻĆŸe bĂœt ve druhĂ©m kroku (bjprovedena stejnĂœm zpĆŻsobem, jak bylo po-psĂĄno vĂœĆĄe u zpĆŻsobu podle prvnĂ myĆĄlen-ky vynĂĄlezu. OdstranÄnĂ zinkovĂœch kation-tĆŻ ze selektivnÄ N-acylovanĂ©ho komplexuaminoglykosidovĂ©ho antibiotika a zinkovĂ©-ho kationtĆŻ takto zĂskanĂ©ho mĆŻĆŸe bĂœt vetĆetĂm kroku (c) tohoto zpĆŻsobu provedenorozliÄnĂœmi postupy, jak bylo popsĂĄno vĂœĆĄe,ÄĂmĆŸ se zĂskĂĄ ÄĂĄsteÄnÄ a selektivnÄ chrĂĄnÄ-nĂœ N-acylovanĂœ derivĂĄt aminoglykosidovĂ©hoantibiotika, kterĂœ je prostĂœ zinkovĂœch ka-tiontĆŻ, a ve kterĂ©m 1-amino- a 3â-amino-nebo 3â-alkylaminoskupiny jsou nechrĂĄnÄ-ny, avĆĄak vĆĄechny jinĂ© aminoskupiny v mo-lekule aminoglykosidu jsou blokovĂĄny acyl-skupinou acylaÄnĂho Äinidla pouĆŸitĂ©ho vkroku (bj tohoto zpĆŻsobu. Tento ÄĂĄsteÄnÄa selektivnÄ chrĂĄnÄnĂœ N-acylovanĂœ derivĂĄtaminoglykosidovĂ©ho antibiotika se potomnechĂĄ reagovat v kroku (dj tohoto zpĆŻsobus esterem nÄkterĂ© alkanoickĂ© kyseliny vzor-ce (VIII j nebo s N-formimidazolem stej-nĂœm zpĆŻsobem jak bylo popsĂĄno vĂœĆĄe pĆizpĆŻsobu podle druhĂ© myĆĄlenky vynĂĄlezu kzĂskĂĄnĂ selektivnĂ 3â-N-acylace ÄĂĄsteÄnÄ N--chrĂĄnÄnĂ©ho derivĂĄtu aminoglykosidovĂ©hoantibiotika bez acylace jeho 1-aminoskupi-ny. V pĂĄtĂ©m kroku (ej tohoto zpĆŻsobu 1-N--nechrĂĄnÄnĂœ a jinĂœ N-plnÄ chrĂĄnÄnĂœ deri-vĂĄt aminoglykosidovĂ©ho antibiotika zĂska-nĂœ v pĆedeĆĄlĂ©m kroku (dj tohoto zpĆŻsobuse nechĂĄ reagovat s nÄkterou a-hydroxy-Ï--aminoalkanoickou kyselinou vzorce (X),zejmĂ©na s 3-amino-2-hydroxypropionovoukyselinou (DL-isoserin, D-isoserin nebo L--isoserinj, nebo s L-4-amino-2-hydroxymĂĄ-selnou kyselinou k acylaci 1-aminoskupinyaminoglykosidovĂ©ho antibiotika 3-amino-2--hydroxypropionyl- nebo 4-amino-2-hydroxy-butyrylskupinou.
Tato 1-N-acylace mĆŻĆŸe bĂœt provĂĄdÄna o-becnÄ zpĆŻsobem popsanĂœm v britskĂ©m pa-tentovĂ©m spise Ä. 1 426 908 nebo v patento-vĂ©m spise Sp. st. am. Ä. 4 001 208 podle kte-rĂ©hokoli znĂĄmĂ©ho zpĆŻsobu syntĂ©zy amidĆŻreakcĂ chrĂĄnÄnĂ©ho derivĂĄtu aminoglykosi-dovĂ©ho antibiotika s nÄkterĂœm isoserinemnebo s L-4-amino-2-hydroxymĂĄselnou kyseli- nou, buÄ v jejĂ volnĂ© formÄ kyseliny, nebove formÄ jejĂho. reaktivnĂho ekvivalentu, na-pĆĂklad aktivnĂho esteru, napĆĂklad dicyk-lohexylkarbodimidesteru, smĂĆĄenĂ©ho anhyd-ridu kyseliny, azidu kyseliny, v nÄkterĂ©m i-nertnĂm organickĂ©m rozpouĆĄtÄdle, napĆĂ-klad dioxanu, dimethoxyethanu, dimethyl-formamidu, tetrahydrofuranu nebo vodnĂ©formÄ tÄchto rozpouĆĄtÄdel. Isoserin a L-4--amino-2-hydroxymĂĄselnĂĄ kyselina mohoubĂœt ty lĂĄtky, jejichĆŸ aminoskupiny byly blo-kovĂĄny skupinou pro ochranu aminoskupin.VhodnĂ© skupiny pro ochranu aminoskupinpro tento ĂșÄel mohou bĂœt stejnĂ© nebo od-liĆĄnĂ© od tÄch, kterĂ© byly pouĆŸity v 1-N--nechrĂĄnÄnĂ©m, ale jinĂ©m N-chrĂĄnÄnĂ©m de-rivĂĄtu aminoglykosidovĂ©ho antibiotika, kte-rĂœ mĂĄ bĂœt 1-N-acylovĂĄn. VĂœhodnĂĄ skupinapro ochranu aminoskupin je t-butoxykarbo-nylskupina, neboĆ„ je pĆĂmo odstranitelnĂĄ re-akcĂ se zĆedÄnou kyselinou, napĆĂklad s vod-nou kyselinou trifluoroctovou, vodnou kyse-linou octovou a zĆedÄnou kyselinou chlo-rovodĂkovou.
Jako skupiny pro ochranu aminoskupinyjsou dĂĄle vhodnĂ© benzyloxykarbonylskupi-na, kterĂĄ se odstranĂ obvyklou hydrogeno-lĂœzou na palladiovĂ©m nebo platinovĂ©m oxi-dovĂ©m katalyzĂĄtoru, jakoĆŸ i ftaloylskupina,kterĂĄ se snadno odstranĂ hydrolĂœzou s hyd-razĂnem.
AcylaÄnĂ reakce kroku 1-N-acylace (ejpĆi zpĆŻsobu podle ÄtvrtĂ© myĆĄlenky pĆedlo-ĆŸenĂ©ho vynĂĄlezu mĆŻĆŸe bĂœt vĂœhodnÄ provĂĄ-dÄna v nÄkterĂ©m vodnĂ©m organickĂ©m roz-pouĆĄtÄdle s pouĆŸitĂm nÄkterĂ©ho aktivnĂhoesteru «-hydroxy-w-aminoalkanoickĂ© kyseli-ny vzorce (Xj. VhodnĂœ aktivnĂ ester mĆŻĆŸebĂœt N-hydroxysukcinimidester isoserinu ne-bo L-4-benzyloxykarbonylamino-2-hydroxy-mĂĄselnĂĄ kyselina, a tento aktivnĂ ester mĆŻ-ĆŸe bĂœt pouĆŸit v mnoĆŸstvĂ od 1 do 2 molĆŻ,vĂœhodnÄ cd 1 do 1,5 molu na 1 mol ami-noglykosidu, kterĂœ mĂĄ bĂœt 1-N-acylovĂĄn. Svodou mĂsitelnĂ© organickĂ© rozpouĆĄtÄdlo po-uĆŸitĂ© v reakÄnĂm prostĆedĂ mĆŻĆŸe bĂœt s vĂœ-hodou dioxan, dimethoxyethan, dimethyl-formamid nebo tetrahydrofuran.
Po kroku (ej se provede krok (f) toho-to zpĆŻsobu k odstranÄnĂ ochrany, to je kodstranÄnĂ vĆĄech zbĂœvajĂcĂch skupin chrĂĄ-nĂcĂch aminoskupiny z produktu 1-N-acyla-ce zĂskanĂ©ho v pĆedeĆĄlĂ©m kroku (ej tohotozpĆŻsobu. OdstranÄnĂ zbĂœvajĂcĂch skupin proochranu aminoskupin se mĆŻĆŸe provĂ©st ob-vyklou technikou. TakovĂĄ zbĂœvajĂcĂ skupi-na chrĂĄnĂcĂ aminoskupiny, kterĂĄ je alkoxy-karbonylovĂ©ho typu, mĆŻĆŸe bĂœt odstranÄnahydrolĂœzou s vodnĂœm roztokem kyseliny tri-fluoroctovĂ© nebo. kyseliny octovĂ© nebo zĆe-dÄnĂœm roztokem kyseliny, napĆĂklad zĆe-dÄnou kyselinou chlorovodĂkovou. TakovĂĄzbĂœvajĂcĂ skupina pro ochranu aminoekupl-ny, kterĂĄ je aralkylkarbonylovĂ©ho typu, na-pĆĂklad benzyloxykarbonyl, se odstranĂ pĆĂ-mo katalytickou hydrogenolĂœzou. KdyĆŸ se 261853 33 34 odstranĂ vĆĄechny zbĂœvajĂcĂ skupiny pro och-ranu aminoskupin z produktu 1-N-acylace zkroku (e] tohoto zpĆŻsobu, zĂskĂĄ se s vy-sokĂœm vĂœtÄĆŸkem ĆŸĂĄdanĂ© l-N-(2-hydroxy-3-aminoprcpionyl j- nebo l-N-(2-hydroxy-4-a-minobutyryl) aminoglykosidovĂ© antibioti-kum. PĆĂklady 1-N- {a-hydroxy-w-aminoalkanc-ylj-aminoglykosidovĂ©ho antibiotika pĆipra-venĂ©ho zpĆŻsobem podle ÄtvrtĂ© myĆĄlenkypĆedloĆŸenĂ©ho vynĂĄlezu jsou uvedeny dĂĄle.
(1 ] 1-N- (L-4-amino-2-hydroxybutyryl) --kanamycin A
(2) 1-N- (L-4-amino-2-hydroxybutyryl) --3â-deoxykanamycin A
(3 ] 1-N- (L-4-amino-2-hydroxybutyryl) --3â,4â-dideoxykanamycin A (4) 1-N- (L-4-amino-2-hydroxybutyryl) --tobramycin (5 j 1-N- (L-4-amino-2-hydroxybutyryl j -dibekacin (6) l-N-(3-amino-2-hydroxypropionylj--dibekacin jinĂ© pouĆŸitĂ zpĆŻsobĆŻ podle prvnĂ a druhĂ©myĆĄlenky pĆedloĆŸenĂ©ho vynĂĄlezu spoÄĂvĂĄ vpĆĂpravÄ l-N-alkylaminoglykosidovĂ©ho an-tibiotika ze vĆĄech N-acylovanĂœch aminogly-kosidovĂœch derivĂĄtĆŻ obsahujĂcĂch nechrĂĄ-nÄnou 1-aminoskupinu, a pĆĂkladem tohotopouĆŸitĂ mĆŻĆŸe bĂœt pĆĂprava netilmycinu ne-bo jeho 1-N-alkylanalogĆŻ ze sisomycinu ai-kylacĂ s niĆŸĆĄĂm alifatickĂœm aldehydem akyanborohydridem.
VynĂĄlez bude dĂĄle osvÄtlen avĆĄak niko-liv omezen tÄmito pĆĂklady. PĆĂklad 1
PĆĂprava 3,6â-di-N-benzyloxykarbonylkana-mycinu A (i) 2,0 g, 4,13 mM kanamycinu A jako vol-nĂ© bĂĄze bylo suspendovĂĄno ve smÄsi 50 mldimetylsulfoxidu a 20 ml tetrahydrofuranua k suspenzi bylo pĆidĂĄno 4 g, 18,1 mM oc-tanu zineÄnatĂ©ho dihydrĂĄtu, naÄeĆŸ byla re-akÄnĂ smÄs protrepĂĄvĂĄna pĆi teplotÄ mĂst-nosti aĆŸ do vytvoĆenĂ homogennĂho rozto-ku. VytvoĆenĂ a rozpuĆĄtÄnĂ zineÄnatĂ©ho kom-plexu kanamycinu A trvalo asi 4â5 hodin.VĂœslednĂœ roztok byl pak ochlazen na 0 °Ca k nÄmu byl pak pĆidĂĄvĂĄn po jednu hodi-nu 0 °C chladnĂœ roztok 2,37 g, 9,5 mM N--benzyloxykarbonyloxysukcinimidu,
O c&H5CH&OCOO~N/] rozpuĆĄtÄnĂ©ho ve smÄsi 40 ml tetrahydrofu-ran-dimethylsulfoxid 1 : 1 obj. ReakÄnĂ roz-tok byl ponechĂĄn stĂĄt pĆi teplotÄ okolĂ po4 hodiny, bÄhem nichĆŸ byl zineÄnatĂœ kom-plex kanamycinu A benzyloxykarbonylovĂĄn,coĆŸ pĆedstavuje acylaci podle prvnĂho hle-diska vynĂĄlezu.
Vzorek odebranĂœ z takto zĂskanĂ© reakÄnĂsmÄsi byl chromatografovĂĄn na tenkĂ© vrst-vÄ silikagelu, pĆiÄemĆŸ jako vyvolĂĄvacĂhoroztoku bylo, pouĆŸito spodnĂ kapalnĂ© fĂĄzesmÄsi chloroform â metanol â 28% vodnĂœÄpavek v obj. pomÄru 1 : 1 : 1, a pĆi chro-matografli dĂĄval hlavnĂ skvrnu ĆŸĂĄdanĂ©hoproduktu pĆi R( = 0,23 a dvÄ nebo tĆi men-ĆĄĂ skvrny, nĂĄleĆŸejĂcĂ vedlejĆĄĂm produktĆŻm,ve vyĆĄĆĄĂch bodech. (ii) VĂœĆĄe uvedenĂœ reakÄnĂ roztok byl vlitdo 500 ml etylĂ©teru a oddÄlenĂœ olej byl nÄ-kolikrĂĄt promyt dalĆĄĂmi objemy etylĂ©teru,ÄĂmĆŸ se zĂskalo 8,8 g hustĂ© sirupovitĂ© lĂĄt-ky. (iii] OdstranÄnĂ kationu zinku ze sirupo-vitĂ© lĂĄtky,, obsahujĂcĂ pĆevĂĄĆŸnÄ zineÄnatĂœkomplex, bylo provedeno nÄkterĂœm z tÄch-to rĆŻznĂœch postupĆŻ: (A) Postup pouĆŸĂvajĂcĂ slabÄ kyselou ka-texovou iontomÄniÄovu pryskyĆici nesoucĂkarboxylovou skupinu âCOOH jako funkÄnĂskupinu, komerÄnÄ dostupnou pod nĂĄzvemâAmberliteâ CG 50 pryskyĆice (HH forma]od. Rohm and Haas Co., (Sp. st. a.]. 60 ml pryskyĆice Amber lit CG 50 H+ for-my bylo- pĆedem dĆŻkladnÄ nasycenou smÄ-sĂ voda â dioxan (2 : 1) a pak naplnÄno,do kolony. Roztok 1 g sirupovitĂ© lĂĄtky roz-puĆĄtÄnĂ© v 20 ml smÄsi voda â dioxan (1 :: 1) byl ponechĂĄn projĂt kolonou, kterĂĄ by-la pak vyvolĂĄna smÄsĂ voda â dioxan (2 : : 1] obsahujĂcĂ 1% kyselinu octovou. EluĂĄtbyl pak sbĂrĂĄn ve frakcĂch. ĆœĂĄdanĂœ 3,6â-di--N-benzyloxykarbonylkanamycin A, kterĂœ bylpozitivnĂ v niuhydrinovĂ© reakci byl eluo-vĂĄn jako prvnĂ z kolony a octan zineÄnatĂœ,kterĂœ byl senzitivnĂ na zbarvenĂ difenyl-karbazidern, byl eluovĂĄn aĆŸ po nÄm. FrakceobsahujĂcĂ ĆŸĂĄdanĂœ produkt byly spojeny azakoncentrovĂĄny do, sucha. Zbytek byl pakpromyt etylĂ©terem, ÄĂmĆŸ se vytÄĆŸilo 340 mgtj. 81 % 3,6â-di-N-benzylGxvkarbonylkana-mycinu A ve formÄ bezbarvĂ© pevnĂ© lĂĄtky.
[ajn25 +76° (c 1, voda â dimetylformamid, 1 : 2).
ElementĂĄrnĂ analĂœza
VypoÄteno pro C-y,HwN4O15 . 2 CH-tCO-,HâH-,O: 51,23 % C, 6,56 % H, 6,29 % N,
Nalezeno: 51,02 % C, 6,71 % H, 6,22 % N. (B) ZpĆŻsob pouĆŸĂvajĂcĂ slabou katexovou 33
IontomÄniÄovou pryskyĆici nesoucĂ karbo-xylanovou skupinu jako funkÄnĂ skupinu,obchodnÄ dostupnou jako ,,Amberiiteâ CG50 pryskyĆice, ĆH// forma od Rohm andHaas Co. 1 g sirupovitĂ© lĂĄtky zĂskanĂ© vĂœĆĄe v pĆĂ-kladu 1 (ii) byl rozpuĆĄtÄn vc 20 ml smÄsivoda â dioxan (1 : 1) a roztok byl nechĂĄnprojĂt kolonou 60 ml pryskyĆice AmberlitCG 50, NIL·/ forma a byl eluovĂĄn lineĂĄr-nĂm gradientem smÄsi voda â dioxan 1. : 1obsahujĂcĂ aĆŸ 0,1 N Äpavek. Nebyl eluovĂĄnĆŸĂĄdnĂœ kation zinku ale pouze ĆŸĂĄdanĂœ pro-dukt, 3,6â-di-N-benzyloxykarbonylkanauiy-cin A. Frakce eluĂĄiu obsahujĂcĂ ĆŸĂĄdanĂ© ben-zyloxykarbouylovanĂ© produkty byly koncentrovĂĄny do sucha pĆiÄemĆŸ bylo zĂskĂĄno323 g, tj. 89 % ĆŸĂĄdanĂ©ho- produktu ve formÄbezbarvĂ© pevnĂ© lĂĄtky.
[ajir5 =-- +86° (c 1, voda - dinietylforma-mld, 1:2).
ElementĂĄrnĂ analĂœza
VypoÄteno pro C-wH^Nz.Ur,. W 1-1-,0(.).: 52,87 % C, 6,30 % II, 7,15 % N,
Nalezeno: 52,50 % C, 6,50 % H, 7,00 % N. (C) ZpĆŻsob vyuĆŸĂvajĂcĂ katexovou ionto-mÄniÄovou pryskyĆici nesoucĂ silnÄ kyseloufunkÄnĂ skupinu âSO-.,Î, komerÄnÄ dostup-nou jako- âDovexâ 50 W X 2 nd Dox Chemi-cal Co. 30 ml pryskyĆice Dowex 50 W X2-H+ forma ve smÄsi voda â dioxan 2 : 1bylo naplnÄno do kolony, kterou pak pro-chĂĄzel roztok 1 g sirupovitĂ© lĂĄtky zĂskanĂ©v pĆĂkladu 1 [ii) v 20 ml smÄsi voda â di-oxan 2 : 1. Kolona pak byla promĂœvĂĄna smÄ-sĂ voda â dioxan 2 : 1 aĆŸ smÄs vy Ă©kajĂcĂ zkolony byla neutrĂĄlnĂ a pak byla provede-na eluce lineĂĄrnĂm gradientem smÄsi vo-da â dioxan 2 : 1 obsahujĂcĂ 0 aĆŸ 1 N Äpa-vek. EluovanĂ© frakce obsahujĂcĂ ĆŸĂĄdanĂœ3,6â-di-N-benzyloxykarbonylkanamycin A by-ly koncentrovĂĄny do sucha za snĂĆŸenĂ©ho tla-ku, ÄĂmĆŸ sc: zĂskalo 311 mg ti, 84 % bĂlĂ©pevnĂ© lĂĄtky, jeĆŸ byla shodnĂĄ lĂĄtkou zĂs-kanou v pĆĂkladu 1 (iii 1 [ B). (D) AlternativnĂ postup pouĆŸĂvajĂcĂ Do-wex 50W X 2
Roztok 1 g sirupovitĂ© lĂĄtky zĂskanĂ© v pĆĂ-kladu 1 (ii) ve 20 ml smÄsi voda - meta-nol 3 : 1 byl nanesen na kolonu 30 ml Do-wexu 50W X 2 v H+ formÄ pĆedem smoÄe-nou smÄsĂ voda â metanol 3 : 1. Kolona pakbyla dobĆe promyta smÄsĂ voda - metanol3 : 1, a pak byla elucvĂĄna gradientem smÄ-si voda â metanol 3 : 1 obsahujĂcĂ 0 aĆŸ 6 Nkyselinu chlorovodĂkovou. AktivnĂ frakce ob-sahujĂcĂ ĆŸĂĄdanĂœ 3,6â-di-N-benzyloxykarhonyI-kanamycin A byly sebrĂĄny a smĂchĂĄny se 36 silnÄ zĂĄsaditou anexovou iontomÄniÄovoupryskyĆicĂ Dowex 1 X 2 v OH formÄ v mnoĆŸ-stvĂ, jeĆŸ bylo schopnĂ© pĆĂmÄs mĂrnÄ okyse-lit.
SmÄs byla zfiltrovĂĄna a filtrĂĄt byl za-koncentrovĂĄn do sucha, ÄĂmĆŸ se zĂskalo286 mg, tj. 72 % ĆŸĂĄdanĂ©ho produktu veformÄ dihydrochloridu.
[ce]jj35 â +79° (c 1, voda â dimetylforma-xnid, 1 : 2). (E) ZpĆŻsob pouĆŸĂvajĂcĂ anexovou ionto-mÄniÄovou pryskyĆici nesoucĂ silnÄ bazic-kĂ© funkÄnĂ kvartĂ©rnĂ amoniovĂ© skupiny,komerÄnÄ' dostupnĂœ jako Dowex 1X2cd. Dow Chemical Co.
Roztok 1 g sirupovitĂ© lĂĄtky zĂskanĂœ v pĆĂ-kladu Ă (ii) ve smÄsi voda â dioxan (1 : : 1) byl nanesen na kolonu 30 ml prysky-Ćice Dowex 1 X 2 v OH formÄ pĆedem pro-mytĂ© smÄsĂ voda â dioxan 1 : 1, a pak by-la kolona vyvolĂĄna smÄsĂ voda â dioxan 1 : : 1 pomÄrnÄ vysokou rychlostĂ. EluovanĂ©frakce obsahujĂcĂ ĆŸĂĄdanĂœ produkt byly sbĂ-rĂĄny a zakoncentrovĂĄny do sucha, ÄĂmĆŸ sezĂskalo 305 mg, tj. 84 % bezbarvĂ© pevnĂ©lĂĄtky totoĆŸnĂ© s lĂĄtkou z pĆĂkladu 1 (iii) (B). (F) Postup pouĆŸĂvajĂcĂ anexovou ionto-mÄniÄovou pryskyĆici se slabÄ bazickĂœmifunkÄnĂmi skupinami, komerÄnÄ dostup-nou jako Dowex WGR, vĂœrobek fy DowChemical Co. 1 g sirupovitĂ© lĂĄtky zĂskanĂ© v pĆĂkladu1. (ii.) byl rozpuĆĄtÄn v 20 ml smÄsi voda â - dioxan 2:1a roztok se nechal protĂ©cikolonou 50 ml Dowexu WGR v bazickĂ© for-irÄ pĆedem vysycenou smÄsĂ voda â· dio-xan 2:1a puk byla provedena eluce smÄsĂvoda - dloxĂ· i 2 : 1. ĆœĂĄdanĂœ 3,6â-di-N-ben-zyloxykarbc.nyikanamycin A byl eluovĂĄn vnÄkterĂœch frakcĂch spolu se stopami zineÄ-eaĂĂœch kationtĆŻ. Tyto frakce byly spojeny azakoncentrovĂĄny do sucha, ÄĂmĆŸ se zĂskalo450 mg bezbarvĂ© pevnĂ© lĂĄtky. fG) Postup pouĆŸĂvajĂcĂ chelĂĄtovou ionto-mÄniÄovou pryskyĆici nesoucĂ slabÄ ky-selĂ© funkÄnĂ skupiny, komerÄnÄ dostup-nou jako Dowex A 1, vĂœrobek fy Dow Che-mical Co., Sp. st. a.
Roztok 1 g sirupovitĂ© lĂĄtky zĂskanĂ© v pĆĂ-kladu 1 (ii) ve smÄsi voda â dioxan 1 :: 1 byl nanesen na kolonu 50 ml DowexuAI, kterĂĄ byla vysycena smÄsĂ voda â dio-xan 1 : 1 obsahujĂcĂ 1% amoniak a pak nĂĄ-sledovala eluce gradientem smÄsi voda â - dioxan 1 : 1 obsahujĂcĂ 0 aĆŸ 1 N amoniak.Frakce obsahujĂcĂ ĆŸĂĄdanĂœ 3,6â-di-N-benzylo-xykarbonylkanamycin A, kterĂ© byly eluovĂĄ-ny pouze v pozdÄjĆĄĂ fĂĄzi, byly spojeny a za-huĆĄtÄny do sucha, ÄĂmĆŸ vzniklo 272 mg tj. 261853 37 74 % ĆŸĂĄdanĂ©ho' produktu ve formÄ pevnĂ©bĂlĂ© lĂĄtky. (H) Postup pouĆŸĂvajĂcĂ Chitosan, ve vodÄnerozpustnĂœ polymer obsahujĂcĂ funkÄnĂskupiny schopnĂ© vazby s kovem, komerÄ-nÄ dostupnĂœ jako vĂœrobek fy Toko KaseiKoyo Co., Ltd. Japonsko'. 100 ml Chitosanu byla dĆŻkladnÄ nasyce-no smÄsĂ voda â metanol 3:1a naplnÄno-do kolony, kterou se nechal protĂ©ci roztok1 g sirupovitĂ© lĂĄtky zĂskanĂ© v pĆĂkladu 1(ii) ve smÄsi voda â metanol (3 : 1). Ko-lona pak byla vyvolĂĄna smÄsĂ voda â me-tanol 3 : 1, pĆiÄemĆŸ ĆŸĂĄdanĂœ 3,6â-di-N-ben-zyloxykarbonylkanamycin A byl eluovĂĄn ja-ko prvnĂ a oct-an zineÄnatĂœ mnohem poz-dÄji. Frakce obsahujĂcĂ ĆŸĂĄdanou lĂĄtku bylyspoijeny a zakoncentrovĂĄny do sucha, ÄĂmĆŸvznikl zbytek, kterĂœ byl rozpuĆĄtÄn ve smÄ-si voda â dioxan 1:1a roztok byl nane-sen na kolonu Amber litu CG 50 v NH4+formÄ pĆedem proimytou smÄsĂ voda â dio-xan 1 : 1. Kolona byla dĆŻkladnÄ promytasmÄsĂ voda â dioxan 1:1a pak byla eluo-vĂĄna gradientem smÄsi voda â dioxan 1 : 1obsahujĂcĂ 0 aĆŸ 0,1 N amoniak. Tyto frakcecitlivĂ© k ninhydrinovĂ© reakci byly spojenya zakoncentrovĂĄny do sucha, ÄĂmĆŸ se zĂska-lo 301 mg, tj. 82 °/o bezbarvĂ© pevnĂ© lĂĄtkyshodnĂ© s lĂĄtkou zĂskanou v pĆĂkladu 1 (iii)(B). (I) Postup pouĆŸĂvajĂcĂ vysokĂ©ho polymerunesoucĂho karboxylovĂ© funkÄnĂ skupiny,obchodnÄ dostupnĂ©ho jako ,,CM-Sepha-dexâ Câ25, kterĂœ je iontomÄniÄem i lĂĄt-kou pro gelovou filtraci a je vytvoĆen zkarboxymetyl-substituovanĂ© dextranovĂ©-ho gelu, vĂœrobek fy Pharmacia Fine Che-mical Co., Ć vĂ©dsko1.
Roztok 1 g sirupovitĂ© lĂĄtky zĂskanĂ© v pĆĂ-kladu 1 (ii) ve smÄsi voda â dioxan 1 : 1se nechal protĂ©ci kolonou 40 ml CM-Sepha-dexu Câ25 v NH4+ formÄ, dĆŻkladnÄ nasy-cenĂ©ho smÄsĂ voda â dioxan 1 : 1. Kolonabyla promyta 200 ml smÄsi voda â dioxan1:1a pak eluovĂĄna gradientem smÄsi vo-da â dioxan 1 : 1 obsahujĂcĂ 0 aĆŸ 0,1 N a-moniak. Z kolony nebyly eluovĂĄny ĆŸĂĄdnĂ©kationty zinku, ale pouze ĆŸĂĄdanĂœ 3,6â-di-N--benzyloxykarbonylkanamycin A. EluĂĄt bylkoncentrovĂĄn do sucha, ÄĂmĆŸ vzniklo 303miligramĆŻ, tj. 82 % bezbarvĂ© pevnĂ© lĂĄtkyshodnĂ© s lĂĄtkou v pĆĂkladu 1 (iii) (BJ. ()) Postup pouĆŸĂvajĂcĂ sirovodĂk jako lĂĄt-ku precipitujĂcĂ zinek. 1 g sirupovitĂ© lĂĄtky zĂskanĂ© v pĆĂkladu 1(ii) byl rozpuĆĄtÄn ve 20 ml smÄsi voda â-â metanol 1 : 1, a k tomuto roztoku bylpak pĆidĂĄn vodnĂœ amoniak a potĂ© bylo za-vedeno dostateÄnĂ© mnoĆŸstvĂ sirovodĂku. Re-akÄnĂ smÄs obsahujĂcĂ sraĆŸeninu sirnĂku zi- 38 neÄnatĂ©ho, kterĂœ se vytvoĆil, byla zfiltrovĂĄ-na na sklenÄnĂ©m filtru s celitovou vloĆŸkoua filtrĂĄt byl zahuĆĄtÄn za snĂĆŸenĂ©ho tlaku,ÄĂmĆŸ vznikla sirupovitĂĄ lĂĄtka, ze kterĂ© popromytĂ etylĂ©terem vznikl pevnĂœ zbytek. Ten-to zbytek byl rozpuĆĄtÄn ve smÄsi voda ââ dioxan 1:1a roztok byl chromatogra-fovĂĄn na kolonÄ 30 ml Amberlitu IRA 900,coĆŸ je silnÄ bazickĂĄ pryskyĆice, vĂœrobek fyRohm and Haas Co., v OH formÄ, za pouĆŸi-tĂ smÄsi voda â dioxan 1 : 1 jako vyvolĂĄ-vacĂho rozpouĆĄtÄdla. EluĂĄt byl shromaĆŸÄovĂĄnve frakcĂch a frakce obsahujĂcĂ 3,6â-di-N--benzyloxykarbonylkanamycin A byly spoje-ny a zahuĆĄtÄny do sucha, ÄĂmĆŸ se zĂskalo235 mg, tj. 64 % bezbarvĂ© pevnĂ© lĂĄtky shod-nĂ© s lĂĄtkou v pĆĂkladu 1 (iii) (B). PĆĂklad 2
PĆĂprava 3,6-di-N-benzyloxykarbonylkana-mycinu A 500 mg, tj. 1,03 mM kanamycinu A veformÄ volnĂ© bĂĄze bylo suspendovĂĄno v 15mililitrech dimetylsulfoxidu a potĂ© bylo- pĆi-dĂĄno 420 mg, tj. 3,09 mM chloridu zineÄna-tĂ©ho a 840 mg, tj. 6,18 mM octanu sodnĂ©hotrihydrĂĄtu. Po desetihodinovĂ©m mĂchĂĄnĂsmÄsi pĆi okolnĂ teplotÄ, byl ke smÄsi ob-sahujĂcĂ vytvoĆenĂœ komplex kanamycin A--zinek pĆidĂĄvĂĄn pomalu asi po jednu hodi-nu roztok 675 mg, tj. 2,27 mM N-benzyloxy-karbanyloxyftalimidu
rozpuĆĄtÄnĂ©ho v 10 ml dimetylsulfoxidu. VĂœ-slednĂĄ smÄs byla ponechĂĄna stĂĄt pĆi tep-lotÄ mĂstnosti 4 hodiny. DĂĄle byla reakÄnĂ smÄs zpracovĂĄna stej-nĂœm zpĆŻsobem, jak bylo popsĂĄno v pĆĂkla-du 1 (iij a (iii) (I), ÄĂmĆŸ se zĂskalo 598miligramĆŻ, tj. 74 °/o 3,6â-di-N-benzyloxykar-bonylkanamycinu A ve formÄ bezbarvĂ© pev-nĂ© lĂĄtky. PĆĂklad 3
PĆĂprava 3,6â-di-N-benzyloxykarbcnylkana-mycinu A 600 mg, tj. 0,95 mM kanamycinu A tetra-hydrochloridu a 150 mg, tj. 3,8 mM hydro-xidu sodnĂ©ho v 15 ml dimetylsulfoxidu by-lo tĆepĂĄno jednu hodinu a pak bylo pĆidĂĄno'1 g, tj. 4,55 mM octanu zineÄnatĂ©ho dihyd-rĂĄtu, a v tĆepĂĄnĂ bylo pokraÄovĂĄno dalĆĄĂch5 hodin. Ke smÄsi obsahujĂcĂ vytvoĆenĂœ kom-plex kanamycinu A-zinek byl pĆidĂĄn bÄhem 261853 39 40 30 minut roztok 545 mg, tj. 2,2 mM N-ben-zyloxykarbonyloxysukcinimidu rozpuĆĄtÄnĂ©hov 5 ml smÄsi dimetylsulfoxidtetrahydrofu-ran 1 : .1. Po protĆepĂĄnĂ vĂœslednĂ© smÄsi pĆiokolnĂ teplotÄ pĆes noc byl pĆidĂĄn etylĂĄ-ter, aby se oddÄlil N-acylovanĂœ zineÄnatĂœkomplex jako precipitĂĄt. PrecipitĂĄt byl pakzpracovĂĄn stejnĂœm zpĆŻsobem jak bylo po-psĂĄno v pĆĂkladu 1 (iiij (H), ÄĂmĆŸ se zĂs-kalo 581 mg, tj. 78 % bezbarvĂ© pevnĂ© lĂĄtky.PĆĂklad 4 PĆĂprava 3.6â-di-N-benzyloxykarbonylkana-
mycinu A (i] 500 mg, tj. 1,03 mM kanamycinu A veformÄ volnĂ© bĂĄze bylo rozpuĆĄtÄno v 20 mlsmÄsi voda â dimetylsulfoxid 1 : 9, pakbylo pĆidĂĄno 1 g, tj. 4,55 mM octanu zineÄ-natĂ©bo dihydrĂĄtu a potom 590 mg, tj. 2,4mM N-benzyloxykarbon.yloxysukcinimidu.
SmÄs se nechala stĂĄt pĆi okolnĂ teplotÄpĆes noc a ke smÄsi bylo pĆidĂĄno velkĂ©mnoĆŸstvĂ etylĂ©teru, ÄĂmĆŸ se oddÄlila vodnĂĄsirupovitĂĄ vrstva, kterĂĄ byla promyta nÄ-kolikrĂĄt etylĂ©terem, ÄĂmĆŸ se zĂskala hustĂĄsirupovitĂĄ vrstva. (ii) Takto zĂskanĂĄ sirupovitĂĄ lĂĄtka bylarozpuĆĄtÄna ve smÄsi voda --- metanol 3 : Ia roztok se nechal projĂt kolonou 200 mlChitosanu. Kolona byla eluovĂĄna smÄsĂ vo-da - metanol 3 : 1 a eluĂĄt byl jĂmĂĄn vefrakcĂch. Frakce pozitivnĂ k ninhydrhmvĂ©reakci byly spojeny a zahuĆĄtÄny do malĂ©-ho objemu. KoncentrĂĄt byl nanesen na ko-lonu Amberlitu CG 50 v NII+ formÄ a kolo-na byla dĆŻkladnÄ promyta smÄsĂ voda - di-oxan 1 : 1 a pak eluovĂĄna gradientem smÄ-si voda â dioxan 1 : 1 obsahujĂcĂ 0 aĆŸ 0,1 NÄpavek.
EluovanĂ© frakce obsahujĂcĂ ĆŸĂĄdanĂœ pro-dukt byly spojeny a zakoncentrovĂĄny do su-cha, ÄĂmĆŸ se zĂskalo 494 mg, tj. 6J % bez-barvĂ© pevnĂ© lĂĄtky, shodnĂ© s lĂĄtkou zĂska-nou v pĆĂkladu 1 (iiij (B). P Ć Ă k la d 5
PĆĂprava 3,6â-di-N-benzytoxykarbonyl-kanamycinu A 500 mg, tj. 1,03 mM kanamycinu A ve for-mÄ volnĂ© bĂĄze bylo rozpuĆĄtÄno ve 20 mlsmÄsi voda â tetrahydrofuran 1 : 3, a ktomu byl pak pĆidĂĄn 1 g, tj. 4,55 mM octa-nu zlneÄnatĂ©ho dihydrĂĄtu a pak bylo pĆi-dĂĄno 590 mg, tj. 2,4 mM N-benzyloxykarbo-nyloxysukcinimidu. SmÄs byla ponechĂĄnastĂĄt pĆi teplotÄ okolĂ pĆes noc a reakÄnĂroztok takto zĂskanĂœ byl zahuĆĄtÄn za snĂĆŸe-nĂ©ho tlaku. Zbytek se nechal protĂ©ci kolo-nou s 200 ml Chitosanu a eluĂĄt byl zpraco-vĂĄn stejnĂœm zpĆŻsobem jako v pĆĂkladu 4(iij, ÄĂmĆŸ se zĂskalo 414 mg, tj. 51 % bez-barvĂ© pevnĂ© lĂĄtky ĆŸĂĄdanĂ© slouÄeniny. PĆĂklad 6'
PĆĂprava 3,6â-di-N-benzyloxykarbonyl-kanamycinu A jij 500 mg, tj. 1,03 mM kanamycinu A veformÄ volnĂ© bĂĄze bylo rozpuĆĄtÄno v 15 mlsmÄsi voda â metanol 1 : 7, a k tomu by-lo pak pĆidĂĄno· 1,5 g, tj. 6,8 mM octanu zi-neÄnatĂ©ho dihydrĂĄtu a pak bylo pĆidĂĄno 590miligramĆŻ, tj. 2,4 mM N-benzyloxykarbonyl-oxysukcinimidu v 7 ml tetrahydrofuranu.SmÄs byla ponechĂĄua stĂĄt pĆi teplotÄ okolĂpĆes noc a reakÄnĂ roztok takto zĂskanĂœ bylzahuĆĄtÄn za snĂĆŸenĂ©ho tlaku. Zbytek se paknechal projĂt kolonou 200 ml Chitosanu aeluĂĄt vychĂĄzejĂcĂ z kolony byl pak zpraco-vĂĄn stejnĂœm zpĆŻsobem jako v pĆĂkladu 4(iij, ÄĂmĆŸ se zĂskalo· 442 mg, tj. 55 % bez-barvĂ© pevnĂ© lĂĄtky ĆŸĂĄdanĂ© slouÄeniny. PĆĂklad 7
PĆĂprava 3,6â-di-N-benzyloxykarbonyl-kanamyciiiu A 500 mg, tj. 1,03 mM kanamycinu A ve for-mÄ volnĂ© bĂĄze bylo suspendovĂĄno v 20 mldimetylsulfoxidu a 272 mg, tj. 1,24 mM oc-tanu zineÄnatĂ©ho dihydrĂĄtu bylo k tĂ©tosmÄsi pĆidĂĄno. SmÄs byla mĂchĂĄna pĆi tep-lotÄ mĂstnosti 10 hodin, pĆiÄemĆŸ se vytvoĆiltĂ©mÄĆ ÄirĂœ roztok, ke kterĂ©mu pak bylo pĆi-dĂĄvĂĄno po malĂœch ÄĂĄstech po asi dvÄ hodi-ny 540 mg, tj. 2,17 mM N-benzyloxykarbo-nyloxysukcinimidu.
Pak se nechala vĂœslednĂĄ smÄs stĂĄt pĆiteplotÄ okolĂ pĆes noc, byl pĆidĂĄn velkĂœ ob-jem etylĂ©teru a oddÄlenĂĄ oiejovitĂĄ lĂĄtka by-la odebrĂĄna a promyta nÄkolikrĂĄt etylĂ©te-rem, ÄĂmĆŸ vznikla hustĂĄ sirupovitĂĄ lĂĄtka.
PomocĂ chromatografie na tenkĂ© vrstvÄsilikagelu se ve vzorku odebranĂ©m ze si-rupovitĂ© lĂĄtky, pĆi pouĆŸitĂ smÄsi chloro-form â metanol â 28% vodnĂœ Äpavek 1 : : 1 : 1, dolnĂ fĂĄze, jako vyvolĂĄvacĂho roz-pouĆĄtÄdla podaĆilo urÄit tyto skvrny: â menĆĄĂ skvrna o R, 0,4â1,3 6â,3â-tetra~N--benzyloxykarbonylkanamycin A, kde sevyvĂjelo zabarvenĂ po postĆiku kyselinousĂrovou a nĂĄsledovnĂ©m zahĆĂĄtĂ; â slabĂĄ skvrna o Rt 0,28; â hlavnĂ skvrna o R( 0,23 â ĆŸĂĄdanĂœ pro-dukt, 3,6â-di-N-benzyloxykarbonylkana-mycin A; â menĆĄĂ skvrna o Rf 0,12 6â-N-benzyloxy-karbonyl-kanamycin A; a â velmi slabĂĄ skvrna o Rf 0 â nezreago-vanĂœ kanamycin A.
Nebyla pozorovĂĄna skvrna odpovĂdajĂcĂ tri-N-benzyloxykarbonylkanamycinu A, kte- rĂĄ by se niÄla objevit pĆi RÂŁ 0,28 aĆŸ 0,4. VĂœĆĄe uvedenĂĄ sirupovitĂĄ lĂĄtka byla roz- puĆĄtÄna ve smÄsi voda â dioxan 1:1a roz- 261853 41 42 tok byl nechĂĄn, projĂt kolonou se 100 mlCM-Sephadexu Câ25 v NH4+ formÄ, pĆedempromytou smÄsĂ voda â dioxan 1 : 1. Pakbyla kolona eluovĂĄna stejnĂœm postupem jakbylo popsĂĄno v pĆĂkladu 1 (iii) (I), pĆiÄemĆŸbyly odstranÄny kationty zinku a ĆŸĂĄdanĂœprodukt byl oddÄlen od jinĂœch produktĆŻ,ÄĂmĆŸ se zĂskalo, 412 mg, tj. 51 % ĆŸĂĄdanĂ©slouÄeniny ve formÄ bezbarvĂ© pevnĂ© lĂĄtky.
Pro srovnĂĄnĂ byl tento postup opakovĂĄns tĂm rozdĂlem, ĆŸe byl octan zineÄnatĂœ di-hydrĂĄt nahrazen 308 g, tj, 1,24 mM octanunikelnatĂ©ho tetrahydrĂĄtu, s tĂm vĂœsledkem,ĆŸe ĆŸĂĄdanĂœ 3,6â-di-N-benzyloxykarbonylkana-mycin A byl zĂskĂĄn jako bezbarvĂĄ pevnĂĄlĂĄtka pouze s vĂœtÄĆŸkem 59 mg, tj. 7,3 %. PĆĂklad 8
PĆĂprava 3,6â-di-N-(p-metoxybenzyloxykar-bonyl) kanamycinu A 500 mg, tj. 1,03 mM kanamycinu A veformÄ volnĂ© bĂĄze bylo suspendovĂĄno ve 12mililitrech dimetylsulfoxidu a k suspenzi bylpĆidĂĄn 1 g, tj. 4,55 mM octanu zineÄnatĂ©hodihydrĂĄtu. SmÄs byla mĂchĂĄna pĆi teplotÄmĂstnosti aĆŸ do vytvoĆenĂ homogennĂho roz-toku, k nÄmuĆŸ pak byl pĆidĂĄn bÄhem 30minut roztok 789 mg, tj. 2,6 mM p-met?xy-karbobenzoxy-p-nitrofenylesteru(pâCHaOCfiH^CH^OCOOCoH^pâNO2) rozpuĆĄ-tÄnĂ©ho v dimetylsulfoxidu. VĂœslednĂĄ smÄs se nechala stĂĄt pĆi tep-lotÄ okolĂ pĆes noc a pak byla zpracovĂĄnastejnĂœm zpĆŻsobem jako v pĆĂkladu 1 (ii) a (iii) (B), ÄĂmĆŸ se zĂskalo 722 mg, tj. 83 %bezbarvĂ© pevnĂ© lĂĄtky ĆŸĂĄdanĂ© slouÄeniny.
[a]n25 +87° (c 1, voda â dimetylformamid, 1 : 2).
ElementĂĄrnĂ analĂœza
VypoÄteno pro C^jH^N-iOiy . V2 HâCO·,: 51,95 % C, 6,33 % H, 6,64 % N,
Nalezeno: 51,56 % C, 6,41 % H, 6,53 % N.PĆĂklad 9
PĆĂprava 6â-N- (t-butoxykarbonyl jkanamy-cin A
StejnĂœm postupem jako bylo popsĂĄno vpĆĂkladu 8, s tĂm rozdĂlem, ĆŸe p-metoxykar-bobenzoxy-p-nitrofenylester byl nahraĆŸen220 mg, tj. 1,54 mM t-butoxykarbonylazidu,byla zĂskĂĄna ĆŸĂĄdanĂĄ slouÄenina ve formÄbezbarvĂ© pevnĂ© lĂĄtky. VĂœtÄĆŸek Äinil 627miligramĆŻ.
[«Id25 = +96° (c 1, voda â dimetylforma- mid, 1 : 2), PĆĂklad 10
PĆĂprava 3,6â-di-N-trifluoroacetylkanamyci-nu A 500 mg, tj. 1,03 mM kanamycinu A veformÄ volnĂ© bĂĄze bylo suspendovĂĄno ve 12mililitrech dimetylsulfoxidu a k suspenzi bylpĆidĂĄn 1 g, tj. 4,55 mM octanu zineÄnatĂ©hodihydrĂĄtu. SmÄs byla mĂchĂĄna pĆi teplotÄmĂstnosti aĆŸ do vytvoĆenĂ homogennĂho roz-toku, ke kterĂ©mu pak byl pĆidĂĄn roztok 1,2 g, tj. 5,1 mM p-nitrofenolesteru kyseli-ny trifluoroctovĂ© rozpuĆĄtÄnĂ© v 10 ml dime-tylsulfcxidu. VĂœslednĂĄ smÄs byla ponechĂĄ-na stĂĄt pĆes noc pĆi teplotÄ okolĂ a pakbyla zpracovĂĄna s etylĂ©terem stejnÄ jako vpĆĂkladu 1 (ii). V Ă©teru nerozpustnĂœ siru-povitĂœ materiĂĄl byl dĂĄle zpracovĂĄn stejnĂœmzpĆŻsobem jako v pĆĂkladu 1 (iii) (A), ÄĂmĆŸse zĂskalo 590 mg (70%) ĆŸĂĄdanĂ© slouÄeniny ve formÄ bezbarvĂ© pevnĂ© lĂĄtky.
[Îż?]Ï 25 +81° (c 1, voda â dimetylformamid, 1 : 2).
ElementĂĄrnĂ analĂœza
VypoÄteno pro C^H^NĂOriF,!. 2 CH iCO-H . H-,0: 38,33 % C, 5,44 % H, 6,88 % N,13,99 % F,
Nalezeno^: 33,03 % C, 5,48 % H, 6,54 % N. PĆĂklad 11
PĆĂprava 3,6â-di-N-fenoxykarbonylkanamy-cinu A 500 mg, tj. 1,03 mM kanamycinu A veformÄ volnĂ© bĂĄze bylo· suspendovĂĄno· vesmÄsi dimetylsulfoxidu, 15 ml a 5 ml tetra-hydrofuranu a k suspenzi byl pĆidĂĄn 1 g,tj. 4,55 mM octanu zineÄnatĂ©ho dihydrĂĄtu apotĂ© byla smÄs mĂchĂĄna pĆi teplotÄ mĂst-nosti aĆŸ do vytvoĆenĂ homogennĂho, rozto-ku. VĂœslednĂœ roztek byl pak ochlazen na0 CC a k tomu byl pak pomalu pĆidĂĄvĂĄnchladnĂœ roztok 0 °C 400 mg, tj. 2,55 mM fe-noxykarbonylchloridu C(iH-,OCOCl ve 3 mltetrahydrofuranu. ReakÄnĂ smÄs byla pĆe-nesena do teploty mĂstnosti za jednu hodi-nu a pak byla nechĂĄna stĂĄt pĆi tĂ©to tep-lotÄ 3 hodiny. Potom byla reakÄnĂ smÄs zpra-covĂĄna s etylĂ©terem stejnÄ jako, v pĆĂkladu1 (ii) a v Ă©teru nerozpustnĂĄ sirupovitĂĄ lĂĄt-ka byla dĂĄle zpracovĂĄna stejnĂœm zpĆŻsobemjako v pĆĂkladu 1 (iii) (A), ÄĂmĆŸ se zĂska-lo 625 mg, tj. 70 % ĆŸĂĄdanĂ© slouÄeniny veformÄ bezbarvĂ© pevnĂ© lĂĄtky.
[ajD25 +73° (c 1, voda â dimetylformamid, 1:2).
ElementĂĄrnĂ analĂœza
VypoÄteno: pro C^H^N^O,-, . 2 CH-CO-Jl . η,Î: 50,11 % C, 6,31 % II, 6,49 % N,
Nalezeno·: 49,77 % (1, 6,60 % H, 6,11 % N. PĆĂklad 12
PĆĂprava 3,6â-di-N-acetylkanainycinu A
ReakÄnĂ smÄs zĂskanĂĄ stejnĂœm postupemjako v pĆĂkladu 8, s vĂœjimkou, ĆŸe bylo pou-ĆŸito 260 mg, tj. 2,6 mM acetanhydridu mĂsto p-metoxykarbobenzoxy-p-nitrofeny] este-ru, byla zpracovĂĄna stejnĂœm zpĆŻsobem ja-ko v pĆĂkladu 1 (iii) (A). Takto bylo pĆi-praveno 525 mg, tj. 72 % ĆŸĂĄdanĂ© slouÄeni-ny ve formÄ bezbarvĂ© pevnĂ© lĂĄtky.
[«]âÂź - I 93 (cmid, 1 : 2). AnalĂœza VypoÄteno 1, voda â dimetylfurma pro C%HWN,.O|:,. 2 Cll.;CĂ),l I.. 11,0: 44,19 % C, 7,13 % II, 7,63 % N, Nalezeno: 44,20 % C, PĆĂklad 13 7,07 % H, 7,85 % N.
PĆĂprava 3,6â-di-N-lorruylkanarnycinu A 500 rng, tj. 1,03 mM kanamyciuu A veformÄ volnĂ© bĂĄze bylo suspendovĂĄno ve 12mililitrech dimetylsulfoxidu a k tĂ©to sus-penzi by] pĆidĂĄn 1 g, tj. 4,55 mM octa nu zi-neÄnatĂ©ho dihydrĂĄtu. SmÄs byla mĂchĂĄnapĆi teploto mĂstnosti aĆŸ do vytvoĆenĂ homo-gennĂho roztoku, ke kterĂ©mu pak bylo pĆi-dĂĄno 690 mg, tj. 4,12 mM p-nitrofenylformĂĄ-tu OHCOD(iH5- -p -NO·,. VĂœslednĂĄ smÄs by-la ponechĂĄna stĂĄt pĆes noc pĆĂ teplotÄ oko-lĂ a pak byla zpracovĂĄna stejbiĂœm zpĆŻsobemjako v pĆĂkladu 1 (iii] (H).
Frakce pozitivnĂ k uinlivdriuovĂ© reakcibyly spojeny, probuldĂĄny plynnĂœm oxidemuhliÄitĂœm a pak zahuĆĄtÄny do sucha. Tak-to se zĂskalo 430 mg, tj. 67 % ĆŸĂĄdanĂ© slou-Äeniny ve formÄ bezbarvĂ© lĂĄtky. fajir5 |101° (c 1, voda).
AnalĂœza
VypoÄteno pro . H9CO·, .11,0: 40,64 % C, "6,50 % H, 9,03 % N,
Nalezeno: 40,43 % C, 6,47 % H 8,83 % N. 44 PĆĂklad 14
PĆĂprava 3,6â-di-N-tosylkanamycinu A 500 mg, tj. 1,03 mM kanamycinu A veformÄ volnĂ© bĂĄze bylo* suspendovĂĄno v 15mililitrech dimetylsulfoxidu a k suspenzi bylpĆidĂĄn 1 g, tj. 4,55 mM octanu zineÄnatĂ©hodihydrĂĄtu. SmÄs byla mĂchĂĄna pĆi teplotÄmĂstnosti aĆŸ do vytvoĆenĂ homogennĂho1 roz-toku, k nÄmuĆŸ pak byl pomalu pĆidĂĄn roz-tok 400 mg, tj. 2,1 mM tosylchloridu v 7 mltetrahydrofuranu. VĂœslednĂĄ smÄs se nechala stĂĄt pĆi teplo-tÄ okolĂ jednu hodinu, a pak bylo pĆidĂĄno200 mg tosylchloridu rozpuĆĄtÄnĂ©ho ve 3,5mililitrech tetrahydrofuranu. ReakÄnĂ smÄsbyla ponechĂĄna dalĆĄĂ dvÄ hodiny v klidu apak byla zpracovĂĄna postupem stejnĂœm ja-ko v pĆĂkladu 1 (iij a (iii) (Aj, pĆiÄemĆŸ sezĂskalo 270 mg, tj. 28 % bezbarvĂ© pevnĂ©lĂĄtky, pĆedstavujĂcĂ ĆŸĂĄdanou slouÄeninu. (α]ÎŻ,25 +68° (c 1, voda â dimetylformamid, 1 : 2).
Ana lĂœzaVypoÄteno pro C39H48N,,Or>S3 . 2 CH3CO2H . Î,Î: 46,44 % C, 6,28 % H, 6,02 % N, 6,89 % S,Nalezeno: 46,31 % C, 5,98 % H, 6,31 % N, 6,55 % S. PĆĂ provĂĄdÄnĂ vĂœĆĄe uvedenĂ©ho reakÄnĂhopostupu bez octanu zineÄnatĂ©ho, se nepoda-Ćilo zĂskat vÄtĆĄĂ mnoĆŸstvĂ bezbarvĂ© pevnĂ©lĂĄtky. P Ć Ă k 1 a d 1 5
PĆĂprava 3,6'-di-N-benzyloxykarbonyl-6â-N-- m et y 1 k a n a ru yc i n u A 500 mg, tj. t,0 mM 6'-N-nietyl-kanamyci-nu A ve hĆme volnĂ© bĂĄze bylo suspendo-vĂĄno ve 12 ml dimetylsulfoxidu a k sus-penzi byl pĆidĂĄn 1 g, tj. 4,55 mM octanu zi-ncÄuatĂ©ho dihydrĂĄtu. SmÄs byla mĂchĂĄnaoĆi teplotÄ mĂstnosti aĆŸ do vytvoĆenĂ homo-gennĂho roztoku, ke kterĂ©mu byl bÄhem 30minut pĆidĂĄn roztok 550 mg, ti. 2,2 mM N--benzyloxykarbonyloxysukcinimidu rozpuĆĄ-tÄnĂ©ho v 5 ml dimetylsulfoxidu tetrahydro-furanu 1:1. VĂœslednĂĄ smÄs byla ponechĂĄna v klidupĆes noc pĆi teplotÄ okolĂ a pak byla zpra-covĂĄna stejnÄ jako v pĆĂkladu 1 (iij a (iii)(A), ÄĂmĆŸ se zĂskalo 720 mg, tj. 79% ĆŸĂĄda-nĂ© slouÄeniny ve formÄ bezbarvĂ© lĂĄtky.
[oĂ],j-5 -1-74° (c 1, voda â dimetylformamid, 1 : 2).
DalĆĄĂm zpracovĂĄnĂm takto pĆipravenĂ© 261853 45 slouÄeniny postupem podobnĂœm jako v pĆĂ-kladu 31 uvedenĂ©m nĂĆŸe, se zĂskal 1-N-[(S)--4-amino-2-hydroxybutyryl]-6â-N-metylka-namycin A. PĆĂklad 16
PĆĂprava 3,6â-di-N-benzyloxykarbonyl-3â-deoxykanamycinu A
Tato slouÄenina ve formÄ bezbarvĂ© pevnĂ©lĂĄtky byla zĂskĂĄna s vĂœtÄĆŸkem 765 mg, tj.82 % opakovĂĄnĂm stejnĂ©ho postupu jako vpĆĂkladu 15, avĆĄak s tĂm rozdĂlem, ĆŸe vĂœ-chozĂ lĂĄtkou bylo 500 mg, tj. 1,07 mM 3â-de-oxykanamycinu A ve formÄ volnĂ© bĂĄze a by-lo pouĆŸito 610 mg, tj. 2,45 mM N-benzyloxy-karboinyloxysukcinimidu.
[a]D25 â +76° (c 1, voda â dimetylforma-mid, 1:2).
AnalĂœza
VypoÄteno pro C-hI-V^Oh . 2 CH.,CO2iH . H..O: 52,16 % C, 6,68 % H, 6,40 % N,
Nalezeno: 51,99 % C, 6,75 % H, 6,20 % N.
DalĆĄĂm zpracovĂĄnĂm takto pĆipravenĂ©slouÄeniny postupem podobnĂœm jako v pĆĂ-kladu 31 se zĂskal l-N-[ (S)-4-amino-2-hyd-roixybutyrylj-3â-deoxykanamycin A.PĆĂklad 17
PĆĂprava 3,6â-di-N-benzyloxykarbonyl-3â-de-oxy-6â-N-metylkanamycinu A ĆœĂĄdanĂĄ slouÄenina byla zĂskĂĄna s vĂœtÄĆŸ-kem 737 mg, tj. 80 % opakovĂĄnĂm stejnĂ©-ho' postupu jako v pĆĂkladu 15 s tĂm rozdĂ-lem, ĆŸe se vychĂĄzelo z 500 mg, tj. 1,04 mM3â-deoxy-6â-N-metylkanamycinu A ve formÄvolnĂ© bĂĄze a bylo pouĆŸito 595 mg, tj. 2,4mM N-benzyloxykarbonyloxysukcinimidu.
[a]035 +73° (c 1, voda â dimetylformamid, 1 : 2).
DalĆĄĂm zpracovĂĄnĂm takto pĆipravenĂ©slouÄeniny se zĂskal l-N-[ (S)-4-amino-2-hyd-roxybutyryl ] -3â-deoxy-6â-N-metyl-kanamy-cin A. PĆĂklad 18
PĆĂprava 3,6â-di-N-beinzytoxykarbonyl-4â-de-oxykanamycinu A
VychĂĄzeje z 500 mg, tj. 1,07 mM 4â-deoxy- kanamycinu A ve formÄ volnĂ© bĂĄze [viz âJournal of Antibioticsâ, Vol. 27, pp. 838â 46 â849 (1974); âBulletin of the Chemical So-ciety of Japan,â, Vol. 50, str. 2 362â2 368(1977)], byla zĂskĂĄna ĆŸĂĄdanĂĄ slouÄenina veformÄ bezbarvĂ© pevnĂ© lĂĄtky s vĂœtÄĆŸkem 666miligramĆŻ, tj. 71% stejnĂœm postupem jakov pĆĂkladu 15, pouze s tĂm rozdĂlem, ĆŸe 580miligramĆŻ, tj. 2,3 mM N-benzyloxykarbonyl-oxysukcinimidu rozpuĆĄtÄnĂ©ho ve 4 ml di-metylsulfoxidu bylo pomalu pĆidĂĄvĂĄno podĂ©le neĆŸ jednu hodinu k homogennĂmu roz-toku.
[a]u25 = +77° (c 1, voda â dimetylĂorm-amid, 1 : 2).
AnalĂœza
VypoÄteno pro Cb^HĂsNĂOis . 2 CH3lCO ,,H . η>Î: 52,16 Ξ/Îż C, 6,68 % H, 6,40 % N,Nalezeno: 51,77 % C, 6,79 % H, 6,31 % N.PĆĂklad 19
PĆĂprava 3,2*,6â-tri-N-benzyloxykarbonyl-kanamycinu B 500 mg, tj. 1,03 mM kanamycinu B veformÄ volnĂ© bĂĄze bylo suspendovĂĄno vesmÄsi 12 ml dimetylsulfoxidu a 4 ml tetra-hydrofuranu a k suspenzi byl pĆidĂĄn 1 g âtj. 4,55 mM octanu zineÄnatĂ©ho dihydrĂĄtu.SmÄs byla mĂchĂĄna pĆi teplotÄ mĂstnosti aĆŸdo vytvoĆenĂ homogennĂho roztoku a pakochlazena na 0 °C. Ke chladnĂ©mu roztokubyl pomalu pĆidĂĄvĂĄn dĂ©le neĆŸ jednu hodi--benzytoxykarbonyloxysukcinimidu rozpuĆĄ-nu chladnĂœ roztok 825 mg, tj. 3,3 mM N-tÄnĂ©ho v 10 ml tetrahydrofuranu â dime-tylsulfoxidu 1 : 1. VĂœslednĂĄ smÄs byla po-nechĂĄna v klidu pĆi 0 °C po dvÄ hodiny apak pĆi teplotÄ okolĂ pĆes noc, naÄeĆŸ bylasmÄs zpracovĂĄna stejnĂœm zpĆŻsobem jako vpĆĂkladu 1 (ii) a (iii) (A), pĆiÄemĆŸ se zĂskalo 740 mg, tj. 70 % ĆŸĂĄdanĂ© slouÄeninyve formÄ bezbarvĂ© pevnĂ© lĂĄtky.
[a]n25 â +63° (c 1, voda dimetyl â for-mamid 1:2).
AnalĂœza
VypoÄteno pro C42H.-,5N.-,O|6 . 2 CHjCCCH . H,O: 53,95 % C, 6,40 % h, 6,84 % N,
Nalezeno: 53,66 % C, 6,67 % H, 6,63 % N.
DalĆĄĂm zpracovĂĄnĂm takto pĆipravenĂ© slouÄeniny postupem podobnĂœm jako v pĆĂ- kladu 31 byl zĂskĂĄn l-N-[ (S)-4-amiiuo-2-hyd- roxybutyryljkanamycin B. 261853 47 48 PĆĂklad 20 PĆĂprava 3,2â,6â tri N bonzyk)<xykĆirl>i mylto-bramycinu 480 mg, tj. 1,03 mM tobramycinu v» for-mÄ volnĂ© bĂĄze bylo suspendovĂĄno ve 12 mldlmetylsulfoxidu a k suspenzi byl pak pĆi-dĂĄn 1 g, tj. 4,55 mM octanu zineÄnatĂ©ho di-hydrĂĄtu. SmÄs byla mĂchĂĄna pĆi teplotÄmĂstnosti po jednu hodinu do vytvoĆenĂ ho-mogennĂho roztoku, ke kterĂ©mu byl pakpĆidĂĄvĂĄn po asi jednu hodinu roztok 850miligramĆŻ, tj. 3,4 mM N-bcnzyloxykarbo-nyloxysukeinimidu, rozpuĆĄtÄnĂ©ho v 10 mlsmÄsi tetrahydrofuran-dimetylsulfoxid 1 : 1.SmÄs byla ponechĂĄna v klidu pĆes noc pĆiteplotÄ okolĂ, a takto, zĂskanĂœ reakÄnĂ roztokbyl zpracovĂĄn s velkĂœm objemem etylĂ©terustejnÄ jako v pĆĂkladu 1 (ii j pĆiÄemĆŸ vznik-la hustĂĄ sirupovitĂĄ lĂĄtka.
SirupovitĂĄ lĂĄtka byla dĂĄle zpracovĂĄnastejnĂœm zpĆŻsobem jako v pĆĂkladu 1 (iiij(A), avĆĄak za pouĆŸitĂ smÄsi voda â dioxan1 : 2 mĂsto 2 : 1, pĆiÄemĆŸ se zĂskalo 810 mg,tj. 78 % ĆŸĂĄdanĂ© lĂĄtky ve formÄ bezbarvĂ©pevnĂ© lĂĄtky.
[a]u25 = 1-65° je 1, voda â dimetylEorma-mid, 1:2].
AnalĂœza VĂœpoÄtem» pro CĂł,HrĂłN ,Or, . 2 CH-CO-dl . H-.O: 54,81 o/o C, 6,50 % II, 6,0> % N,
Nalezeno: 54,77 % C, 6,71 % II, 6,88 % N.
Takto zĂskanĂĄ slouÄenina muĆŸe bĂœt dĂĄlezpracovĂĄna postupem podobnĂœm jako vpĆĂkladu 31 za vzniku l-N-[(S)-4-amino-2--hydroxybutyryl ] -tobramycinu. PĆĂklad 21 PĆipravĂ! 3,2â,6â-tri-N -benzyloxykarbonyl-G1--N-metyltobramycinu ĆœĂĄdanĂĄ slouÄenina ve formÄ bezbarvĂ©pevnĂ© lĂĄtky byla zĂskĂĄna s vĂœtÄĆŸkem 890miligramĆŻ, tj. 84 % stejnĂœm postupem jakov pĆĂkladu 20 s tĂm rozdĂlem, ĆŸe se vychĂĄ-zelo z 500 mg, tj. 1,04 mM Bâ-N-metyltobra-mycinu ve formÄ volnĂ© bĂĄze.
[a]D25 ~ -I 63° (c 1, voda -- dimetylforma·mid, 1 : 2). PĆĂklad 22
PĆĂprava 3,2â,6â-tri-N-benzyloxykarbonyI-4â--deoxykanamycinu B
VychĂĄzeje z 480 mg, tj. 1,03 mM 4-deoxy- kanamycinu B ve formÄ volnĂ© bĂĄze [viz âBulletin of the Chemical Society of Japanâ,Vol. 50, pp. 2 362--2 368 (1977)], byla zĂs-kĂĄna ĆŸĂĄdanĂĄ slouÄenina ve formÄ bezbar-vĂ© pevnĂ© lĂĄtky s vĂœtÄĆŸkem 815 mg, tj. 79proÄ., stejnĂœm postupem jako, v pĆĂkladu20.
[«]i,-5 463° (c 1, voda â dimetyiformamiĂș, 1 : 2], PĆĂklad 23 PĆĂprava 3,2â,6â-tri-N-benzyloxykarbonyl-dibekacin 000 mg, tj. 1,33 mM dibekacinu (3â,4â-di-deoxykanamycinu B] ve formÄ volnĂ© bĂĄzebylo suspendovĂĄno v 15 ml dimetylsulfoxi-du a suspenze byla mĂchĂĄna aĆŸ do- vytvoĆe-nĂ roztoku, ke kterĂ©mu bylo pĆidĂĄno 1,4 g,tj. 6,4 mM dihydrĂĄtu octanu zineÄnatĂ©ho,naÄeĆŸ se v mĂchĂĄnĂ pokraÄovalo. K vĂœsled-nĂ©mu roztoku se pomalu pĆidĂĄval po asijednu hodinu roztok 1,1 g, tj. 4,4 mM N--benzylnxykarbonyloxysukcinimidu ve 12 mldimetylsulfoxidu a smÄs byla ponechĂĄna vklidu pĆi teplotÄ okolĂ pĆes noc. Pak byl sreakÄnĂm roztokem smĂchĂĄn velkĂœ objemetylĂ©teru, aby se oddÄlila olejovitĂĄ sedlina,obsahujĂcĂ zejmĂ©na N-benzyloxykarbonylo-vanĂœ zineÄnatĂœ komplex jako ĆŸĂĄdanĂœ pro-dukt a ÄĂĄst dimetylsulfoxidu, a pak bylapromyta etyiĂ©terem, ÄĂmĆŸ se zĂskal sirupo-vitĂœ materiĂĄl.
Tato sirupovitĂĄ lĂĄtka byl opakovanÄ pro-myta vodou, ÄĂmĆŸ se rozpadl N-acylovanĂœzineÄnatĂœ komplex a volnĂ© kationty zinkubyly odstranÄny spolu s pĆŻvodnÄ existujĂ-cĂm nadbytkem c-ctanu zineÄnatĂ©ho. Taktobylo zĂskĂĄno 1,1 g ve vodÄ nerozpustnĂ© pev-nĂ© lĂĄtky, pĆedstavujĂcĂ N-acylovanĂœ dibeka-cin. PevnĂĄ lĂĄtka byla chromatograf ovĂĄnana tenkĂ© vrstvÄ silikagelu za pouĆŸitĂ smÄsichloroform â etanol â 18% vodnĂœ Äpa-vek 1:1:1, spodnĂ fĂĄze, jako vyvĂjecĂhorozpouĆĄtÄdla, pĆiÄemĆŸ se vytvoĆila jedinĂĄskvrna pĆi Rf 0,3, coĆŸ znamenĂĄ, ĆŸe pevnĂĄlĂĄtka byla tvoĆena pĆevĂĄĆŸnÄ 3,2â,6â-tri-N--benzyloxykarbonyldibekacinem se stopouzinku.
DalĆĄĂm zpracovĂĄnĂm ĆŸĂĄdanĂ© slouÄeninypostupem podobnĂœm jako v pĆĂkladu 31 sezĂskal l-N-[[S)-4-amino-2-hydroxybutyryl]--dibekacin.
DalĆĄĂ purifikace surovĂ©ho produktu zĂs-kanĂ©ho- postupem uvedenĂœm vĂœĆĄe, byla pro-vĂĄdÄna tak, ĆŸe slouÄenina byla promĂœvĂĄnaroztokem 3 M Äpavku, ÄĂmĆŸ se zĂskal pro-dukt neobsahujĂcĂ pĆĂmÄsi iontĆŻ zinku.
[q:]d25 +71° (c 1, voda â dimetylformamid, 1 : 2). PĆĂklad 24 PĆĂprava 3,2â,6â-trl-N-benzyloxykarbonyl-6â- -N-metyldibekacinu 267033 49 500 mg, tj. 1,07 mM 6â-N-metyldibekacinuve formÄ volnĂ© bĂĄze a 1,2 g, tj. 5,45 mM di-hydrĂĄtu octanu zineÄnatĂ©ho bylo rozpuĆĄtÄ-no! ve 20 ml dimetylsulfoxldu, a k tomu by-lo pomalu pĆidĂĄvĂĄno po asi 30 minut 910miligramĆŻ, tj. 3,6 mM N-benzyloxykarbonyl-oxysukcinimidu. ReakÄnĂ roztok byl pone-chĂĄn stĂĄt v klidu pĆi teplotÄ okolĂ pĆes noca pak byl zpracovĂĄn stejnĂœm zpĆŻsobem ja-ko v pĆĂkladu 23, ÄĂmĆŸ se zĂskalo 910 mg ĆŸĂĄ-danĂ© slouÄeniny, jeĆŸ byla v podstatÄ ÄistĂĄ.
DalĆĄĂm zpracovĂĄnĂm takto zĂskanĂ© slou-Äeniny se zĂskal l-N-[ (S j-4-amino-2-hydro-xybutyryl ] -6â-N-metyldibekacin. PĆĂklad 25
PĆĂprava 3,2â-di-N-benzyloxykarbonylkana-mycinu C ĆœĂĄdanĂĄ slouÄenina ve formÄ zbarvenĂ©pevnĂ© lĂĄtky byla zĂskĂĄna s vĂœtÄĆŸkem 730miligramĆŻ, tj. 79 % stejnĂœm postupem ja-ko v pĆĂkladu 1 (i), (ii) a (iiij (A), s tĂmrozdĂlem, ĆŸe se vychĂĄzelo z 500 mg, tj. 1,03mM kanamycinu C ve formÄ volnĂ© bĂĄze.
[«ji,-5 +75° (c 1, voda â dimetylformamid, 1 : 2). DĂĄle byla takto zĂskanĂĄ slouÄenina zpra-covĂĄna, pĆiÄemĆŸ byl zĂskĂĄn l-N-[(Sj-4-ami-no-2-hydroxybutyryl]-kanamycin C.PĆĂklad 26
PĆĂprava 6â-N-benzyloxykarbonylkanamyci-nu A 500 mg, tj. 1,03 mM kanamycinu A ve for-mÄ volnĂ© bĂĄze bylo suspendovĂĄno ve 20 mldimetylsulfoxidu a k suspenzi bylo, pĆidĂĄ-no 0,5 g, tj. 2,3 mM octanu zineÄnatĂ©ho di-hydrĂĄtu. SmÄs byla mĂchĂĄna pĆi teplotÄmĂstnosti aĆŸ do vytvoĆenĂ homogennĂho roz-toku, ke kterĂ©mu pak bylo pĆidĂĄno 283 mg,tj. 1,13 mM N-benzyloxykarbonytoxysukcin-imidu. VĂœslednĂĄ smÄs byla ponechĂĄna v kli-du pĆes noc pĆi teplotÄ okolĂ a pak bylazpracovĂĄna stejnĂœm zpĆŻsobem jako v pĆĂ-kladu 1 (ii) a (iiij (Ij ÄĂmĆŸ se zĂskalo 556miligramĆŻ ĆŸĂĄdanĂ© slouÄeniny ve formÄ bez-barvĂ© pevnĂ© lĂĄtky.
[a]D2R =, +g2« (C 1, voda). PĆĂklad 27 PĆĂprava 6â-N-benzyloxykarbonyldibekacinu
StejnĂœm postupem jako v pĆĂkladu 26, bylo zĂskĂĄno 382 mg ĆŸĂĄdanĂ© slouÄeniny za 80 pouĆŸitĂ 500 mg dibekacinu ve formÄ volnĂ©bĂĄze, 12 ml dimethylsulfoxidu, 0,7 g octanuzineÄnatĂ©ho dihydrĂĄtu a 305 mg N-benzyl-oxykarbonyloxysukcinimidu.
[ ce]D25 +105° (c 0,5, voda). PĆĂklad 28
PĆĂprava 3,2â,6â-tri-N-benzyloxykarbonyl--3â,4â-dideoxy-3â-enokanamycinu B 500 mg, tj. 1,11 mM 3â,4â-dideoxy-3â-eno-kanamycinu B ve formÄ volnĂ© bĂĄze (vizâBulletin of the Chemical Society of Ja-panâ, Vol. 50, ppd 1 580â1 583 (1977)] by-lo rozpuĆĄtÄno· ve 12 ml dimetylsulfoxidu ak roztoku byl pĆidĂĄn 1 g, tj. 4,55 mM di-hydrĂĄtu octanu zineÄnatĂ©ho, a roztok bylmĂchĂĄn po jednu hodinu. K vĂœslednĂ©mu roz-toku bylo pĆidĂĄvĂĄno pomalu pĆes 30 minut870 mg, tj. 3,49 mM N-benzylotxykarbonyl-oxysukcinimidu. SmÄs byla ponechĂĄna v kli-du pĆes noc pĆi teplotÄ okolĂ a reakÄnĂ roz-tok takto zĂskanĂœ byl zpracovĂĄn s velkĂœmobjemem etylĂ©Ăeru stejnÄ jako v pĆĂkladu 1(ii) pĆiÄemĆŸ se zĂskala hustĂĄ sirupovitĂĄ lĂĄt-ka.
SirupovitĂĄ lĂĄtka byla dĂĄle zpracovĂĄnastejnÄ jako v pĆĂkladu 1 (iiij (Bj, avĆĄak zapouĆŸitĂ smÄsi voda â dioxan 1 : 2 mĂsto·2 : 1, ÄĂmĆŸ se zĂskalo· 784 mg ĆŸĂĄdanĂ© slou-Äeniny ve formÄ bezbarvĂ© pevnĂ© lĂĄtky.
[a]D25 + 30° (c 1, voda â dimetylformamid, 1 : 2). PĆĂklad 29 PĆĂprava 3,2â,6â-tri-N-benzyloxy-karbonylsi-somicinu ĆœĂĄdanĂĄ slouÄenina ve formÄ bezbarvĂ©pevnĂ© lĂĄtky byla zĂskĂĄna s vĂœtÄĆŸkem 780miligramĆŻ stejnĂœm postupem jako v pĆĂkla-du 28, avĆĄak s tĂm rozdĂlem, ĆŸe se vychĂĄze-lo z 500 mg, tj. 1,12 mM sisomicinu ve for-mÄ volnĂ© bĂĄze.
[ ofjir5 = +110â (c 1, voda â diraetylforma-mid, 1:2). PĆĂklad 30 PĆĂprava 3,2â,6â-tri-N-benzyloxykarbonyl-gentamicinĆŻ 787 mg ĆŸĂĄdanĂ© slouÄeniny bylo zĂskĂĄnove formÄ bezbarvĂ© pevnĂ© lĂĄtky stejnĂœmpostupem jako v pĆĂkladu 28, s tĂm rozdĂ-lem, ĆŸe se vychĂĄzelo z 500 mg smÄsi genta-micinĆŻ C, Cla, C2 atd.
CZECHOSLOVAK SOCIALISTIC
(19)
DESCRIPTION OF THE INVENTION
PATENT 261853 (Î ) (B2)
INVENTORY OFFICE DISCOVERY (22) Enrolled 12 11 79 (21) (PV 7711-79) (32) (31) (33) Priority 11 11 78 (138402) Japan (51) Int. Cl.4C 07 H 15/234 (40) Published 15 06 88 (45) Published 15 07 89 (72)
The author of UMEZAWA HAMAO, UMEZAWA SUMIO, TOKIO, TSUCHIYA TSUTOMU, TAKAGI YASUSHI, JIKIHARA ΀ÎÎÎ, KANAGAWA (Japan) (73)
Patent holder ZAIDAN HOJIN BISEIBUTSU KAGAKU KENKYU KAI, TOKIO (Japan) (54) Method for the production of selectively acylated N-targeted derivative of aminoglycoside antibiotic 1
The present invention relates to a process for the preparation of a selectively acylated N-protected derivative of aminoglycoside antibiotic.
In particular, the present invention relates to a novel process for the production of a selectively protected N-acyl derivative aminoglycoside dibiotic, in which some amino acids or alkylamino groups in certain positions. aminoglycoside molecules are selectively protected or blocked by an acyl group. The invention therefore relates to a novel method for the selective protection of certain amino or alkylamino groups at certain positions of the aminoglycosidic anti-biotics and finds a major use in the manufacture of a selectively protected N-acylated aminoglycoside derivative having a deoxystreptamine structure comprising 3 ' an aminoglycosyl moiety linked to the 6-hydroxy group of the deoxystreptamine moiety in the aminoglycoside molecule.
The aminoglycoside antibiotic useful in the present invention can be precisely defined as an aminoglycoside anti-biotic consisting of 6-O- (3 "-amino- or 3" -alkylamino-3 "-deoxyglycosyl) -2-deoxy-streptainine, which may optionally contain a 4-O- (6'-aminoglycosyl) substituent. Typical examples are kanamycins, gent-2 amicins, sisomicin, netilmicin and verdamycin.
It is known that aminoglycoside antibiotics, such as kanamycins, are substances containing several amino and hydroxy groups that have a relatively high and different degree of reactivity. Many types of polysynthetic aminoglycoside antibiotics that are derived from the original aminoglycoside antibiotics have been synthetically synthesized. In the semisynthesis of these derivatives, it is often necessary or advantageous to ensure that some amino groups and / or some hydroxy groups in the starting aryoglycoside antibiotic are selectively protected by at least one suitable protecting group.
Various useful methods have been developed for the selective protection of amino and / or hydroxy groups in the aminoglycosideantibiotic, which as such are useful for the selective protection of the hydroxy group. However, the pro-selective protection of some selected amino groups from a number of amino groups that are in the aminoglycoside antibiotic, these known methods are either difficult to perform or require some complex operations. This is due to the fact that all amino groups in the aminoglycoside antibiotic 261853 281853 '' rozdĂl do not have a larger difference in reactivity. A typical example is the 6'-amino group of kanamycin A, but such an amino- or methylamino group that is bound to a particular carbon atom, which in turn is bound to only one carbon atom in the amino-glycoside molecule, produces a higher reactivity than an amino- or methylamino-group that is bonded to a carbon atom that is bonded to two or more carbons in that amino-glycoside molecule. For this reason, the first type of amino- or methylamino group is much more capable of reacting with an acylating agent that contains an acyl group intended to be introduced as an amino-protecting group than the latter type of amino- or methyl-amino group so that the N-protected derivative having the first type of amino- or methylamino group preferably protected by an acyl group may be produced with a higher yield than otherwise N-protected derivatives. A few years ago, some of the inventors of the present invention have found that when amino and hydroxy groups are adjacent to each other in pairs in the steric configuration of the aminoglycoside antibiotic molecule, the amino and hydroxy groups may be selectively combined with each other to form a cyclic carbamate by reaction with sodium hydride so that a pair consisting of an amino group and a hydroxy group can be protected simultaneously in the cyclic carbamate by the non-blocking of other amino groups present in the same molecule [see Journal of Antibiotics 25, 12, 741-742 [1972], -ts Nos. 3 925 354 and 3 965 0B9],
Recently, Nagabhushan et al. found that when a divalent transition metal salt (M + + J selected from divalent copper, nickel, cobalt, and cadmium is reacted with an aminoglycoside antibiotic in the 4-0-- (aminoglycosyl) group in an inert organic solvent) ) -6-O- [aminoglycosyl) -2-deoxystreptamine, represented by kanamycins, gentamicins and sisomicin, this divalent transition metal cation is complexed with a pair consisting of an amino group and a hydroxy group, which are especially in the conjugate configuration in an aminoglycoside molecule thereby forming a transition metal cationic complexaminoglycoside antibiotic (Japanese Patent Publication No. Ser-52-153 944 and US Patent No. 4,136,254, issued Jan. 23, 1979 J. In this cationic with the transition metal complex of the amino-glycoside antibiotic, the complexed amino group is protected by a divalent transition cation. with an acyl-containing acylating agent, only non-complexed amino groups which are not protected by the divalent metal cation can be acylated to the complex, so that a selective N-protection with an acyl group is achieved. This is shown below with reference to kanamycin A as an example. If the divalent transition metal cation (M + +) is selected from the group consisting of bivalent copper, nickel, cobalt and cadmium, it is reacted with kanamycin A, the complexation reaction of the divalent co-cation (M + + J occurs between the 1-amino group of a 2) "-Hydroxy group and between the 3" -amino group and the 4 "-hydroxy group of the canancin A molecule, as shown by formula ec:
Thus, in the above-mentioned complexation reaction, it is clear that at least 2 moles of the transition metal per 1 mol of kanamycin A are required. In the resulting metal complex, the 1-amino and 3 ' -amino groups are blocked simultaneously. When this complex of formula I * is treated with an acylating agent containing a macyl group which is suitable for protecting an amino group, in the usual synthesis of polypeptides, only the uncomplexed 3-amino- and 6'-amino groups are acylated to give 3,6 ' di-N-acylated derivative [Journal of American Chemical Society, 100, 5 253-5254 [1978J],
The present inventors have taken into account the above, but have conducted further in-house investigations of the interaction of other diverse metal cations with aminoglycoside antibiotics such as kanamycin A and kanamycin B, as well as semi-synthetic derivatives of aminoglycoside antibiotics. As a result, although the cation of divalent zinc exhibits substantially different cationic behavior from the above set comprising divalent cations of nickel, cobalt, copper and cadmium, the zinc cation is capable of complex complexing and protecting both the 1-amino-or 1-alkylamino and 3 "-amino or 3" -alkylamino group of aminoglycoside compounds, for example kanamycin A, B or C, which contains a deoxystrepta-261853 mine moiety comprising a 3 "-aminoglycosyl group or a 3" -alkylaminoglycosyl moiety a group attached to the 6-hydroxy group of said deoxystreptamine moiety.
According to Nagabhushan et al. one would expect that when the cation of divalent nickel, divalent cobalt, divalent copper or divalent cadmium is reacted with kanamycin, for example. B, a metal complex salt of cannabin B of formula II should be formed
This assumption can be underpinned by Nagabhushana et al. in the above-mentioned "American ChemicalSociety Journal", according to which pairs of vicinal amino-hydroxy groups could form versatile complexes with divalent transition cations due to the fact that kauamycin B contains three pairs of initial aminohydroxy groups between position-mi 1 and 2 ", between the positions 2 'and 3' and between the positions 2" and 3 "of the kanamycin B molecule.
However, it has now been found that when secanamycin B is reacted with a zinc cation, the cananamin B complex formed with the zinc salt contains free 2'-amino- and 3'-hydroxy groups which are not blocked by the zinc cation, as opposed to the assumption Nagabhushana et al. Although the complexing reaction of the zinc cation with the 2'-amino- and 3'-hydroxy group occurs, the complex formation force is great, so that in practice the 2'-amino- and 3'-hydroxy groups are not blocked. Thus, when the kanamycin B complex with zinc cation is acylated by reaction with N-benzyloxycarbonyloxy succinimide to introduce a benzyloxycarbonyl group as an amino-protecting acyl group, a tri-3,2 ', 6'-N-acylated derivative is formed in which the 3-amino, 3-, 2'- and 6'-amino groups have been acylated, in fact at a higher yield than the otherwise N-acylated derivatives, but can then in fact not be obtained 3 , 6'-di-N-acylated derivative (see Example 19J below. This experimental fact shows that zinc cation has a different behavior to the above-mentioned cations of the four transition metals, especially in that zinc cation does not form complexes) with the vicinal pair of the 2'-amine- and the 3'-hydroxy group.
As another example, when kanamycin A is reacted with a zinc cation followed by acylation with a benzyloxycarbonyl group (referring to formula T above), the major acylation product is observed to form 3,6'-di-N-benzyloxycarbonylkanamycin And, if the zinc cation is present in an amount slightly above 1 mole per mole of cancanine A. In this case, it is noted that this acylation reaction produces 1,3, -6 ', 3 "-tetra- N-benzyloxacarbonyl derivative alkamycin A and, at the same time, non-acylated starting kanamycin A but at the same time producing a tri-N-benzyloxycarbonylene derivative of kanamycin A with only a low yield, although the mechanism of reaction of Nagabhushan et al. that the tri-N-benzyloxycarbonyl derivative is formed with a higher yield than other N-acylated derivatives (see Example 7 below). U.S. Pat. No. 4,136,254 to Nagabhu-sahan et al. Have reported that the divalent divalent transition metal, e.g., copper, nickel, cobalt, etc., must be used in a total amount of at least 2 moles per mole of kanamycin A to form a complex kanamycin salt A with a transition metal as shown in Formula I * above.
The present inventors have shown that a woman, in contrast to the cationic metals of the four transition metals, is able to achieve the blocking of the 1-amino- and 3 ' -amino group of kanamycin when the cation is used in a total amount of at least 1 molar per 1 mole of kanamycin A. It was further found that when a nickel salt was used in the reaction at a rate of less than 1 mole per mole of kanamycin A, followed by acylation of the resulting complex of the kanamycin A salt with nickel by a benzyloxycarbonyl group, only a very small amount yielding 3,6'-di-N-benzyloxycarbonyl-kanamycin A, which compound could be obtained with significant yield of cyanide complex of kanamycin A with zinc (see Example 7 below). From the foregoing, it has been concluded that the cesium cation creates a mechanism for the formation of a complex with some aminoglycoside, which is a different mechanism of complex formation with the divalent nickel, cobalt, copper and cadmium complex and that the cationic complex of the zinc zinc aminoglycoside has a stabilizing complex, which is different from the stability of the aminoglycoside caustic complex with divalent nickel, cobalt, copper or cadmium. For complexation of the zinc cation with the aminoglycoside antibiotic 261853 7, the zinc cation may be used in the form of a zinc salt which has the advantage of being inexpensive and probably not a source of environmental pollution.
Therefore, the present inventors have found that when the zinc cation is reacted in an inert organic solvent with an aminoglycoside antibiotic containing a deoxystreptamine moiety comprising a 3-aminoglycosyl or 3-alkylaminoglycosyl moiety linked to the 6-hydroxy group of the deoxystreptamine moiety optionally comprising an aminoglycosyl group linked to the 4-hydroxy group of the deoxy-streptamine moiety, the zinc cation forms a complex with aminohydroxyl group pairs located at certain positions, depending on the properties of the aminoglycoside antibiotic, when the cationic complex of the aminoglycoside antibiotic with seasoning is reacted with an acylating agent containing an acyl group usually used to introduce a protective amino group in the synthesis of the polypeptides, the acylating agent acylating at least one of the amino amino groups in the aminoglycosides mantibiotics that do not form a complex and are therefore not blocked by the zinc cation so that the acylated amino group is protected, and further that when the resulting acylation product, i.e., the cationic complex of the aminoglycoside antibiotic with zinc containing the acylated amino or acylated amino groups is reacted with a suitable zinc cation removing agent from this acylation product, the zinc complex is disrupted to provide a selectively protected N-acylated amino-glycoside derivative of the zinc-uncomplexed amino group or amino groups have been selectively protected with an acyl group.
The invention thus provides a process for the preparation of a selectively acylated N-protected derivative of an aminoglycoside antibiotic.
The present invention provides a process for the production of a selectively acylated N-protected derivative of an aminoglycoside antibiotic comprising 4-O- (aminoglycosyl) -6-O- (3 "-amino-or 3" -methylamino-3 "-deoxyglycosyl) -2-deoxystreptamine in which the 1-amino- and 3-amino groups are unprotected, but all other amino groups are protected with an amino-protecting acyl group of formula I wherein R 'is hydrogen or ethyl, G is formyl , C 2 -C 5 alkanoyl, C 2 -C 5 trifluoroalkanoyl, C 1 -C 4 alkoxycarbonyl, 1 to 4 carbon phenyloxycarbonyl, 1 to 4 carbonyl phenylalkyloxycarbonyl or C 1 -C 4 methoxyphenylalkoxycarbonyl groups. 4 carbon atoms of alkoxy, Q 1 represents an N-protected aminoglycosyl group of the formula IIIa
W is hydroxy or N-protected amino -NHG wherein G is as defined above, X is hydrogen or hydroxy, Y is hydrogen or hydroxy, Z 'is hydrogen, hydroxy, N -protected amino group of formula -NHG or N-protected alkylamino group of formula R '
âN
G in which G is as defined above and R 'is methyl, Z' is a hydrogen atom or a methyl group, or QL is an N-protected 3 ', 4'-dideoxy-3' - eno-aminoglycosyl group of the general formula IIIa
261833 10 wherein G is as defined above, or Q 1 is an N-protected 3 ', 4'-dideoxy-4'-one-aminoglycosyl group of the general formula IVa
R "-CH-NHG
wherein R '' is a hydrogen atom or a methyl group and G is as defined above, is a 3 "-amino-3" -deoxy dycosyl group of the formula Va
wherein M is hydroxy or hydrogen and M; is hydroxy or hydrogen, or Q2 is a 3 ' -methylamino-3 ' -deoxyglycosyloxy group of the formula VIa.
in which
R 'represents a hydrogen atom or a methyl group, wherein the zinc cation salt with an inorganic or organic acid is reacted with a self-glycotsid antiblotics of formula VII wherein R' is hydrogen or ethyl, Q 3 is an aminoglycosyl group of general formula IIb
wherein W 'is hydroxy or amino, X is hydrogen or hydroxy, Y is hydrogen or hydroxy, Z''is hydrogen, hydroxy, amino or methylamino of the formula --NHR, wherein R 'is methyl, Z' is hydrogen or methyl or Q 3 is 3 ', 4'-dideoxy-3'-eno-aminoglycosyl group IIIb
or 'Q3 is a 3', 4'-dideoxy-4'-eno-aminoglycosyl group of formula IVb 261853 11
wherein R '' represents a hydrogen atom or a methyl group and Q 4 represents a 3 "-amino-3" -deoxyglycosyl group or a 3 "-methylamino-3" -deoxy-glycosyl group identical to the above-mentioned group Q 2 of the general formula And / or V1 at a molar ratio of at least 1 part molar, preferably 2 to 6 parts molar zinc cation salt with an inorganic or organic acid per 1 molar aminoglycoside antibiotic of formula VII at a temperature between -10 ° C. and 100 ° C in an inert organic solvent selected from dimethyl sulfoxide, aqueous dimethyl sulfoxide, dimethylformamide, aqueous dimethylformamide, a mixture of dimethylsulfoxide and dimethylformamide, tetrahydrofuran, aqueous tetrahydrofuran, methylene, aqueous methanol, ethanol and ethanol, optionally in the presence of sodium acetate to form the cationic complex of the aminoglycoside antibiotic with the seed, then the cationic complexaminoglycoside antibiotic is zinc it may be reacted with an acylating agent selected from the group consisting of the carboxylic acid of the formula IVa RRCOOH (IVa) wherein R5 represents a hydrogen atom, an alkyl group having 1 to 4 carbon atoms, a trifluoroalkyl group having 1 to 4 carbon atoms, or with the halide, anhydride or active ester of the aforementioned carboxylic acid of formula IVa, the chloroformate of formula IVb R6O-CO-Cl (IVb) p-nitrophenyl carbonate of formula IVc RfO-CO-O-C6H5-p-NO ·, ( IVc) active N-hydroxysuccinimide ester of formula IVd 12 0 0 (IVd) and azidoformiate of formula IVeReO â CO â N 3 (IVe) in which samples
R 6 is as defined above and R 6 is C 1 -C 4 alkyl, phenyl, C 1 -C 4 alkyl, or C 1 -C 4 p-methoxyphenylalkyl; C, for the acylation of uncomplexed amino groups present in the cationic complex of the aminoglycoside antibiotic with zinc and thus to form a cationic complex of the N-acylated aminoglycoside antibiotic with zinc, and the zinc cationic complex of the N-acylated aminoglycoside antibiotic is then left with water or with an aqueous or non-aqueous polar organic solvent selected from methanol, ethanol, liquid ammonia, ethylamine and triethylamine, or with hydrogen, alkali metal sulfide or alkaline earth metal or water ammonium hydroxide or cation exchange resin containing carboxylic or sulfonic acid functions or an anion exchange resin containing ammonium functions, or a chelating agent containing metal chelating functions or with chitin or chitosan as a water-insoluble higher polymer comprising a compound capable of combining, at a temperature between -10 and 100 ° C, removing the zinc cations from the complex and forming an N-acylated amino glycoside antibiotic of formula I.
The process of the invention is suitable for the production of a selectively acylated N-protected aminoglycoside antibiotic derivative of cyano of some amino groups other than the 1- and 3 ' -amino groups of the starting aminoglycoside antibiotic, and such selectively N-protected derivative is a suitable prochemical the synthesis of 1-N-amino acylated derivatives of aminoglycoside antibiotics, for example kanamycin C 1, including amikacin ("Journal of Antibiotics" 25, 695-708 (1972)], which has been shown to be an effective antibacterial drug in recent years 261853 13 11. N-aminoacylated derivatives of aminoglycoside antibiotics include numerous derivatives of aroinoglycosides, such as kanamycin A, cananycin B, kanamycin C, gentamicins, sisicin, and others, as well as their various deoxy derivatives, but all of which are consistent with that their J-amino group is acylated with some -hydroxy-Ï-aminoalkanoyl group (see U.S. Pat. Nos. 3,781,268, 3,939,143.3,940,382, and 4,001,208). Due to this 1-N-aminocacylation, aminoglycoside antibiotics receive antibacterial activity against resistant bacteria against which the current antibiotics are not effective and these aminoglycoside antibiotics also obtain improved antibacterial activity against broader bacterial areas compared to existing aminoglycoside antibiotics.
The method of the present invention will now be described in more detail.
The aminoglycoside antibiotic to be reacted with a zinc cation to form a zinc complex, which may also be referred to as the zinc complex salt of the present invention, includes such aminoglycoside antibiotics containing a deoxy-streptamine structure whose 6-hydroxy group is substituted with 3-amino -glycosyl or 3-alkylaminoglycosyl group, and whose 4-hydroxy group may be optionally substituted with one of the aminoglycosyl groups. More specifically, the aminoglycoside anti-biotic used in the present invention to form a zinc cation complex may be defined as containing 6-O- (3 "-amino- or 3" -alkylamino-3 "-deoxyglycosyl j -2- In addition, the aminoglycoside antibiotic may be any 1-N-alkylaminoglycoside, for example netilmicin, as examples of aminoglycoside antibiotics useful in the present invention. of the invention can be mentioned a group of kanamycin A antibiotics, such as kanamycin A, 6 * -N-alkyl-kanamycin A, especially 6'-N-methylkanamycin A, 3'-deoxycanamycin A, 6'-N-methyl-3'-deoxycanamycin A, 4'-deoxycanamycin A, 6'-N-methyl-4'-deoxycanamycin A, 3 ', 4'-dide-oxycanamycin A (see Japanese Patent Application Nos. 11 402/79), and 6 "- deoxy- or 4 ", 6" -dide-oxycanamycin A (see Japanese Patent Application No. 34 733/79), a group of kanamycin B antibiotics, i.e. kanamycin B alone 3'-deoxy-kanamycin B (i.e. tobramycin), 4'-deoxy-kanamycin B, 3 ', 4'-dideoxycanamycin B (i.e. dibecacin), 3', 4'-dideoxy-3'-eno-kanamate -cin B. 6'-N-methyl-3 ', 4'-dideoxycanamycin B, a group of kanamycin C antibiotics, that is, kanamycin C, 3'-deoxycanaraycin C, 3', 4'-dideoxycanamycin C, gentamicins A , B and C, verdamicin, sisomicin and netilmicin (i.e., 1-N-ethylsisomycin), as well as other known aminoglycosides.
The method of the first idea of the present invention is applicable not only to so-called aminoglycoside antibiotics, which are not yet known and will be discovered in the future, but also to novel semisynthetic aminoglycoside derivatives of antibiotics which will in future be manufactured by chemical transformation of known amino-glycoside antibiotics .
Typical examples of aminoglycoside antibiotics for use in the present invention are kanamycin A, kanamycin B, kanamycin C, deoxy derivatives of such cancers, and their 6'-N-alkyl derivatives, which are defined by this general formula.
wherein R 1 is hydroxy or amino, R 2 and R 3 are each hydrogen or hydroxy, and R 4 is hydroxy or amino or alkylamino containing alkyl of 1 to 4 carbon atoms, especially methylamino.
In order to form the zinc cation aminoglycoside antibiotic complex by reacting the zinc cation aminoglycoside antibiotic of the present invention, the corresponding aminoglycoside antibiotic, either in the form of its free base or in the form of its acid addition salt, is dissolved or suspending in a suitable organic solvent and adding the appropriate zinc salt to at least 1 mole per mole of the aminoglycoside antibiotic used. Any conventional organic solvent may be used if the zinc complex formed after the addition of the zinc salt is at least partially soluble therein. Preferably, however, the large volume of polar organic solvent and, in particular, the larger volume of water should be avoided since the presence of a polar organic solvent and water would reduce the stability of the resulting complex of the amino-glycoside antibiotic and zinc cation so that the subsequent acylation reaction would introduce 261853 13 The protective amino group could give a disappointing result.
Thus, it is desirable to use an organic solvent having a high solubility, for example dimethyl sulfoxide as the solvent in which the zinc complex is to be formed, but it is also possible to use aqueous dimethyl sulfoxide, dimethylformamide, aqueous dimethylformamide, a mixture of dimethylsulfoxide and dimethylformamide, tetrahydrofuran, aqueous tetrahydrofuran, as well as lower alkanol as methanol, ethanol and aqueous methanol.
The zinc cation may be added in the form of a zinc salt to the system in which the complex is formed. For this purpose, a zinc salt formed by the reaction of zinc cation with a conventional organic or inorganic acid can be used in the present invention. However, it is generally preferred to use a zinc salt of some weak acid, for example zinc acetate, since it is common that among the metal complexes containing an amino group, the non-quaternary amine group complex with the metal salt is more stable than the ammonium-type complex of the metal salt, and that the use of a zinc salt and a weak acid does not normally result in the formation of a relatively unstable complex of aminammonium-containing metal. If a zinc salt of an acid, such as zinc chloride, is used, the desired complexzinc can also be formed, but it is preferable to add a weakly alkali salt, for example sodium acetate, to the salt to neutralize the reaction medium. Similarly, it is desirable to add a certain amount of sodium acetate or sodium hydroxide as a neutralizing agent when the starting aminoglycoside antibiotic is used in the form of its additive salt with a strong acid, for example hydrochloric acid. In this case, the use of an unnecessary excess of neutralizing agent should be avoided, as otherwise the precipitation of the hydroxybenzoate will be impaired, thereby compromising the formation of the complex. For example, when the aminoglycoeide antibiotic tetrahydrochloride is used to form a complex, preferably 4 moles of sodium hydroxide are added to neutralize the reaction mixture.
If the total molar amount of zinc used is at least equal to the molar amount of the aminoglycoside antibiotic, the complex formation reaction can take place. However, it is preferred to use zinc salts in a quantity substantially greater than 1 mole per 1 mol of aminoglycoside antibiotic, so that the reaction to form the complex is shifted in favor of complex creation. A favorable yield of zinc complex can be obtained by using zinc salt in an amount of 2.3 to 6 moles per mole of aminoglycoside, but in the case of zinc salts 4 to 5 moles per mole of aminoglycoside is most preferred. The time required for the complete complex formation reaction after the addition of the zinc salt can vary depending on the organic solvent used and can be "instantaneous" (using an aqueous organic solvent) up to 20 hours. The complex formation reaction may normally take place at room temperature, but heating or cooling may be performed.
In this way, a solution or suspension containing a zinc complex and an amino-glycoside antibiotic is prepared to which an acylating agent having an acyl group for addition to the complex is added as a protective amino group.
The acylating agent used in the process of the present invention may be a conventional amino protecting agent, and is used to allow free, complex non-forming amino groups in the resulting complex of aminoglycoside antibiotic and zinc cation to be acylated and blocked and acyl group acyl group.
The acylating group may be an alkanoyl group, an aroyl group, an alkoxycarbonyl group, an aralalkoxycarbonyl group, an aryloxycarbonyl group, an alkylsulfonyl group, an arylalkylsulfonyl group, or an arylsulfonyl group, which are all common amino protecting groups.
The acylating agent useful for this radical may also be a carboxylic acid of formula (IVa): R 5 COOH (IVa) wherein R 5 is hydrogen, alkyl, especially C 1-6 alkyl, aryl, especially phenyl, or aralkyl, especially benzyl, and optionally further substituted, or any halide, anhydride or active ester of said carboxylic acid of formula (IVa), or a chloro-format of this general formula (IVb): R 5 O âCO â Cl (IVb) or p-nitrophenyl carbonate of this general formula (IVc): R 50 -CO-O-CGHv-p-NO., (IVc) or the active N-hydroxysuccinimide ester of this formula (IVd):
Or ild (IVd) or azidoformate of the formula (IVe): R 50 -CO-N 9 (IVe) 261853 17 18 wherein R 'is as defined above, or sulfonic. an acid of the formula (IVf):
Rhsn-ii (ivf) wherein R 1; yeah and hydrogen, alkylnpine. in particular, an alkyl group having from 6 to 6 carbon atoms, an aryl group. in particular phenyl, or aralkyl, in particular phenylalkyl, for example benzyl, and these may be optionally substituted, or the halide, anhydride or active ester of the sulfonic acid. it is therefore clear that the acylation reaction protection of amino groups according to the present invention is acylation in a broad sense, including, for example, formylation, acetylation, propionylation, trifluoroacetylation, benzyl oxycarbonylation, p-methoxybenzyloxycarbonylation, t-butoxycarbonylation, phenexycarbony- tosylation, mesylation and other equivalent reactions.
Particular examples of useful acylating agents are acetoxyformyl, p-nitrophenyl formate, acetic anhydride, acetyl chloride, propionic anhydride, trifluoroacetic acid p-nitrophenol ester, trifluoroacetic acid ester, N-benzyloxycarbonylaminocide (a significant active ester). , N-benzyloxycarbonyloxyphthalimide, benzyloxycarbonyl chloride, p-methoxybenzyloxycarbonyloxy-p-nitrophenyl, t-butoxycarbonylazide, phenoxycarbonyl chloride, tosyl chloride, mesyl chloride and others,
The acylating agent, either as such or as a solution in a solvent such as tetrahydrofuran or dimethyl sulfoxide or a mixture thereof, may be added to the solution or suspension containing the aminodycoside antibiotic complex and zinc. Typically, the amount of acylating agent may be greater than or equal to the excess of the number of unexploded amino groups with which the acylating agent is to react. However, in some cases, the amount of acylating agent added may be from about a molar amount and about three times higher than the number of uncomplexed amino groups.
The acylating agent may be added, either at once or in portions over about 2-3 hours, although it may usually be added within 30 minutes to one hour. The acylation may be carried out at a temperature of from -20 ° C to 100 ° C, but may normally be carried out at a temperature in the range of 0 ° C to room temperature. In some cases, the temperature of the reaction may be kept low during the addition of the acylating agent and then gradually increased as the acylation proceeds. Normally, the acylation reaction may be carried out in a sieve in an organic solvent in which an aminoglycoside antibiotic complex with a zinc cation is formed.
This acylation of the zinc complex forms an N-acylated zinc complex, i.e. a complex of zinc cations with a selectively N-acylated derivative of cminoglycoside antibiotic.
According to the method of the first idea of the present invention, the acylation of the zinc cation complex of the aminoglycoside antibiotic is followed by a step of removing the zinc cation from the N-acylated zinc complex, namely, the zinc complex is decomposed to obtain an actively protected N-acylated derivative of an aminoglyccside antibiotic which is zinc cations.
To remove zinc cations from the N-acylated zinc complex, it is necessary to react N-acylated zinc complex with a suitable reagent that removes the zinc cation from said N-acylated zinc complex. There are many ways to use this. The first method consists in reacting the zinc precipitating agent capable of converting the zinc cation to a zinc compound water-soluble, for example zinc sulfide, zinc hydroxide or zinc carbonate, while the N-acylated zinc complex remains dissolved in mixtures for acylation reactions where the zinc cation complex of the aminoglycoside antibiotic was acylated, or after being transferred to a new solution in fresh organic solvent from said mixture for the acylation reaction. The zinc precipitating agent useful in the first process may be hydrogen sulfide, some alkali metal sulfide, ammonium sulfide, some alkaline earth sulfide, for example, calcium sulfide, and alkali metal carbonate, for example sodium carbonate, or ammonium hydroxide. In some cases, the removal of zinc cations from the N-acylated zinc complex can only be accomplished by the addition of water. According to the first method of adding a zinc precipitating agent to a solution of the N-acylated zinc complex, precipitation of the insoluble zinc compound formed from the zinc cations is relatively rapid and the precipitate can be removed by filtration. The N-acylated derivative of the amino-glycoside antibiotic, which then remains in the filtrate solution, can be obtained by concentration of the solution or extraction from the solution, and if necessary can be purified. For example, chromatography in a column of silica gel can be used for purification. The second method is to complete the above-mentioned mixture for acylation reaction (i) by concentrating or concentrating it to dryness by evaporating the solvent, or (ii) diluting with a liquid diluent, which may also be carried out with a new solution of the N-acylated zinc complex of the transferred fresh volume an organic solvent such that an oily or solid sediment, concentrate or residue is obtained, whereupon the desired N-acylated aminoglycoside derivative is obtained in some way from 261853 19. The liquid diluent useful in this second method is water or such an organic liquid in which either the N-acylated zinc complex or the structure of the N-acylated derivative of the aminoglycoside antibiotic of said N-acylated zinc complex is only or only low solubility.
First of all, according to the aforementioned second method, the acylation reaction mixture comprising an N-acylated zinc complex (non-ionic solution of N-acylated zinc complex transferred to an organic solvent) is concentrated or concentrated to dryness to give an oily or solid deposit or residue. . When an inorganic solvent, such as dimethylsulfoxide, etc., is difficult to evaporate for the reaction environment, for the N-acylation of the zinc complex, it is possible that the reaction mixture proacylation containing the N-acylated zinc complex is mixed with the organic liquid diluent, e.g. with an ethyl ether such that the hardly volatile organic solvent is dissolved in or diluted with this diluent, whereby a solid or oil containing N-acylated zinc complex is deposited therefrom. In this way, an oily or solid deposit is obtained which is normally a mixture consisting of (i) an N-acylated zinc complex, i.e. a zinc cation complex with an N-acylated α-minoglycoside antibiotic derivative, (ii) N- an acylated derivative of aminoglycoside antibiotic deprived of decomposition of the complex-forming linkage in a portion of the N-acylated zinc complex due to the substantial absence of an organic solvent environment; (iii) from a certain amount of inorganic zinc formed by complex-forming digestion coupling in a portion of the N-acylated zinc complex, (iv) from a certain amount of zinc salt that has been added initially as excess and has remained unreacted by the formation of the complex, and possibly (v) the residual amount of organic solvent used in previous operations. The above-mentioned oily or solid residue (the aforementioned mixture) may then be treated by any of the processes (a), (b) and (c) below. (a) The oily or solid deposit or the abovementioned mixture is mixed with or by a polar organic solvent, an aqueous polar organic solvent or a mixture of polar organic solvents, which is a polar organic liquid disintegrating complex of the zinc cations in the N- the acylated zinc complex present in said deposit or residue, and wherein the portions of the zinc salt liberated and initially unreacted are soluble but in which the desired N-acylated aminoglycoside derivative antibiotic is insoluble. In this way, the N-acylated zinc complex is broken down to release the zinc compounds therefrom, allowing the zinc cations to dissolve and extract as zinc salt with water or aqueous inorganic solvent, and leaving the desired N-acylated amino glycoside antibiotic derivative in a non-zinc salt. residual to be recovered. The residue may optionally be purified by redissolving it in an organic solvent. The polar organic solvent useful in this process (a) is, for example, methanol, ethanol, liquid ammonia, ethylamine and triethylamine. These polar organic solvents and water serve as a zinc removal agent. (b) Alternatively, the oily or solid composition (the aforementioned mixture) is blended with a different kind of polar organic solvent, either anhydrous or aqueous, to decompose the zinc cation complex formation in the N-acylated zinc complex present in said soil or residue and in which the zinc salt is not soluble, but the desired N-acylated aminoglycoside derivative is soluble so that the N-acylated zinc complex is decomposed to release the N-acylated aminoglycoside antibiotic derivative therefrom to allow dissolution and extraction by said polar organic solvent and thus salt separation. zinc which is released but remains undissolved in said polar organic solvent. In this way, a solution of the desired N-acylated aminoglycoside antibiotic derivative in a polar organic solvent is obtained and collected, it can be purified, for example, chromatographically with the following concentration of purified solution to separate the desired N-acylated product. (c) According to a further alternative, the oily or solid deposit or residue (the aforementioned mixture) obtained in the aforementioned second method can be redissolved as a whole in a suitable organic solvent containing a proportion of water when the whole residue is or a water-soluble or substantially water-soluble residue. The solution thus obtained can then be subjected to a chromatographic process in which the liberated zinc salt and the released N-acylated aminoglycoside antibiotic derivative can be recovered separately from the solution. The present inventors have found that various types of ion-exchange resins, anion exchange and chelating and water-insoluble high polymers containing combinations capable of combining with metal, such as chitin or chito-san, are useful for this chromatographic process. . Suitable cation exchange resin grades for this purpose are those which carry carboxy groups (âCOOH 1 as interchange functions, and those containing sulfonyl groups (âSO: iH as exchange functions. in which the cation exchange resin containing the carboxyl functions for the above-mentioned chromatographic process, the above-mentioned oily or solid residue or residue (the aforementioned mixture) is dissolved in a suitable aqueous organic solvent, for example 10% to 90% by volume of water and ethanol. water, or a mixture of water and an dioxane containing from 10% to 90% by volume of water, and the resulting solution is introduced into a column of said cation exchange resin, and the column is then washed with an additional amount of the above-mentioned organic solvent, followed by induction of e- The eluent uses the amount of the above aqueous organic As the acid, a weak organic acid, for example, acetic acid, or a dilute inorganic acid, such as dilute hydrochloric acid, may be used.
As a base, α-inorganic hydroxide can be used for most cases. The concentration of the acid or base in the developing solvent (the eluent may preferably be from 0.01 to 5% by weight of the developing solution. The desired N-acylated aminoglycoside derivative may be separated from the complexing zinc cations during the process of development). Since the resin used for the exchangers has different adsorption affinities with respect to the desired N-acylated aminoglycoside and zinc cations, it is firstly different from that of the other for bonding with the resin for bonding with the resin. zinc-free N-acylated aminoglycoside which can then be concentrated to obtain the desired N-acylated aminoglycoside antibiotic derivative.
If the cation exchange resin containing the sulfonyl functions is mentioned above, the separation and extraction of the desired N-acylated aminoglycoside antibiotic agent can be accomplished in the same manner as above, since to separate the N-acylated aminoglycoside cd com -Plexing Zinc Cations is the same mechanism. On the other hand, a weakly or strongly alkaline anion exchange resin is used for the chromatin-graphical process, the proportion of N-acylated aminoglycoside in the N-acylated zinc complex which contains one or more non-acylated amino groups is not normal they are adsorbed weakly or strongly by base-resins for the anion exchange due to ionic repulsion between them, so that the development of an anion exchange resin column with a suitable aqueous organic solvent allows the N-acylated aminoglycoside antibiotic derivative to elute while the zinc cations remain column.
Performing the chromatographic process using a chelate exchange resin capable of combining with zinc cation, a solution of the above-mentioned solid or solid residue or residue (above) in an aqueous organic solvent is introduced into the resin column of the chelate exchange the developing solvent is then eluted from the column, while the zinc cations remain bound in the resin to exchange the chelates. The water-soluble high polymer containing metal-combining functions, such as chitin and chitonil, can be used in the same manner as when using a resin to exchange chelates. (dj Furthermore, a process in which the aforementioned acylation reaction mixture in which zinc complex acylation was performed to protect amino groups is directly introduced into a column of cation exchange resin or anion exchange, chelating agent, or water insoluble exchange) a high polymer having a combination function of the combination so that the N-acylated zinc complex is adsorbed by the resin or by the high polymer, which may then be a washbasin organic solvent containing or not containing acid or base as indicated in process (cj, followed by similar operations) as in process (c), thereby removing zinc cations from the N-acylated zinc complex, as well as extracting the desired N-acylated aminoglycoside antibiotic derivative. The N-acylated derivative of the α-minoglycoside antibiotic of the above-mentioned acylation with reakciobsahujĂcĂ N-acylated zinc komplexzpracuje immediately adding water as dy, if the desired N-acyl-derivative of derivatization of the aminoglycoside antibiotic is not soluble in water.
An example of an N-acylated derivative of an α-minoglyccside antibiotic which is substantially insoluble in water may be 3,2 ', 6'-tri-N-benzyloxycarboinyldibecacin. In this case, when the acylation reaction mixture containing the N-acylated zinc complex containing the N-acylated derivative of the aminoglycoside substantially insoluble in the water is immediately mixed with water, the complexing bonding in the Na-linked zinc complex and the N-acyl- the aminoglycoside derivative is precipitated as a solid while the zinc salt formed from the released zinc cations remains in solution so that the desired N-acylated derivative of the 281853 24 23 aminoglycoside antibiotic as a substantially pure product can be recovered separately from the zinc salt.
As mentioned above, the N-acylation, namely the reaction to protect amino groups, is carried out with the zinc complex of the aminoglycoside antibiotic in accordance with the first idea of the present invention and the complex of zinc zinc mono-, di-, tri- or the poly-N-acylated derivative of aminoglycoside thus formed is one in which the zinc cations used are complexed to the structure of the N-acylated aminoglycoside derivative. If the desired N-acylated aminoglycoside derivative is insoluble or poorly soluble in the water, simply adding water to the mixture causes an N-acylated zinc complex containing acylation reaction such that the water-insoluble N-acylated aminoglycoside derivative precipitates as a solid, whereas the release cations are removed from the mixture by dissolution in water as in the fourth process described in the preceding paragraph (ej). The precipitate thus obtained, which is insoluble in water, can be used immediately as starting material for the following reactions for More generally, although the N-acylated aminoglycoside antibiotic derivative is sometimes soluble or partially soluble in water, and therefore the N-acylated aminoglycoside derivative can be recovered only at a substantially reduced yield if a simple water addition process is used immediately Therefore, a better result can be obtained by using either of the above procedures (bj and (cj, in which the N-acylated zinc complex, that is, the zinc cation complex with N-acy The separated aminoglycoside derivative formed in the N-acylation reaction is first separated from the acylation reaction mixture, the N-acylated zinc complex thus separated is then dissolved in water or some aqueous organic solvent and the resulting solution is further dissolved. processed to remove zinc cations therefrom. One simple method of removing zinc cations that is generally applicable is that in which the hydrogen sulfide or alkali metal sulfide is reacted as a zinc cation precipitating agent to precipitate it as zinc sulfide, which is one method of the first procedure described in (a) above. However, the zinc sulfide sometimes precipitates as a colloidal deposit, which is not very filtered and, in addition, hydrogen sulfide and sulfide alkali metals have an unpleasant odor not suitable for use in commercial use of the process. Therefore, the present inventors have conducted extensive research to create a practical way to remove zinc cations from the zinc complex without the use of sulfides, and have succeeded in developing an efficient and easy way to remove zinc cations by using the above-mentioned exchange resins. ions or other polymeric material, such as in processes (c) and (d) described above. These processes (cj and dj are commercially advantageous and valuable since they are easy to perform, give high zinc deprotection efficiency and give a high yield of the desired N-acylated aminoglycoside antibiotic derivative.
Finally, the methods and procedures described above for treating the N-acylated zinc complex can be summarized as follows: (i) A zinc cation complex with a selectively N-acylated aminoglycoside antibiotic derivative; separating the pro-acylation reaction from the mixture before reacting with one zinc removal cation reagent from the complex. (ii) The zinc cathode complex with a selectively N-acylated aminoglycoside derivative antibiotic is separated from the mixture for acylation by extraction with an organic solvent, evaporation of the organic solvent from the acylation re-mixture or dilution of the mixture for (iii) The zinc cation complex is mixed selectively with N-acylated amino-glycoside derivative antibiotic once with water or some polar organic solvent; either anhydrous or aqueous to serve as a zinc cation removal agent. This polar organic solvent is either one of which the zinc salt is soluble, but in which the N-acylated aminoglycoside antibiotic derivative is insoluble or one in which the zinc salt is insoluble but in which the N-acylated derivative is aminogly -coside antibiotic soluble. NIV: The zinc cation complex with the N-acylated aminoglycoside derivative of the edible antibiotic is again dissolved in an organic solvent containing some water, and the resulting solution is subjected to a chromatographic process using a cation exchange resin, anion exchange, exchange or a water-insoluble polymer containing functional groups capable of being combined with a metal serving as a zinc cation removal agent. (v) The acylation reaction mixture is passed through a column of cation exchange resin, anion exchange, exchanges, or a water insoluble polymer comprising a metal-combining function to adsorb the zinc cation complex with the N-acylated derivative; the minoglycoside antibiotic, and the column is then developed with some aqueous organic 261853 25 28 solvent containing or not containing an acid or base, and the eluate is collected in fractions, followed by extraction of the fraction containing the desired selectively N-acylated amino dykoside derivative; containing no zinc cations. (vij If the desired N-acylated α-aminoglycoside antibiotic derivative is insoluble or substantially insoluble in water, the acylation reaction is directly mixed with the water so that said derivative precipitates out of the zinc salt which remains dissolved. (vii) The acylation reaction mixture is reacted immediately with hydrogen sulfide, alkali metal sulfide, or alkaline earth sulfide to precipitate zinc calions such as zinc sulfide or ammonium hydroxide to precipitate zinc sulfates. In the zinc complex mentioned in the first idea of the invention, the zinc salts are principally linked to the 1-amino and 3 ' -amino-aminoglycoside antibiotic complex and hence the N-acylation of the aminoglycoside antibiotic complex with the zinc cation followed by by removing the zinc cations, normally the N-acylated amino-glycoside derivative is preferred antibiotics wherein the amino or alkylamino groups other than the 1-amino and the 3-amino groups are protected by an alkyl group.
When the N-acylated derivative of the aminoglycoside antibiotic thus obtained is acylated according to the first idea of the present invention with an .alpha.-hydroxy-.alpha.-aminoalkanic acid according to the method described in U.S. Pat. No. 3,939,143, followed by removal of the remaining protecting groups by protecting the amino groups from the resulting 1-N-acylated product to provide a semi-synthetic 1-N-acylated amine glycoside antibiotic known as a useful antibacterial agent.
The synthesis of 1-N-acylated aminoglycoside antibiotics is now described with reference to the exemplary use of canaraycin A as a goat. When kanamycin A is used as the starting material in the process of the invention, it is initially blocked by 1-amino- and 3-amino-complexes by complexation with seasoning cations upon formation of its zinc complex. Thus, if the kanamycin A complex with the zinc cation is acylated with a suitable acylating agent of the present invention or another amino blocking agent, the non-complexed 3-amino- and 6'-amino groups of the kanamyl A moiety can be protected by the acyl group used. agents or other species of α-nosculitis blocking group. Subsequent removal of the complexing zinc cations from the zinc cation N-acylated 10 'kanamycin A complex, the resulting N-acylated kanamycin A derivative is reacted with an acylating agent having an acyl group to be introduced into the 1-amine; This acyl fill then only reacts with the unblocked 1-amino- and 3-amino groups of kanamycin A. In this context, the 1-amino group is normally somewhat re-more active than the 3-amino group, so The desired 1-N-acylated derivative of kanamycin A can be obtained with a slightly higher yield than the 3 "-N-acylated cananycin A derivative. Subsequent cancellation of the N-protection of the 1-N-acylated kanamycin derivative of Atakto obtained by 1-N acylated cannabinic A as a desired end product. When using the process according to the first idea of the present invention, it can be seen that the desired 1-N-acylkanamycin A can be obtained at a higher yield compared to the case where unprotected kanamycin A or 6 The N-protected kanamycin A is reacted directly with an acylating agent for 1-N-acylation of kanamycin A. If no N-protecting kanamycin is reacted with any 1-N-acylating agent it is found that mixed N-acylated products comprising a very small proportion, usually from 1% to a few% by weight, are found in this case. the desired 1-na-cyano product.
When using the method of the first aspect of the present invention for kanamycin of the above general formula (III), some or all of the amino acids other than the 1-amino- and 3-amino-amino groups of the used kanamycin are protected, giving the N-acylated derivative kanamycin corresponding to this general formula (V):
where
Rfa is hydroxy, amino (âNH 1), âNHCOR 5, or âNHCO · OR 5, âNHSO 2 R 6, R <is hydroxy, âNHCO3, group 261853 2? 28 R8 /
âN COR5 group - NHCO.OR5, group R8 /
âNC â OR5 âNHSO2Rfi or R8 /
SO2R ° R3 and R! are as defined above in relation to formula (III), R 7 is âCOR 5, - CO.OR 5 or âSO 2 R 6, R 5 and R G are as defined above for formulas (IVa) to (IVf), and R 8 is an alkyl group, especially an alkyl group having 1 to 4 carbon atoms. When the method of the first aspect of the present invention is applied to some kanamycin, a N-protected derivative of kanamycin of formula (V) is usually obtained, wherein all amino groups other than the amino and / or alkylamino groups present in the positions 1- and 3-kanamycin molecules are blocked.
However, when the acyl group to be introduced as an amino blocking group is relatively broad in its spherical size, for example t-butoxycarbonyl, or when the molar amount of acylating agent used in the reaction is less than the stoichiometric amount desirable for the acylation of non-complexing amino groups in the kanamycin molecule, although the acyl group acylating agent is customary in size, or when the acylation reaction is stopped on the scale, an N-protected derivative of kanamycin is obtained in which the number of acylated amino groups in the molecule kanamycin is smaller than in the above case, and then in particular cases such a limited N-acylated derivative of kanamycin is obtained, in which the 6'-amino- or 6'-alkylamino group is exceptionally acylated, by that of 6 ' the amino- or 6'-alkylamino group is more reactive than the other amino group in the kanamycin molecule. The N-acylated derivative of kanamycin of general formula (V) is a significant intermediate useful in the semi-synthetic preparation of various kinds of kanamycin derivatives. The compound of formula (V) has an increased value as the intermediate for chemical synthesis, for example, when a 1-N-acylated aminoglycoside antibiotic active against kanamycin-resistant bacteria is introduced into the process, acylation of the 1-amino-group of the compound of formula (V) any α-hydroxy-Ï-aminoalkanoic acid followed by deprotection of the blocked amino and / or alkylamino groups of the resulting 1-N-acylated product. For example, if the intermediate compound of formula (V) is to be acylated with an acyl group, for example (S) -4-benzyloxycarbonyl-amino-2-hydroxybutyryl, the compound of formula (5) may be in a suitable solvent, e.g. aqueous tetrahydrofuran, reacted with an appropriately substituted butyric acid or an equivalent reactive derivative thereof, for example an active ester, for example N-hydroxysuccinimide ester. N-hydroxyphthalimide ester or p-nitrophenol ester to form the 1-N-acylation product. The subsequent removal of the benzyloxycarbonyl and protecting group (R 7) in formula (V) from the 1-N-acylation product can be accomplished by the conventional N-deprotection technique, for example, either by hydrolysis with acid or base, or by reduction with a reducing metal, or by catalytic hydrogenolysis with water. by reducing the sodium radical in the liquid ammonia to obtain a semisynthetic derivative of kanamycin having a (S) -4-amino-2-hydroxybutyryl group linked to the 1-amino group of kanamycin and active against the lower bacteria, corresponding to this general formula (VI): ".
Wherein R 1, R 2, R 2, and R 3 'are as defined above in relation to formula (III). In the above method, a generally N-protected derivative of some N -hydroxy-w-aminoalkanoic acid of formula (VII): HOOCCH (CH 2) can be used.
OH (VII) wherein n is equal to 1, 2 or 3, instead of (S) -4-benzylcarbonylbonylainino-2-hydroxybutyric acid, thereby obtaining a derivative of 1-N- (R) -w hydroxy-.alpha.-aminoalkynoyl-.beta.-cyanine.
The process according to the invention makes it possible to prepare a 1-N-acylated aminogycoside antibiotic which is known as a semi-synthetic antibacterial composition in high yield. Accordingly, the present invention further provides a process for the preparation of 1N- (α-hydroxy-Ï-aminoalkanoyl aminoglycoside antibiotic starting from a known aminoglycoside antibiotic, wherein the amino-cationic complex is initially produced by the above process). zinc glycoside antibiotics, a partially protected N-acylated α-minoglycoside antibiotic derivative in which the 1-amino- and 3 3-amino- or 3--alkylamino groups are not protected and all other amino groups are protected, whereupon the 1-N-unprotected and otherwise N-fully protected derivative by the selective 3'-acylation process described in US Patent No. 261,859 and moves the 1-amino group of the 1-N-unprotected and otherwise N-fully the protected derivative obtained in the preceding step 3 "-N-acylation, acylating with α-hydroxy-ÎČ-amino amino acid, in particular 3-amino-2-hydroxypropionic acid (isoserine) or 4-amino-2-hydroxybutyric acid finally, protecting groups from the N-acy-left product are removed.
Method of production lN- (thihydroxy- Îł-amino-alkynoyl derivative of aminoglycoside antibiotic containing 6-O- (3 "-amino-or 3" -alkylamino-3 "-deoxyglycosyl) -2-deoxystreptamine moiety, optionally containing a 4-O-aminoglycosyl group is carried out in a manner that (also zinc cations are reacted with an aminoglycoside antibiotic in an inert organic solvent to form a zinc cation complex with an aminoglycoside antibiotic, (bj with a zinc cation complex with an aminoglycoside antibiotic that has been formed in The above-described step (a) is reacted in situ in an inert organic solvent with an acylating agent having an acyl group to be introduced as an amine protecting group to form a complex of zinc cations with a selectively N-acylated aminoglycoside derivative of the antibiotic originally uncomplexed amino groups acylated; (c) selectively N-acylated aminoglyc derivative the osidic antibiotic with the seasoning cation obtained in the above-described step (bj is reacted with a reagent to remove zinc cations from the N-acylated zinc complex, to form partially and selectively protected N-acylated aminoglycoside derivative antibiotics which is free of zinc cations and wherein the 1-amino- and 3-amino- or 3'-alkylamino groups are unprotected, but all other amino groups of the aminoglycoside molecule are protected by an acyl group, (d i partially and selectively protected by Na) the -cylated derivative obtained in the above-described step (cj is reacted with an ester of some alkanoic acid which corresponds to (VIIIa):
Ra â C â RbII o (VIII) in which
R a is hydrogen or dihaloalkyl or (C 1 -C 6) trihaloalkyl and R b is (C 1 -C 6) alkyloxy, especially benzyloxy or an aryloxy group, especially phenoxy or N-formylimidazole, as an inert acylating agent an organic solvent to selectively acylate the 3 "-amino- or 3" -alkylamino group with an acyl group of RCO-said acylating agent to form a 1-N-unprotected and other N-fully protected aminoglycoside derivative of antibiotic, in which all amino groups other than The 1-amino group is acyl protected, (e) 1-N-unprotected and the other N-protected derivative obtained in the above-described step (dj is reacted with some α-hydroxy-Ï-aminoalkanoic acid of formula (IX j: HOOC) -CH (CH2-NH2
OH (IX) wherein m is 1 or 2, or with an equivalent reactive derivative of this acid, the amino group of which is either unprotected or protected, to acylate the 1-amino group of said 1-N-unprotected derivative, (f) whereupon the remaining amino-protecting groups 261853 31 32 are removed from the 1-N-acylation product obtained in the above-described step (s) in a conventional manner for deprotection. Next, the implementation of the method according to the third idea of the present invention will be described in more detail.
Aminoglycoside antibiotics that are suitable as a starting material for the first step (a) of this process are the same as the anti-biotics described above in the method of the first invention. The reaction of the complexing zinc cations with the aminoglycoside antibiotic is carried out in the same manner as described above. The acylation of the aminoglycoside antibiotic complex with the zinc cation obtained in the first step (i may be in the second step (as described above for the method of the first invention. Removal of zinc cations from selectively N-acylated complex aminoglycoside antibiotic and zinc) The cations thus obtained can be the third step (c) of this process by various methods, as described above, thereby obtaining a partially and selectively protected zinc cation-free N-acylated aminoglycoside derivative and wherein the 1-amino - and the 3 "-amino-or 3" -alkylamino groups are unprotected, but all other amino groups in the aminoglycoside molecule are blocked by the acyl group of the acylating agent used in the step (bj of this process. This partially selectively protected N-acylated derivative of aminoglycoside antibiotic is descendants react in step (dj of this process with an ester of some alkanoic acid (VIII j or with N-formimidazole in the same manner as described above adapted according to the second idea of the invention to obtain a selective 3 '-N-acylation of a partially N-protected aminoglycoside derivative without acylation) its 1-amino group. In the fifth step (ej of this process 1-N - unprotected and another N-fully protected aminoglycoside antibiotic derivative obtained in the previous step (dj of this method is reacted with some α-hydroxy-Ï-aminoalkanoic acid of formula (X ), in particular with 3-amino-2-hydroxypropionic acid (DL-isoserine, D-isoserine or L-isoserinj, or with L-4-amino-2-hydroxy-acid to acylate 1-amino-aminoglycoside antibiotic 3-amino-2 - hydroxypropionyl- or 4-amino-2-hydroxy-butyryl.
This 1-N-acylation can be carried out generally as described in British Patent Specification 1 426 908 or in the patent application Sp. st. am. No. 4,001,208, according to any known method for the synthesis of amides by reacting a protected aminoglycoside antibiotic derivative with an isoserine or with L-4-amino-2-hydroxybutyric acid, either in its free acid form or in its form. a reactive equivalent, for example an active ester, for example a dicyclohexylcarbodimide ester, a mixed acid anhydride, an acid azide, in an inert organic solvent, for example dioxane, dimethoxyethane, dimethylformamide, tetrahydrofuran or an aqueous form of these solvents. Isoserine and L-4-amino-2-hydroxybutyric acid may be those whose amino groups have been blocked by an amino protecting group. Suitable amino protecting groups for this purpose may be the same or different from those used in 1 -N-unprotected but another N-protected aminoglycoside antibiotic derivative to be 1-N-acylated. A preferred group for protecting amino groups is t-butoxycarbonyl, since it is directly removable by reaction with dilute acid, for example aqueous trifluoroacetic acid, aqueous acetic acid, and dilute hydrochloric acid.
Furthermore, benzyloxycarbonyl groups, which are removed by conventional hydrogenolysis on palladium or platinum oxide catalysts, as well as phthaloyl groups which are readily removed by hydrolysis with hydrazine, are suitable as amino protecting groups.
The acylation reaction of step 1-N-acylation (according to the method of the fourth idea of the present invention may advantageously be carried out in an aqueous organic solvent using any active ester of the? -Hydroxy? -Aminoalkanoic acid formula (Xj. the active ester may be an isoserine N-hydroxysuccinimide ester or L-4-benzyloxycarbonylamino-2-hydroxy-butyric acid, and the active ester may be used in an amount of from 1 to 2 moles, preferably from 1 to 1.5 moles per 1 The water miscible organic solvent used in the reaction medium can preferably be dioxane, dimethoxyethane, dimethylformamide or tetrahydrofuran.
After step (f) of this process is carried out to remove the protection, i.e., the removal of all remaining amino protecting groups from the 1-N-acylation product obtained in the previous step (removal of the remaining amino protecting groups) The remaining amino protecting group, which is an alkoxycarbonyl type, can be removed by hydrolysis with an aqueous solution of trifluoroacetic acid or acetic acid or dilute acid solution, for example dilute hydrochloric acid. The so-called aminoalkyl protecting group, which is an aralkylcarbonyl type, for example benzyloxycarbonyl, is removed directly by catalytic hydrogenolysis When all of the remaining amino protecting groups from the 1-N-acylation product of the step are removed by 261853 33 34 ( e] of this process, the desired 1N- (2-hydroxy-3-aminoprcpion) is obtained in high yield. Examples of 1-N- {α-hydroxy-Ï-aminoalkanecyl] -aminoglycoside antibiotic prepared by the method of the fourth embodiment of the present invention are given below. further.
(1] 1-N- (L-4-amino-2-hydroxybutyryl) -kanamycin A
(2) 1-N- (L-4-amino-2-hydroxybutyryl) -3'-deoxycanamycin A
(3) 1-N- (L-4-amino-2-hydroxybutyryl) -3 ', 4 ' -dideoxycanamycin A (4) 1-N- (L-4-amino-2-hydroxybutyryl) -tobramycin ( 5 1-N- (L-4-amino-2-hydroxybutyryl-1-dibecacin (6) 1 N- (3-amino-2-hydroxypropionyl) dibecacin other uses of the methods of the first and second embodiments of the present invention reside in the preparation of an N-alkylaminoglycoside an antibiotics from all N-acylated aminoglycoside derivatives containing an unprotected 1-amino group, and examples of this may be the preparation of netilmycin or its 1-N-alkylanalogs from sisomycin by ligation with a lower aliphatic aldehyde and cyanoborohydride.
The invention will be further illustrated by the following examples. Example 1
Preparation of 3,6'-di-N-benzyloxycarbonylcanancin A (i) 2.0 g, 4.13 mM of kanamycin A as the free base was suspended in a mixture of 50 billion dimethyl sulfoxide and 20 ml tetrahydrofuran and 4 g was added to the suspension, 18.1 mM zinc acetate, dihydrate, whereupon the reaction mixture was shaken at room temperature until a homogeneous solution was formed. The formation and dissolution of the zinc complex of kanamycin A lasted for about 4-5 hours. The resulting solution was then cooled to 0 ° C and a cold solution of 2.37 g, 9.5 mM N was added thereto for one hour. - benzyloxycarbonyloxysuccinimide,
H5CH & OCOO-N / O dissolved in a mixture of 40 ml of tetrahydrofuran-dimethyl sulfoxide 1: 1 by volume The reaction solution was allowed to stand at ambient temperature for 4 hours, during which time the zinc complex of kanamycin A was benzyloxycarbonyl, represents acylation according to the first aspect of the invention.
The sample obtained from the reaction mixture thus obtained was chromatographed on a thin layer of silica gel, using a lower liquid phase of chloroform-methanol-28% aqueous ammonia in a ratio of 1: 1: 1 as the starting solution, giving a major spot of the desired product during chromatography. at R (= 0.23 and two or three smaller spots belonging to by-products at higher points. (ii) The above reaction solution was poured into 500 ml of ethyl ether and the separated oil was washed several times with additional volumes of ethyl ether, thereby (iii) Removal of the zinc cation from a predominantly zinc-containing syrup was carried out by any of the following various procedures: (A) A procedure using a weakly acidic cation exchange resin carboxyl group -COOH as a functional group, commercially available under the name "Amberlite" CG 50 resin (HH form 60 ml of Amber lit CG 50 H + formate was pre-saturated with a water-dioxane mixture (2: 1) and then filled into columns. A solution of 1 g of the syrupy substance dissolved in 20 ml of water-dioxane (1: 1) was passed through a column which was then induced with water-dioxane (2: 1) containing 1% acetic acid. The desired 3,6'-di-N-benzyloxycarbonylkanamycin A, which was positive in the hydrine reaction, was eluted first from the column and zinc acetate sensitive to diphenylcarbazidene was eluted after it. The fractions containing the desired product were combined and concentrated to dryness, then the residue was washed with ethyl ether to yield 340 mg (81%) of 3,6 ' -di-N-benzyl < / RTI >
[α] 25 D + 76 ° (c 1, water-dimethylformamide, 1: 2).
Elementary analysis
Calculated for Cy, HwN4O15. 2 CH-C0-, H-H-, O: 51.23% C, 6.56% H, 6.29% N,
Found: 51.02% C, 6.71% H, 6.22% N. (B) Weak Cation Exchange Method 33
Ion exchange resin carrying a carboxy group as a functional group, commercially available as "Amberite" CG50 resin, HH // form from Rohm and Haas Co. 1 g of the syrupy substance obtained above in Example 1 (ii) was dissolved in 20 ml of a 1: 1 mixture of water-dioxane and the solution was passed through a column of 60 ml of AmberlitCG 50 resin, NIL · / form, and eluted with a linear mixture gradient water-dioxane 1.: 1 containing up to 0.1 N ammonia. No zinc cation was eluted but only the desired product, 3,6'-di-N-benzyloxycarbonylcanoic acid A. The elution fractions containing the desired benzyloxycarbonylated products were concentrated to dryness to yield 333 g, i.e. 89% of the desired product in in the form of a colorless solid.
[.alpha.] D @ 20 = + 86 DEG (c 1, water-dinethylformamide, 1: 2).
Elementary analysis
Calculated for C-wH ^Nz.Ur ,. W 1-1-, 0 (.): 52.87% C, 6.30% II, 7.15% N,
Found: 52.50% C, 6.50% H, 7.00% N. (C) A method using a cation-exchange ion exchange resin bearing a strongly acid-functional group -SO-, Î, commercially available as "Dovex" 50 WX 2 nd Dox Chemi-cal Co. Ltd 30 ml of Dowex 50 W X2-H + form in water-dioxane 2: 1 was charged to the column, followed by a solution of 1 g of syrupy substance obtained in Example 1 [ii] in 20 ml of water-di-oxane 2: 1 The column was then washed with a 2: 1 mixture of water-dioxane until the column mixture was neutral and then eluted with a linear gradient of 2: 1 v / v -dioxane containing 0 to 1 N particles. The eluted fractions containing the desired 3, 6 ' -di-N-benzyloxycarbonylkanamycin A were concentrated to dryness under reduced pressure to give 311 mg of the white solid which was identical to that obtained in Example 1 (84%). iii 1 [B]. (D) Alternative procedure using Do-wex 50W X 2
A solution of 1 g of the syrupy substance obtained in Example 1 (ii) in 20 ml of water-methanol 3: 1 was applied to a column of 30 ml of Dowex 50W X 2 in H + form with a pre-wetted mixture of water-methanol 3 The column was then washed well with a water-methanol 3: 1 mixture and then eluted with a water-methanol gradient of 3: 1 containing 0 to 6 N hydrochloric acid. The active fractions containing the desired 3,6'-di-N-benzyloxycarbonyl-kanamycin A were collected and mixed with 36 strongly basic anion exchange ion exchange resin Dowex 1 X 2 in an OH form in an amount which was able to acidify slightly.
The mixture was filtered and the filtrate was concentrated to dryness to give 286 mg, i.e. 72% of the desired product as the dihydrochloride.
[α] D 25 - + 79 ° (c 1, water-dimethylformamide, 1: 2). (E) A method using an anion exchange ion exchange resin bearing strongly basic functional quaternary ammonium groups, commercially available as Dowex 1X2cd. Dow Chemical Co.
A solution of 1 g of the syrupy substance obtained in Example I (ii) in a water-dioxane (1: 1) mixture was applied to a column of 30 ml of Dowex 1 X 2 resin in an OH form of a pre-washed water-dioxane 1 mixture. : 1, and then the column was induced with a 1: 1 water-dioxane mixture at a relatively high rate. The eluted fractions containing the desired product were collected and concentrated to dryness to give 305 mg, i.e. 84% colorless solid identical to Example 1 (iii) (B). (F) Procedure using anion exchange ion exchange resin with weakly basic functional groups, commercially available as Dowex WGR, manufactured by DowChemical Co. 1 g of the syrupy substance obtained in Example 1. (ii.) was dissolved in 20 ml of a water-dioxane 2: 1 mixture and the solution was left with a counter-column of 50 ml of Dowex WGR in a basic pre-soaked water-dio-xane mixture 2: 1 and the puck was eluted with a water-oxalic mixture. 2: 1. The desired 3,6'-di-N-benzyloxycarbonynyanamycin A was eluted in some fractions along with traces of zinc cations. These fractions were combined and concentrated to dryness to give 450 mg of a colorless solid. fG) A process using a chelate ion exchange resin bearing weakly acidic functionalities, commercially available as Dowex A 1, manufactured by Dow Chemical Co., Sp. st. and.
A solution of 1 g of the syrupy substance obtained in Example 1 (ii) in a 1: 1 water-dioxane mixture was loaded onto a 50 ml DowexAI column, which was saturated with a 1: 1 water-dioxane mixture containing 1% ammonia and then followed by elution with a 1: 1 water-dioxane gradient containing 0 to 1 N ammonia. The fractions containing the desired 3,6'-di-N-benzyloxycarbonylkanamycin A which were eluted at a later stage were pooled and - thickened to dryness to give 272 mg, i.e. 261853 37, 74% of the desired product as a white solid. (H) A Chitosan process, a water-insoluble metal-binding functional group polymer, commercially available as a Toko KaseiKoyo Co., Ltd. product. Japan'. 100 ml of Chitosan was thoroughly saturated with water-methanol 3: 1 and charged to the column, which was passed through a solution of 1 g of the syrupy substance obtained in Example 1 (ii) in a water-methanol (3: 1) mixture. The column was then induced with a 3: 1 water-methanol mixture, with the desired 3,6'-di-N-benzyloxycarbonylkanamycin A being eluted first and zinc acetate much later. The fractions containing the desired substance were combined and concentrated to dryness to give a residue which was dissolved in a 1: 1 mixture of water-dioxane and the solution was loaded onto a column of Amber lithium CG 50 in NH4 + form prewashed with water-dioxane 1 1. Wash the column thoroughly with 1: 1 water-dioxane and then elute with a 1: 1 water-dioxane gradient containing 0 to 0.1 N ammonia. These fractionated to ninhydrin reactions were combined and concentrated to dryness to give 301 mg, i.e. 82%, of a colorless solid, consistent with the compound obtained in Example 1 (iii) (B). (I) A process employing a high polymeric carboxyl functional group, commercially available as "CM-Sephondex" C-25, which is both an ion exchanger and a gel filtration material and is formed by a carboxymethyl-substituted dextran gel, manufactured by Pharmacia. Fine Chemical Co., Sweden1.
A solution of 1 g of the syrupy substance obtained in Example 1 (ii) in a 1: 1 mixture of water-dioxane was passed through a column of 40 ml of CM-Sephadex C-25 in NH 4 + form, thoroughly saturated with water-dioxane 1: 1 The column was washed with 200 mL of water-dioxane 1: 1 and then eluted with a gradient of 1: 1 v / v dioxane containing 0 to 0.1 N α-mono. No zinc cations were eluted from the column, but only the desired 3,6'-di-N-benzyloxycarbonylkanamycin A was eluted. The eluate was concentrated to dryness to give 303 milligrams, ie 82% of a colorless solid identical to Example 1 (iii) (BJ). ()) Procedure using hydrogen sulfide as a zinc precipitating substance. 1 g of the syrup-like material obtained in Example 1 (ii) was dissolved in 20 ml of water-methanol 1: 1 and aqueous ammonia was added thereto, followed by sufficient hydrogen sulfide. The reaction mixture containing the zinc sulphide precipitate that was formed was filtered on a celite pad filter, and the filtrate was concentrated under reduced pressure to give a syrupy solid from which the ethyl ether was washed with a solid residue. This residue was dissolved in a 1: 1 water-dioxane solution and the solution was chromatographed on a 30 ml Amberlite IRA 900 column, a strongly basic resin, using fyRohm and Haas Co., in the OH form, using water-dioxane 1: 1 as the developing solvent. The eluate was collected and fractions containing 3,6'-di-N-benzyloxycarbonylkanamycin A were pooled and concentrated to dryness to give 235 mg, i.e. 64% of a colorless solid identical to that of Example 1 (iii) ) (B). Example 2
Preparation of 3,6-di-N-benzyloxycarbonylcananine A 500 mg, i.e. 1.03 mM of kanamycin A free base form was suspended in 15 ml of dimethyl sulfoxide followed by addition of 420 mg, i.e. 3.09 mM of zinc chloride and 840 mg, i.e. 6.18 mM sodium acetate tetrahydrate. After stirring the mixture for 10 hours at room temperature, a solution of 675 mg, i.e. 2.27 mM N-benzyloxycarbanyloxyphthalimide, was slowly added to the mixture containing the formed kanamycin A-zinc complex for about one hour.
dissolved in 10 ml of dimethyl sulfoxide. The resulting mixture was allowed to stand at room temperature for 4 hours. Next, the reaction mixture was treated in the same manner as described in Example 1 (iij and (iii) (I) to give 598 mg, i.e. 74 °, of 3,6'-di-N-benzyloxycarbone. of bonkancanamycin A as a colorless solid Example 3
Preparation of 3,6'-di-N-benzyloxycarbonylaminoanine A 600 mg, i.e. 0.95 mM kanamycin A tetrahydrochloride and 150 mg, i.e. 3.8 mM sodium hydroxide in 15 ml dimethyl sulfoxide was shaken one hour, and then 1 g, i.e., 4.55 mM zinc acetate dihydrate, was added and shaking was continued for another 5 hours. A solution of 545 mg, i.e. 2.2 mM of N-benzyloxycarbonyloxy-succinimide dissolved in 5 ml of a dimethylsulfoxidetrahydrofuran 1: 1 mixture was added to the mixture containing the formed kanamycin A-zinc complex during 261853. After shaking the resulting mixture at ambient temperature overnight, ethyl acetate was added to separate the N-acylated zinc complex as a precipitate. The precipitate was then treated in the same manner as described in Example 1 (iiij (H)) to give 581 mg, i.e. 78% of a colorless solid. Example 4 Preparation of 3,6'-di-N-benzyloxycarbonylcanoate
mycine A (i) 500 mg, i.e. 1.03 mM kanamycin A free base form was dissolved in 20 ml of water-dimethylsulfoxide 1: 9, 1 g, i.e. 4.55 mM zinc acetate dihydrate and then 590 g was added mg, i.e. 2.4 mM N-benzyloxycarbonyl-oxysuccinimide.
The mixture was allowed to stand at room temperature overnight and a large amount of ethyl ether was added to the mixture to separate the aqueous sulfurous layer, which was washed several times with ethyl ether, to obtain a thick-layered layer. (ii) The syrup thus obtained was dissolved in water-methanol 3: The solution was passed through a 200 ml Chitosan column. The column was eluted with 3: 1 methanol / methanol and the eluate collected in fractions. The neat hydrotreatment fractions were pooled and concentrated to a small volume. The concentrate was loaded onto a Amberlite CG 50 column in NII + form and the column was washed thoroughly with a 1: 1 water-dioxane mixture and then eluted with a 1: 1 mixture of water-dioxane containing 0 to 0.1 ammonia.
The eluted fractions containing the desired product were combined and concentrated to dryness to give 494 mg, i.e. 6%, of a colorless solid, identical to that of Example 1 (iiij (B)). la la 5
Preparation of 3,6'-di-N-benzyloxycarbonyl-kanamycin A 500 mg, i.e. 1.03 mM of kanamycin A in the free base form, was dissolved in 20 ml of a 1: 3 mixture of water-tetrahydrofuran and 1 g was added. 4.55 mM of the magnesium dihydrate acetate, and then 590 mg, i.e. 2.4 mM of N-benzyloxycarbonyl oxysuccinimide, were added. The mixture was left to stand at room temperature overnight and the reaction solution was concentrated under reduced pressure. The residue was passed through a pad of 200 ml of Chitosan and the eluate was treated in the same manner as in Example 4 (iij to give 414 mg, i.e. 51% of the colorless solid of the desired compound.
Preparation of 3,6'-di-N-benzyloxycarbonyl-kanamycin A is 500 mg, i.e. 1.03 mM kanamycin A, the free base form was dissolved in 15 ml of water-methanol 1: 7 mixture and then added. 5 g, i.e. 6.8 mM of zinc acetate dihydrate, and then 590 ”l, i.e. 2.4 mM of N-benzyloxycarbonyl-oxysuccinimide in 7 ml of tetrahydrofuran was added. The mixture was allowed to stand at ambient temperature over night and the reaction solution thus obtained it was concentrated under reduced pressure. The residue was passed through a column of 200 ml of Chitosan column-eluting eluate then treated in the same manner as in Example 4 (iij to give 442 mg, i.e. 55% of the colorless solid of the title compound).
Preparation of 3,6'-di-N-benzyloxycarbonyl-kanamycin A 500 mg, i.e. 1.03 mM kanamycin A in the free base form was suspended in 20 billion dimethyl sulfoxide and 272 mg, i.e. 1.24 mM zinc acetate. dihydrate was added to the mixture. The mixture was stirred at room temperature for 10 hours, whereupon a clear solution was formed which was then added in small portions for about two hours to 540 mg, i.e. 2.17 mM N-benzyloxycarbonyl oxysuccinimide.
The resulting mixture was then allowed to warm to ambient temperature overnight, large ethyl ether was added and the separated oily substance was collected and washed several times with ethyl ether to give a thick syrup.
Thin-layer chromatography on silica gel, using chloroform-methanol-28% aqueous ammonia 1: 1: 1, lower phase, as the developing solvent, was used to determine the following spots: - smaller R, 0.4-1.3 6 ', 3 "-tetra-N-benzyloxycarbonylkanamycin A stain, where coloring develops after spraying with acid and subsequent heating; Weak spot Rt 0.28; - the main spot of R (0.23 - the desired product, 3,6'-di-N-benzyloxycarbonylcanoate A; - the minor spot of Rf 0.12 6'-N-benzyloxycarbonyl-kanamycin A; - very weak Rf 0 spot - unreacted kanamycin A.
No spot corresponding to tri-N-benzyloxycarbonylkanamycin A was observed which would appear at R 0,2 0.28 to 0.4. The above syrup was dissolved in a 1: 1 water-dioxane mixture and allowed to pass through the column with 100 ml of CM-Sephadex C-25 in NH 4 + form, pre-washed with 1: 1 water-dioxane. eluted by the same procedure as described in Example 1 (iii) (I), removing zinc cations and separating the desired product from other products to give 412 mg, i.e. 51% of the desired compound as a colorless solid.
For comparison, this procedure was repeated except that the zinc acetate di-hydrate was replaced with 308 g, ie, 1.24 mM octanecarbonate tetrahydrate, with the result that the desired 3,6'-di-N-benzyloxycarbonylcanoate mycine A was obtained as a colorless solid only with a yield of 59 mg, i.e. 7.3%. Example 8
Preparation of 3,6'-di-N- (p-methoxybenzyloxycarbonyl) kanamycin A 500 mg, i.e. 1.03 mM kanamycin A free base form was suspended in 12 ml dimethyl sulfoxide and 1 g, 4.55 mM, was added to the suspension. zinc acetate hydrate. The mixture was stirred at room temperature until a homogeneous solution was formed to which a solution of 789 mg (2.6 mM p-methylcarbobenzoxy-p-nitrophenyl ester) was added over 30 minutes (p-CH 2 OC 1 H 2 CH 2 OCOOCOCl 3). p-NO 2) dissolved in dimethyl sulfoxide. The resulting mixture was allowed to stand at ambient temperature overnight and then treated in the same manner as in Example 1 (ii) and (iii) (B) to give 722 mg, i.e. 83% of the colorless solid of the title compound.
[α] 25 D + 87 ° (c 1, water-dimethylformamide, 1: 2).
Elementary analysis
Calculated for C ^ jH ^ NN -Oiyi. H 2 CO ": 51.95% C, 6.33% H, 6.64% N,
Found: 51.56% C, 6.41% H, 6.53% N. Example 9
Preparation of 6'-N- (t-butoxycarbonylkanamycin A
In the same manner as described in Example 8, except that the p-methoxycarbobenzoxy-p-nitrophenyl ester was replaced by 220 mg, i.e. 1.54 mM t-butoxycarbonylazide, the desired compound was obtained as a colorless solid. The yield was 627 milligrams.
[Î] 25 D = + 96 ° (c 1, water - dimethylformamide, 1: 2), Example 10
Preparation of 3,6'-di-N-trifluoroacetylkanamycin A 500 mg, i.e. 1.03 mM of kanamycin A free base form was suspended in 12 ml of dimethylsulfoxide and 1 g of 4.55 mM zinc acetate hydrate was added to the suspension. The mixture was stirred at room temperature until a homogeneous solution was formed, to which a solution of 1.2 g, i.e. 5.1 mM, of p-nitrophenol trifluoroacetic acid ester dissolved in 10 ml of dimethylsulfoxide was added. The resulting mixture was left to stand overnight at ambient temperature and then treated with ethyl ether as in Example 1 (ii). The ether-insoluble sulfurous material was further treated in the same manner as in Example 1 (iii) (A) to give 590 mg (70%) of the desired compound as a colorless solid.
[α] D 25 + 81 ° (c 1, water-dimethylformamide, 1: 2).
Elementary analysis
Calculated for C ^ HH ^ NNĂOriFF,.. 2 CHCO-H. % H, 5.44;% N, 6.88;
Found: C, 33.03; H, 5.48; N, 6.54. Example 11
Preparation of 3,6'-di-N-phenoxycarbonylkanamycin A 500 mg, i.e. 1.03 mM kanamycin A free base form, was suspended in a total of dimethyl sulfoxide, 15 ml and 5 ml of tetrahydrofuran and 1 g was added to the suspension, i.e. 4.55 mM zinc acetate dihydrate and then the mixture was stirred at room temperature until a homogeneous solution was formed. The resulting solution was then cooled to 0 ° C and then a cold 0 ° C solution of 400 mg, i.e., 2.55 mM phenoxycarbonyl chloride C (1 H-, OCOCl in 3 mL tetrahydrofuran) was slowly added. The reaction mixture was treated with ethyl ether as in Example 1 (ii) and the ether-insoluble syrup was further treated in the same manner as in Example 1 (Example 1). iii) (A) to give 625 mg, i.e. 70% of the desired compound in the form of a colorless solid.
[α] D 25 + 73 ° (c 1, water-dimethylformamide, 1: 2).
Elementary analysis
Calculated for C ^ HH ^ NN ^O, ·. 2 CH-CO-J1. Π·, Î: 50,11% C, 6,31% II, 6,49% N,
Found: 49.77% (1, 6.60% H, 6.11% N. Example 12)
Preparation of 3,6'-di-N-acetylkanainycin A
The reaction mixture obtained in the same manner as in Example 8 except that 260 mg, i.e. 2.6 mM acetic anhydride was used in place of p-methoxycarbobenzoxy-p-nitrophenylether, was treated in the same manner as in Example 1 (iii) (A). 525 mg (72%) of the desired compound is thus obtained as a colorless solid.
[I] - Âź 93 (cmid, 1: 2). Analysis Calculated 1, water - dimethylfurma for C% HWN, .O | 11.0: 44.19% C, 7.13% II, 7.63% N, Found: 44.20% C, Example 13 7.07% H, 7 , 85% N
Preparation of 3,6'-di-N-lorruylcanarnycine A 500 rng, i.e. 1.03 mM kanamycin A free base form was suspended in 12 ml dimethylsulfoxide and 1 g, i.e. 4.55 mM octae nu added to this sus-pension. zinc dihydrate. The mixture was stirred at room temperature until a homogeneous solution was formed, to which was added 690 mg, i.e. 4.12 mM, p-nitrophenylformate OHCOD (1H5-ÎČ-NO ·). left to stand overnight at room temperature and then treated in the same manner as in Example 1 (iii) (H).
Fractions positive for ulylivdrius reaction were pooled, gassed with gaseous carbon dioxide, and then concentrated to dryness. This gave 430 mg, i.e. 67% of the desired compound as a colorless substance. fajir 5 101 ° (c 1, water).
Analysis
Calculated for. H9CO · .11.0: C, 40.64; H, 6.50; N, 9.03.
Found: 40.43% C, 6.47% H 8.83% N. 44 Example 14
Preparation of 3,6'-di-N-tosylkanamycin A 500 mg, i.e. 1.03 mM of kanamycin A free base form was suspended in 15 ml of dimethyl sulfoxide and 1 g of 4.55 mM zinc acetate hydrate was added to the suspension. The mixture was stirred at room temperature until a homogeneous solution was formed, to which was slowly added a 400 mg solution, i.e. 2.1 mM tosyl chloride in 7 ml tetrahydrofuran. The resulting mixture was allowed to stand at ambient temperature for one hour, and 200 mg of tosyl chloride dissolved in 3.5 ml of tetrahydrofuran was added. The reaction mixture was allowed to stand for a further two hours and was then worked up in the same manner as in Example 1 (iij and (iii) (Aj, obtaining 270 mg, i.e. 28% colorless solid, representing the desired compound). + 68 ° (c 1, water-dimethylformamide, 1: 2).
Analysis Calculated for C39H48N ,, Or> S3. 2 CH3CO2H. H, 6.28; N, 6.02; N, 6.89. Found: C, 46.31; H, 5.98; N, 6.31. 55% S. In the above-mentioned zinc acetate-free reaction procedure, a large amount of colorless solid was not obtained. Example 1 5
Preparation of 3,6 ' -di-N-benzyloxycarbonyl-6 ' -N ' -methylanine A 500 mg, i.e. 0.1 mM 6 ' -N-methyl-kanamycin A in the loose the base was suspended in 12 ml of dimethyl sulfoxide and 1 g, i.e. 4.55 mM of zinc acetate dihydrate was added to the suspension. The mixture was stirred at room temperature until a homogenous solution was formed to which a 550 mg solution was added over 30 minutes. 2.2 mM N-benzyloxycarbonyloxy-succinimide dissolved in 5 ml of tetrahydrofuran 1: 1 dimethyl sulfoxide. The resulting mixture was allowed to stand overnight at ambient temperature and then treated as in Example 1 (iij and (iii) (A) to yield 720 mg, i.e. 79% of the desired compound as a colorless substance.
[α] D 25 -1-74 ° (c 1, water-dimethylformamide, 1: 2).
Further treatment of the thus prepared 261853 45 compound by a procedure similar to that described in Example 31 below gave 1-N - [(S) -4-amino-2-hydroxybutyryl] -6'-N-methylkamycin A. 16
Preparation of 3,6'-di-N-benzyloxycarbonyl-3'-deoxycanamycin A
This compound as a colorless solid was obtained in a yield of 765 mg, i.e. 82% by repeating the same procedure as in Example 15, except that the starting material was 500 mg, i.e. 1.07 mM 3'-oxycanamycin A in free base form and 610 mg, ie 2.45 mM N-benzyloxycarboinyloxysuccinimide was used.
[α] D 25 - + 76 ° (c 1, water-dimethylformamide, 1: 2).
Analysis
Calculated for C-hI-V ^ Oh. 2 CH 2 CO 2 H. % H, 6.68;% N, 6.40.
Found: 51.99% C, 6.75% H, 6.20% N.
Further work-up of the compound thus prepared by a procedure similar to Example 31 afforded 1 'N - [(S) -4-amino-2-hydroxy-butyryl] -3'-deoxycanamycin A. Example 17
Preparation of 3,6'-di-N-benzyloxycarbonyl-3'-deoxy-6'-N-methylkanamycin A The desired compound was obtained with a yield of 737 mg, i.e. 80% by repeating the same procedure as in the example. 15, except that starting from 500 mg, i.e. 1.04 mM 3'-deoxy-6'-N-methylkanamycin A in the form of the free base, 595 mg, i.e. 2.4 mM N-benzyloxycarbonyloxy succinimide, was used.
[α] 035 + 73 ° (c 1, water-dimethylformamide, 1: 2).
Further treatment of the thus prepared compound gave 1N - [(S) -4-amino-2-hydroxybutyryl] -3'-deoxy-6'-N-methyl-kanamycin A. Example 18
Preparation of 3,6'-di-N-beyloxycarbonyl-4'-deoxycanamycin A
Starting from 500 mg, ie 1.07 mM 4'-deoxy-kanamycin A in free base form [see Journal of Antibiotics, Vol. 27, s. 838-46-849 (1974); Bulletin of the Chemical So-ciety of Japan, Vol. 50, pp. 2,362-2,368 (1977)], the desired compound was obtained in the form of a colorless solid with a yield of 666 mg, i.e. 71% by the same procedure as in Example 15, except that 580 mg, i.e. 2.3 mM N-benzyloxycarbonyl-oxysuccinimide dissolved in 4 ml of dimethyl sulfoxide was added slowly over a period of more than one hour to a homogeneous solution.
[α] D 25 = + 77 ° (c 1, water-dimethylformamide, 1: 2).
Analysis
Calculated for Cb ^HĂsNNĂOis. 2 CH 3 CO 2 H. %: 52.16% C, 6.68% H, 6.40% N, Found: 51.77% C, 6.79% H, 6.31% N. Example 19
Preparation of 3.2 *, 6'-tri-N-benzyloxycarbonyl-kanamycin B 500 mg, i.e. 1.03 mM kanamycin B free base, was suspended in all 12 ml of dimethyl sulfoxide and 4 ml of tetrahydrofuran and 1 g was added to the suspension. i.e. 4.55 mM zinc acetate dihydrate. The mixture was stirred at room temperature until a homogeneous solution was formed and cooled to 0 ° C. To a cold solution, a cold solution of 825 mg, i.e. 3.3 mM N-thion in 10 ml of tetrahydrofuran-dimethylsulfoxide 1: 1 was slowly added over more than one hour benzyloxycarbonyloxy succinimide. 0 ° C for two hours and then at ambient temperature overnight, then treated in the same manner as in Example 1 (ii) and (iii) (A) to obtain 740 mg, i.e. 70% of the desired compound in the form of a colorless solid.
[α] 25 25 + 63 ° (c 1, water dimethylformamide 1: 2).
Analysis
Calculated for C 42 H 56 N 3 O 6. 2 CH 3 CCCH. H, O: 53.95% C, 6.40% H, 6.84% N,
Found: 53.66% C, 6.67% H, 6.63% N.
Further treatment of the compound thus prepared by a procedure similar to that described in Example 31 gave 1N- [(S) -4-amino-2-hydroxy-butyryl] -canamycin B. 261853 47 48 Example 20 Preparation of 3.2 ', 6' tri N bonzyk) The microcrystalline 480 mg, i.e. 1.03 mM tobramycin in the free base was suspended in 12 ml of methyl sulfoxide and 1 g, i.e. 4.55 mM of zinc acetate dihydrate was added to the suspension. . The mixture was stirred at room temperature for one hour to form a homogeneous solution to which was then added for about one hour a solution of 850 milligrams, i.e. 3.4 mM of N-benzyloxycarbonyl oxysuineimide dissolved in 10 ml of tetrahydrofuran-dimethylsulphoxide 1: 1 mixture. The reaction solution was treated with a large volume of ethyl ether as in Example 1 (dense syrupy material).
The syrup was further processed in the same manner as in Example 1 (iiij (A) but using water-dioxane 1: 2 instead of 2: 1 to obtain 810 mg, i.e. 78% of the desired substance as a colorless solid).
[α] 25 D = 1-65 ° is 1, water-dimethylformamide, 1: 2].
Calculation Analysis »for Có, HróN, Or,. 2 CH-CO-d1. H-O: 54.81 o / o C, 6.50% II, 6.0>% N,
Found: 54.77% C, 6.71% II, 6.88% N.
The compound thus obtained can be further processed by a process similar to that of Example 31 to give 1N - [(S) -4-amino-2-hydroxybutyryl] -tobramycin. Example 21 Prepare! 3,2 ', 6'-tri-N-benzyloxycarbonyl-G1-N-methyltobramycin The desired compound in the form of a colorless solid was obtained in a yield of 890 mg, i.e. 84% by the same procedure as in Example 20 except that from 500 mg, i.e. 1.04 mM B'-N-methylbutabycin free base.
[α] D 25 -I 63 ° (c 1, water-dimethylformamide, 1: 2). Example 22
Preparation of 3,2 ', 6'-tri-N-benzyloxycarbonyl-4' - deoxycanamycin B
Starting from 480 mg, ie 1.03 mM 4-deoxy-kanamycin B free base [see Bulletin of the Chemical Society of Japan, Vol. 50, s. 2 362-2 368 (1977)], the desired compound was obtained as a colorless solid in a yield of 815 mg, i.e. 79% by the same procedure as in Example 20.
[Î] D -5 463 ° (c 1, water-dimethylformamide, 1: 2), Example 23 Preparation of 3,2 ', 6'-tri-N-benzyloxycarbonyl-dibecacin 000 mg, ie 1.33 mM dibecacin Free base (3 ', 4'-deoxycanamycin B) was suspended in 15 ml of dimethylsulfoxide and the suspension was stirred until a solution was added to which 1.4 g, ie 6.4 mM, was added zinc dihydrate dihydrate was added and a solution of 1.1 g, i.e. 4.4 mM of N-benzylcarbonylcarbonyl succinimide in 12 ml of dimethylsulfoxide was slowly added to the resulting solution for one hour, and the mixture was allowed to rest at ambient temperature overnight. Then, a large volume of ethyl ether was mixed with the reaction solution to separate the oily sediment containing, in particular, the N-benzyloxycarbonylated zinc complex as the desired product and a portion of the dimethyl sulfoxide, and then washed with ethyl ether to give a syrupy material.
This syrup was repeatedly washed with water to disintegrate the N-acylated zinc complex, and the free zinc cations were removed along with the initially existing zinc c-ct. In this way, 1.1 g of a water-insoluble solid was obtained, representing N-acylated dibecin. The solid was chromatographed on a thin layer silica gel using chloroform-ethanol-18% aqueous 1: 1: 1 low-phase solvent as the solvent to form a single spot at Rf 0.3, indicating that the solid was predominantly 3,2 ', 6'-tri-N-benzyloxycarbonyldibecacin is stopped.
Further treatment of the desired compound with a procedure similar to Example 31 yielded 1N - [[S] -4-amino-2-hydroxybutyryl] -dibecacin.
Further purification of the crude product obtained as described above was carried out by washing the compound with a 3 M ammonia solution to give a product free of zinc ions.
[α] D 25 + 71 ° (c 1, water-dimethylformamide, 1: 2). Example 24 Preparation of 3,2 ', 6'-tr-N-benzyloxycarbonyl-6'-N-methyldibecacin 267033 49 500 mg, ie 1.07 mM 6'-N-methyldibecacin in free base form and 1.2 g, i.e., 5.45 mM of zinc acetate dihydrate was dissolved! in 20 ml of dimethylsulfoxide, while slowly adding 910 mg, i.e. 3.6 mM of N-benzyloxycarbonyl-oxysuccinimide, for about 30 minutes. The reaction solution was allowed to stand at ambient temperature overnight and then treated in the same manner as in Example 23 to give 910 mg of the desired compound, which was essentially pure.
Further treatment of the compound so obtained gave 1N- [(S-4-amino-2-hydroxybutyryl] -6'-N-methyldibecacin).
Preparation of 3,2'-di-N-benzyloxycarbonylcanolamine C The desired compound in the form of a colored solid was obtained in a yield of 730 mg, i.e. 79% by the same procedure as in Example 1 (i), (ii) and (iiij (A)). ), but starting from 500 mg, ie 1.03 mM kanamycin C free base.
[Î] D = + 75 ° (c 1, water-dimethylformamide, 1: 2). Next, the compound thus obtained was worked up to obtain 1N - [(S-4-amino-2-hydroxybutyryl] -kanamycin C. Example 26).
Preparation of 6'-N-benzyloxycarbonylkanamycin A 500 mg, i.e. 1.03 mM kanamycin A in the free base form, was suspended in 20 billion dimethyl sulfoxide and 0.5 g, i.e. 2.3 mM, was added to the suspension. zinc acetate dihydrate. The mixture was stirred at room temperature until a homogeneous solution was formed, to which was then added 283 mg, i. 1.13 mM N-benzyloxycarbonyloxy-succinimide. The resulting mixture was allowed to cool overnight at ambient temperature and then treated in the same manner as in Example 1 (ii) and (iiij (1j) to give 556 mg of the desired compound as a colorless solid).
[a] D 2 R =, + g 2 «(C 1, water). Example 27 Preparation of 6'-N-benzyloxycarbonyldibecacin
In the same manner as in Example 26, 382 mg of the desired compound was obtained in 80 using 500 mg of dibecacin free base, 12 ml of dimethylsulfoxide, 0.7 g of acetic acid dihydrate and 305 mg of N-benzyloxycarbonyloxy succinimide.
[α] D 25 + 105 ° (c 0.5, water). Example 28
Preparation of 3,2 ', 6'-tri-N-benzyloxycarbonyl-3', 4'-dideoxy-3'-enocannamycin B 500 mg, ie 1.11 mM 3 ', 4'-dideoxy-3'-eno -kanamycin B free base (see "Bulletin of the Chemical Society of Ja-pan", Vol. 50, ppd 1,580-1,583 (1977)] was dissolved in 12 ml of dimethyl sulfoxide and 1 g was added to the solution. i.e., 4.55 mM of zinc acetate dihydrate, and the solution was stirred for one hour, followed by the slow addition of 870 mg, ie 3.49 mM N-benzyl-oxycarbonyl-oxysuccinimide, over 30 minutes to the resulting solution. overnight at ambient temperature and the reaction solution thus obtained was treated with a large volume of ethyl ether as in Example 1 (ii) to obtain a thick syrupy substance.
The syrup was further processed as in Example 1 (iiij (Bj but using water-dioxane 1: 2 instead of 2: 1 to give 784 mg of the desired compound as a colorless solid).
[α] D 25 + 30 ° (c 1, water-dimethylformamide, 1: 2). Example 29 Preparation of 3,2 ', 6'-tri-N-benzyloxycarbonylsomycin The desired compound as a colorless solid was obtained in a yield of 780 milligrams in the same manner as in Example 28 but with the difference that from 500 mg, i.e. 1.12 mM sisomicin free base.
[.alpha.] D @ 20 = + 110 DEG (c 1, water-diethylformamide, 1: 2). Example 30 Preparation of 3,2 ', 6'-tri-N-benzyloxycarbonyl-gentamicins 787 mg of the desired compound was obtained as a colorless solid in the same manner as in Example 28, except that 500 mg of genta-micine mixture was used C, Cla, C2 etc.
Claims (14)
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CS829399A CS261859B2 (en) | 1978-11-11 | 1982-12-20 | Method of aminoglycoside antibiotic's selectively protected n-acylated derivative preparation |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP13840278A JPS5564598A (en) | 1978-11-11 | 1978-11-11 | Preparation of aminoglycoside antibiotic having selectively protected amino group |
Publications (2)
Publication Number | Publication Date |
---|---|
CS771179A2 CS771179A2 (en) | 1988-06-15 |
CS261853B2 true CS261853B2 (en) | 1989-02-10 |
Family
ID=15221109
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CS797711A CS261853B2 (en) | 1978-11-11 | 1979-11-12 | Method of aminoglycoside antibiotic's selectively acylated n-protected derivative production |
Country Status (5)
Country | Link |
---|---|
JP (1) | JPS5564598A (en) |
BE (1) | BE879925A (en) |
CS (1) | CS261853B2 (en) |
HU (1) | HU183050B (en) |
ZA (1) | ZA795691B (en) |
Families Citing this family (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4136254A (en) * | 1976-06-17 | 1979-01-23 | Schering Corporation | Process of selectively blocking amino functions in aminoglycosides using transition metal salts and intermediates used thereby |
EP0446670B1 (en) * | 1990-03-08 | 1994-09-21 | Biochimica Opos Spa | Method of preparation of amikacin precursors |
KR100450607B1 (en) * | 2002-07-11 | 2004-09-30 | êČœëì ìœ ìŁŒìíìŹ | Process for preparing Galamin |
-
1978
- 1978-11-11 JP JP13840278A patent/JPS5564598A/en active Granted
-
1979
- 1979-10-24 ZA ZA00795691A patent/ZA795691B/en unknown
- 1979-11-09 BE BE2/58198A patent/BE879925A/en not_active IP Right Cessation
- 1979-11-09 HU HU79ZA414A patent/HU183050B/en unknown
- 1979-11-12 CS CS797711A patent/CS261853B2/en unknown
Also Published As
Publication number | Publication date |
---|---|
JPS631319B2 (en) | 1988-01-12 |
JPS5564598A (en) | 1980-05-15 |
BE879925A (en) | 1980-03-03 |
ZA795691B (en) | 1980-11-26 |
CS771179A2 (en) | 1988-06-15 |
HU183050B (en) | 1984-04-28 |
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