CN115232177A - 一种痢疾志贺氏菌10型o-抗原寡糖的化学合成方法 - Google Patents
一种痢疾志贺氏菌10型o-抗原寡糖的化学合成方法 Download PDFInfo
- Publication number
- CN115232177A CN115232177A CN202210933673.7A CN202210933673A CN115232177A CN 115232177 A CN115232177 A CN 115232177A CN 202210933673 A CN202210933673 A CN 202210933673A CN 115232177 A CN115232177 A CN 115232177A
- Authority
- CN
- China
- Prior art keywords
- reaction
- benzyl
- building block
- formula
- shigella dysenteriae
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000000034 method Methods 0.000 title claims abstract description 69
- 241000607764 Shigella dysenteriae Species 0.000 title claims abstract description 61
- 150000002482 oligosaccharides Chemical class 0.000 title claims abstract description 61
- 229940007046 shigella dysenteriae Drugs 0.000 title claims abstract description 61
- 239000000427 antigen Substances 0.000 title claims abstract description 55
- 238000003786 synthesis reaction Methods 0.000 title abstract description 61
- 229920001542 oligosaccharide Polymers 0.000 title abstract description 52
- 238000006243 chemical reaction Methods 0.000 claims abstract description 165
- 150000002772 monosaccharides Chemical class 0.000 claims abstract description 52
- 238000006206 glycosylation reaction Methods 0.000 claims abstract description 33
- 239000002994 raw material Substances 0.000 claims abstract description 6
- -1 2-naphthylmethyl Chemical group 0.000 claims description 71
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 51
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 40
- 238000010511 deprotection reaction Methods 0.000 claims description 33
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 31
- 239000002808 molecular sieve Substances 0.000 claims description 22
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 21
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 21
- 239000000243 solution Substances 0.000 claims description 19
- AYRSYEFPXJUEOE-UHFFFAOYSA-N ethylsulfanyl propanoate Chemical compound CCSOC(=O)CC AYRSYEFPXJUEOE-UHFFFAOYSA-N 0.000 claims description 17
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 claims description 16
- WJKHJLXJJJATHN-UHFFFAOYSA-N triflic anhydride Chemical compound FC(F)(F)S(=O)(=O)OS(=O)(=O)C(F)(F)F WJKHJLXJJJATHN-UHFFFAOYSA-N 0.000 claims description 16
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 15
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 claims description 15
- 125000001797 benzyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])* 0.000 claims description 14
- 229940076788 pyruvate Drugs 0.000 claims description 14
- LCTONWCANYUPML-UHFFFAOYSA-N Pyruvic acid Chemical compound CC(=O)C(O)=O LCTONWCANYUPML-UHFFFAOYSA-N 0.000 claims description 13
- 125000005647 linker group Chemical group 0.000 claims description 13
- KZMGYPLQYOPHEL-UHFFFAOYSA-N Boron trifluoride etherate Chemical group FB(F)F.CCOCC KZMGYPLQYOPHEL-UHFFFAOYSA-N 0.000 claims description 12
- 239000012190 activator Substances 0.000 claims description 12
- 125000006239 protecting group Chemical group 0.000 claims description 12
- 229940107700 pyruvic acid Drugs 0.000 claims description 12
- UTQNKKSJPHTPBS-UHFFFAOYSA-N 2,2,2-trichloroethanone Chemical group ClC(Cl)(Cl)[C]=O UTQNKKSJPHTPBS-UHFFFAOYSA-N 0.000 claims description 10
- LQZMLBORDGWNPD-UHFFFAOYSA-N N-iodosuccinimide Chemical compound IN1C(=O)CCC1=O LQZMLBORDGWNPD-UHFFFAOYSA-N 0.000 claims description 10
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 claims description 10
- 239000003153 chemical reaction reagent Substances 0.000 claims description 10
- URGAHOPLAPQHLN-UHFFFAOYSA-N sodium aluminosilicate Chemical compound [Na+].[Al+3].[O-][Si]([O-])=O.[O-][Si]([O-])=O URGAHOPLAPQHLN-UHFFFAOYSA-N 0.000 claims description 10
- YQTCQNIPQMJNTI-UHFFFAOYSA-N 2,2-dimethylpropan-1-one Chemical group CC(C)(C)[C]=O YQTCQNIPQMJNTI-UHFFFAOYSA-N 0.000 claims description 9
- 125000002777 acetyl group Chemical group [H]C([H])([H])C(*)=O 0.000 claims description 9
- 125000003236 benzoyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C(*)=O 0.000 claims description 9
- 125000002668 chloroacetyl group Chemical group ClCC(=O)* 0.000 claims description 9
- 125000005524 levulinyl group Chemical group 0.000 claims description 9
- WQDUMFSSJAZKTM-UHFFFAOYSA-N Sodium methoxide Chemical compound [Na+].[O-]C WQDUMFSSJAZKTM-UHFFFAOYSA-N 0.000 claims description 8
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 claims description 8
- 238000006722 reduction reaction Methods 0.000 claims description 8
- ITMCEJHCFYSIIV-UHFFFAOYSA-N triflic acid Chemical compound OS(=O)(=O)C(F)(F)F ITMCEJHCFYSIIV-UHFFFAOYSA-N 0.000 claims description 8
- FTVLMFQEYACZNP-UHFFFAOYSA-N trimethylsilyl trifluoromethanesulfonate Chemical compound C[Si](C)(C)OS(=O)(=O)C(F)(F)F FTVLMFQEYACZNP-UHFFFAOYSA-N 0.000 claims description 8
- 230000004913 activation Effects 0.000 claims description 7
- 238000005858 glycosidation reaction Methods 0.000 claims description 7
- XONPDZSGENTBNJ-UHFFFAOYSA-N molecular hydrogen;sodium Chemical compound [Na].[H][H] XONPDZSGENTBNJ-UHFFFAOYSA-N 0.000 claims description 7
- 230000008569 process Effects 0.000 claims description 7
- KWKAKUADMBZCLK-UHFFFAOYSA-N 1-octene Chemical compound CCCCCCC=C KWKAKUADMBZCLK-UHFFFAOYSA-N 0.000 claims description 6
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 claims description 6
- 125000004923 naphthylmethyl group Chemical group C1(=CC=CC2=CC=CC=C12)C* 0.000 claims description 6
- 230000021736 acetylation Effects 0.000 claims description 5
- 238000006640 acetylation reaction Methods 0.000 claims description 5
- 125000000852 azido group Chemical group *N=[N+]=[N-] 0.000 claims description 5
- MSWZFWKMSRAUBD-UHFFFAOYSA-N beta-D-galactosamine Natural products NC1C(O)OC(CO)C(O)C1O MSWZFWKMSRAUBD-UHFFFAOYSA-N 0.000 claims description 5
- MSWZFWKMSRAUBD-QZABAPFNSA-N beta-D-glucosamine Chemical compound N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O MSWZFWKMSRAUBD-QZABAPFNSA-N 0.000 claims description 5
- 125000005519 fluorenylmethyloxycarbonyl group Chemical group 0.000 claims description 5
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 5
- 230000009467 reduction Effects 0.000 claims description 5
- ITMCEJHCFYSIIV-UHFFFAOYSA-M triflate Chemical compound [O-]S(=O)(=O)C(F)(F)F ITMCEJHCFYSIIV-UHFFFAOYSA-M 0.000 claims description 5
- ZGEGCLOFRBLKSE-UHFFFAOYSA-N 1-Heptene Chemical compound CCCCCC=C ZGEGCLOFRBLKSE-UHFFFAOYSA-N 0.000 claims description 4
- AFFLGGQVNFXPEV-UHFFFAOYSA-N 1-decene Chemical compound CCCCCCCCC=C AFFLGGQVNFXPEV-UHFFFAOYSA-N 0.000 claims description 4
- LIKMAJRDDDTEIG-UHFFFAOYSA-N 1-hexene Chemical compound CCCCC=C LIKMAJRDDDTEIG-UHFFFAOYSA-N 0.000 claims description 4
- JRZJOMJEPLMPRA-UHFFFAOYSA-N 1-nonene Chemical compound CCCCCCCC=C JRZJOMJEPLMPRA-UHFFFAOYSA-N 0.000 claims description 4
- DCTOHCCUXLBQMS-UHFFFAOYSA-N 1-undecene Chemical compound CCCCCCCCCC=C DCTOHCCUXLBQMS-UHFFFAOYSA-N 0.000 claims description 4
- JYFHYPJRHGVZDY-UHFFFAOYSA-N Dibutyl phosphate Chemical compound CCCCOP(O)(=O)OCCCC JYFHYPJRHGVZDY-UHFFFAOYSA-N 0.000 claims description 4
- 238000006859 Swern oxidation reaction Methods 0.000 claims description 4
- 239000002253 acid Substances 0.000 claims description 4
- HUMNYLRZRPPJDN-UHFFFAOYSA-N benzaldehyde Chemical compound O=CC1=CC=CC=C1 HUMNYLRZRPPJDN-UHFFFAOYSA-N 0.000 claims description 4
- 125000000484 butyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 4
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 claims description 4
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 claims description 4
- CTSLXHKWHWQRSH-UHFFFAOYSA-N oxalyl chloride Chemical compound ClC(=O)C(Cl)=O CTSLXHKWHWQRSH-UHFFFAOYSA-N 0.000 claims description 4
- 125000001436 propyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 4
- 238000010791 quenching Methods 0.000 claims description 4
- LPXPTNMVRIOKMN-UHFFFAOYSA-M sodium nitrite Chemical compound [Na+].[O-]N=O LPXPTNMVRIOKMN-UHFFFAOYSA-M 0.000 claims description 4
- 239000000126 substance Substances 0.000 claims description 4
- 230000002194 synthesizing effect Effects 0.000 claims description 4
- 125000001981 tert-butyldimethylsilyl group Chemical group [H]C([H])([H])[Si]([H])(C([H])([H])[H])[*]C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H] 0.000 claims description 4
- 125000000037 tert-butyldiphenylsilyl group Chemical group [H]C1=C([H])C([H])=C([H])C([H])=C1[Si]([H])([*]C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H])C1=C([H])C([H])=C([H])C([H])=C1[H] 0.000 claims description 4
- LBRJCAJLGAXDKP-UHFFFAOYSA-N 1-(benzenesulfinyl)piperidine Chemical group C=1C=CC=CC=1S(=O)N1CCCCC1 LBRJCAJLGAXDKP-UHFFFAOYSA-N 0.000 claims description 3
- VYWSYEDVFVGRGG-UHFFFAOYSA-N 2,4,6-tritert-butylpyrimidine Chemical compound CC(C)(C)C1=CC(C(C)(C)C)=NC(C(C)(C)C)=N1 VYWSYEDVFVGRGG-UHFFFAOYSA-N 0.000 claims description 3
- LCTONWCANYUPML-UHFFFAOYSA-M Pyruvate Chemical compound CC(=O)C([O-])=O LCTONWCANYUPML-UHFFFAOYSA-M 0.000 claims description 3
- AGEZXYOZHKGVCM-UHFFFAOYSA-N benzyl bromide Chemical group BrCC1=CC=CC=C1 AGEZXYOZHKGVCM-UHFFFAOYSA-N 0.000 claims description 3
- KCXMKQUNVWSEMD-UHFFFAOYSA-N benzyl chloride Chemical compound ClCC1=CC=CC=C1 KCXMKQUNVWSEMD-UHFFFAOYSA-N 0.000 claims description 3
- 229940073608 benzyl chloride Drugs 0.000 claims description 3
- 125000001584 benzyloxycarbonyl group Chemical group C(=O)(OCC1=CC=CC=C1)* 0.000 claims description 3
- 125000001246 bromo group Chemical group Br* 0.000 claims description 3
- 239000003795 chemical substances by application Substances 0.000 claims description 3
- HPNMFZURTQLUMO-UHFFFAOYSA-N diethylamine Chemical compound CCNCC HPNMFZURTQLUMO-UHFFFAOYSA-N 0.000 claims description 3
- TXVLFCLSVCYBIV-UHFFFAOYSA-M dimethyl(methylsulfanyl)sulfanium;trifluoromethanesulfonate Chemical compound CS[S+](C)C.[O-]S(=O)(=O)C(F)(F)F TXVLFCLSVCYBIV-UHFFFAOYSA-M 0.000 claims description 3
- 238000005886 esterification reaction Methods 0.000 claims description 3
- 125000001153 fluoro group Chemical group F* 0.000 claims description 3
- 239000000937 glycosyl acceptor Substances 0.000 claims description 3
- XJTQJERLRPWUGL-UHFFFAOYSA-N iodomethylbenzene Chemical compound ICC1=CC=CC=C1 XJTQJERLRPWUGL-UHFFFAOYSA-N 0.000 claims description 3
- 239000012280 lithium aluminium hydride Substances 0.000 claims description 3
- OIRDBPQYVWXNSJ-UHFFFAOYSA-N methyl trifluoromethansulfonate Chemical group COS(=O)(=O)C(F)(F)F OIRDBPQYVWXNSJ-UHFFFAOYSA-N 0.000 claims description 3
- TVMXDCGIABBOFY-UHFFFAOYSA-N n-Octanol Natural products CCCCCCCC TVMXDCGIABBOFY-UHFFFAOYSA-N 0.000 claims description 3
- 125000001624 naphthyl group Chemical group 0.000 claims description 3
- LXNAVEXFUKBNMK-UHFFFAOYSA-N palladium(II) acetate Substances [Pd].CC(O)=O.CC(O)=O LXNAVEXFUKBNMK-UHFFFAOYSA-N 0.000 claims description 3
- YJVFFLUZDVXJQI-UHFFFAOYSA-L palladium(ii) acetate Chemical compound [Pd+2].CC([O-])=O.CC([O-])=O YJVFFLUZDVXJQI-UHFFFAOYSA-L 0.000 claims description 3
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 claims description 3
- UYWQUFXKFGHYNT-UHFFFAOYSA-N phenylmethyl ester of formic acid Natural products O=COCC1=CC=CC=C1 UYWQUFXKFGHYNT-UHFFFAOYSA-N 0.000 claims description 3
- 230000000171 quenching effect Effects 0.000 claims description 3
- 239000012279 sodium borohydride Substances 0.000 claims description 3
- 229910000033 sodium borohydride Inorganic materials 0.000 claims description 3
- 238000001308 synthesis method Methods 0.000 claims description 3
- XXRCUYVCPSWGCC-UHFFFAOYSA-N Ethyl pyruvate Chemical compound CCOC(=O)C(C)=O XXRCUYVCPSWGCC-UHFFFAOYSA-N 0.000 claims description 2
- AZFNGPAYDKGCRB-XCPIVNJJSA-M [(1s,2s)-2-amino-1,2-diphenylethyl]-(4-methylphenyl)sulfonylazanide;chlororuthenium(1+);1-methyl-4-propan-2-ylbenzene Chemical compound [Ru+]Cl.CC(C)C1=CC=C(C)C=C1.C1=CC(C)=CC=C1S(=O)(=O)[N-][C@@H](C=1C=CC=CC=1)[C@@H](N)C1=CC=CC=C1 AZFNGPAYDKGCRB-XCPIVNJJSA-M 0.000 claims description 2
- DHKHKXVYLBGOIT-UHFFFAOYSA-N acetaldehyde Diethyl Acetal Natural products CCOC(C)OCC DHKHKXVYLBGOIT-UHFFFAOYSA-N 0.000 claims description 2
- YFHNDHXQDJQEEE-UHFFFAOYSA-N acetic acid;hydrazine Chemical compound NN.CC(O)=O YFHNDHXQDJQEEE-UHFFFAOYSA-N 0.000 claims description 2
- 239000007864 aqueous solution Substances 0.000 claims description 2
- CNNBRFOPQDCGFV-UHFFFAOYSA-N benzyl 2-oxopropanoate Chemical compound CC(=O)C(=O)OCC1=CC=CC=C1 CNNBRFOPQDCGFV-UHFFFAOYSA-N 0.000 claims description 2
- ZAZUOXBHFXAWMD-UHFFFAOYSA-N butyl 2-oxopropanoate Chemical compound CCCCOC(=O)C(C)=O ZAZUOXBHFXAWMD-UHFFFAOYSA-N 0.000 claims description 2
- 229940117360 ethyl pyruvate Drugs 0.000 claims description 2
- CWKLZLBVOJRSOM-UHFFFAOYSA-N methyl pyruvate Chemical group COC(=O)C(C)=O CWKLZLBVOJRSOM-UHFFFAOYSA-N 0.000 claims description 2
- 239000003960 organic solvent Substances 0.000 claims description 2
- QNGNSVIICDLXHT-UHFFFAOYSA-N para-ethylbenzaldehyde Natural products CCC1=CC=C(C=O)C=C1 QNGNSVIICDLXHT-UHFFFAOYSA-N 0.000 claims description 2
- 125000004817 pentamethylene group Chemical group [H]C([H])([*:2])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[*:1] 0.000 claims description 2
- 239000004304 potassium nitrite Substances 0.000 claims description 2
- 235000010289 potassium nitrite Nutrition 0.000 claims description 2
- ILPVOWZUBFRIAX-UHFFFAOYSA-N propyl 2-oxopropanoate Chemical compound CCCOC(=O)C(C)=O ILPVOWZUBFRIAX-UHFFFAOYSA-N 0.000 claims description 2
- YWAKXRMUMFPDSH-UHFFFAOYSA-N propyl ethylene Natural products CCCC=C YWAKXRMUMFPDSH-UHFFFAOYSA-N 0.000 claims description 2
- 235000010288 sodium nitrite Nutrition 0.000 claims description 2
- SHRKDVQQQPFSIY-UHFFFAOYSA-M tetrabutylazanium;nitrite Chemical compound [O-]N=O.CCCC[N+](CCCC)(CCCC)CCCC SHRKDVQQQPFSIY-UHFFFAOYSA-M 0.000 claims description 2
- 125000001483 monosaccharide substituent group Chemical group 0.000 claims 7
- 230000003213 activating effect Effects 0.000 claims 2
- 125000003356 phenylsulfanyl group Chemical group [*]SC1=C([H])C([H])=C([H])C([H])=C1[H] 0.000 claims 1
- 230000013595 glycosylation Effects 0.000 abstract description 7
- 238000011161 development Methods 0.000 abstract description 6
- 229960005486 vaccine Drugs 0.000 abstract description 5
- 238000002360 preparation method Methods 0.000 abstract description 4
- 238000013461 design Methods 0.000 abstract description 2
- 238000001514 detection method Methods 0.000 abstract description 2
- 239000003814 drug Substances 0.000 abstract description 2
- 238000005457 optimization Methods 0.000 abstract 2
- 229940079593 drug Drugs 0.000 abstract 1
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 48
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 33
- 230000015572 biosynthetic process Effects 0.000 description 29
- 150000004044 tetrasaccharides Chemical group 0.000 description 24
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 18
- XMPZTFVPEKAKFH-UHFFFAOYSA-P ceric ammonium nitrate Chemical compound [NH4+].[NH4+].[Ce+4].[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O XMPZTFVPEKAKFH-UHFFFAOYSA-P 0.000 description 15
- 239000000370 acceptor Substances 0.000 description 14
- 150000002016 disaccharides Chemical class 0.000 description 14
- 239000002904 solvent Substances 0.000 description 14
- FPGGTKZVZWFYPV-UHFFFAOYSA-M tetrabutylammonium fluoride Chemical compound [F-].CCCC[N+](CCCC)(CCCC)CCCC FPGGTKZVZWFYPV-UHFFFAOYSA-M 0.000 description 14
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 13
- 150000001875 compounds Chemical class 0.000 description 12
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 11
- 238000002114 high-resolution electrospray ionisation mass spectrometry Methods 0.000 description 11
- XKRFYHLGVUSROY-UHFFFAOYSA-N Argon Chemical compound [Ar] XKRFYHLGVUSROY-UHFFFAOYSA-N 0.000 description 10
- 239000012043 crude product Substances 0.000 description 10
- 238000010898 silica gel chromatography Methods 0.000 description 10
- 239000011734 sodium Substances 0.000 description 10
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 9
- 239000003208 petroleum Substances 0.000 description 9
- KRHYYFGTRYWZRS-UHFFFAOYSA-N Fluorane Chemical compound F KRHYYFGTRYWZRS-UHFFFAOYSA-N 0.000 description 8
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 8
- 239000011541 reaction mixture Substances 0.000 description 8
- DKGAVHZHDRPRBM-UHFFFAOYSA-N tertiry butyl alcohol Natural products CC(C)(C)O DKGAVHZHDRPRBM-UHFFFAOYSA-N 0.000 description 8
- 239000000706 filtrate Substances 0.000 description 7
- 229920006395 saturated elastomer Polymers 0.000 description 7
- 150000004043 trisaccharides Chemical class 0.000 description 7
- HZNVUJQVZSTENZ-UHFFFAOYSA-N 2,3-dichloro-5,6-dicyano-1,4-benzoquinone Chemical compound ClC1=C(Cl)C(=O)C(C#N)=C(C#N)C1=O HZNVUJQVZSTENZ-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
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 6
- WMFOQBRAJBCJND-UHFFFAOYSA-M Lithium hydroxide Chemical compound [Li+].[OH-] WMFOQBRAJBCJND-UHFFFAOYSA-M 0.000 description 6
- 239000000348 glycosyl donor Substances 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 6
- 235000017557 sodium bicarbonate Nutrition 0.000 description 6
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 5
- 229910052786 argon Inorganic materials 0.000 description 5
- 238000010276 construction Methods 0.000 description 5
- 238000004821 distillation Methods 0.000 description 5
- 239000000386 donor Substances 0.000 description 5
- 238000001914 filtration Methods 0.000 description 5
- 239000000741 silica gel Substances 0.000 description 5
- 229910002027 silica gel Inorganic materials 0.000 description 5
- 229910052938 sodium sulfate Inorganic materials 0.000 description 5
- 235000011152 sodium sulphate Nutrition 0.000 description 5
- RMVRSNDYEFQCLF-UHFFFAOYSA-N thiophenol Chemical compound SC1=CC=CC=C1 RMVRSNDYEFQCLF-UHFFFAOYSA-N 0.000 description 5
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- UFWIBTONFRDIAS-UHFFFAOYSA-N Naphthalene Chemical compound C1=CC=CC2=CC=CC=C21 UFWIBTONFRDIAS-UHFFFAOYSA-N 0.000 description 4
- 241000607768 Shigella Species 0.000 description 4
- 230000009286 beneficial effect Effects 0.000 description 4
- 229910052799 carbon Inorganic materials 0.000 description 4
- 150000002148 esters Chemical class 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 4
- 150000004676 glycans Chemical class 0.000 description 4
- 239000011259 mixed solution Substances 0.000 description 4
- 229910052757 nitrogen Inorganic materials 0.000 description 4
- ILMRJRBKQSSXGY-UHFFFAOYSA-N tert-butyl(dimethyl)silicon Chemical group C[Si](C)C(C)(C)C ILMRJRBKQSSXGY-UHFFFAOYSA-N 0.000 description 4
- PMMURAAUARKVCB-ZXXMMSQZSA-N (2S,4R,5S,6R)-6-(hydroxymethyl)oxane-2,4,5-triol Chemical compound OC[C@H]1O[C@H](O)C[C@@H](O)[C@@H]1O PMMURAAUARKVCB-ZXXMMSQZSA-N 0.000 description 3
- 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 description 3
- PZUPAGRIHCRVKN-UHFFFAOYSA-N 5-[5-[3,4-dihydroxy-6-[(3,4,5-trihydroxyoxan-2-yl)oxymethyl]-5-[3,4,5-trihydroxy-6-[(3,4,5-trihydroxyoxan-2-yl)oxymethyl]oxan-2-yl]oxyoxan-2-yl]oxy-3,4-dihydroxy-6-[(3,4,5-trihydroxyoxan-2-yl)oxymethyl]oxan-2-yl]oxy-6-(hydroxymethyl)oxane-2,3,4-triol Chemical compound OCC1OC(O)C(O)C(O)C1OC1C(O)C(O)C(OC2C(C(O)C(OC3C(C(O)C(O)C(COC4C(C(O)C(O)CO4)O)O3)O)C(COC3C(C(O)C(O)CO3)O)O2)O)C(COC2C(C(O)C(O)CO2)O)O1 PZUPAGRIHCRVKN-UHFFFAOYSA-N 0.000 description 3
- PNNNRSAQSRJVSB-FSIIMWSLSA-N 6-deoxy-L-glucose Chemical compound C[C@H](O)[C@H](O)[C@@H](O)[C@H](O)C=O PNNNRSAQSRJVSB-FSIIMWSLSA-N 0.000 description 3
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 3
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 3
- 241000607760 Shigella sonnei Species 0.000 description 3
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 229940098773 bovine serum albumin Drugs 0.000 description 3
- 150000001720 carbohydrates Chemical group 0.000 description 3
- 238000009903 catalytic hydrogenation reaction Methods 0.000 description 3
- 229920001429 chelating resin Polymers 0.000 description 3
- 238000001816 cooling Methods 0.000 description 3
- 230000008878 coupling Effects 0.000 description 3
- 238000010168 coupling process Methods 0.000 description 3
- 238000005859 coupling reaction Methods 0.000 description 3
- UQLDLKMNUJERMK-UHFFFAOYSA-L di(octadecanoyloxy)lead Chemical compound [Pb+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O UQLDLKMNUJERMK-UHFFFAOYSA-L 0.000 description 3
- 238000003745 diagnosis Methods 0.000 description 3
- 238000010790 dilution Methods 0.000 description 3
- 239000012895 dilution Substances 0.000 description 3
- 125000003147 glycosyl group Chemical group 0.000 description 3
- 239000001257 hydrogen Substances 0.000 description 3
- 229910052739 hydrogen Inorganic materials 0.000 description 3
- 238000011534 incubation Methods 0.000 description 3
- 239000003456 ion exchange resin Substances 0.000 description 3
- 229920003303 ion-exchange polymer Polymers 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 238000001819 mass spectrum Methods 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 239000008363 phosphate buffer Substances 0.000 description 3
- 229920001282 polysaccharide Polymers 0.000 description 3
- 239000005017 polysaccharide Substances 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 229940115939 shigella sonnei Drugs 0.000 description 3
- QRUBYZBWAOOHSV-UHFFFAOYSA-M silver trifluoromethanesulfonate Chemical compound [Ag+].[O-]S(=O)(=O)C(F)(F)F QRUBYZBWAOOHSV-UHFFFAOYSA-M 0.000 description 3
- 238000001228 spectrum Methods 0.000 description 3
- 230000000707 stereoselective effect Effects 0.000 description 3
- 238000004809 thin layer chromatography Methods 0.000 description 3
- GHYOCDFICYLMRF-UTIIJYGPSA-N (2S,3R)-N-[(2S)-3-(cyclopenten-1-yl)-1-[(2R)-2-methyloxiran-2-yl]-1-oxopropan-2-yl]-3-hydroxy-3-(4-methoxyphenyl)-2-[[(2S)-2-[(2-morpholin-4-ylacetyl)amino]propanoyl]amino]propanamide Chemical compound C1(=CCCC1)C[C@@H](C(=O)[C@@]1(OC1)C)NC([C@H]([C@@H](C1=CC=C(C=C1)OC)O)NC([C@H](C)NC(CN1CCOCC1)=O)=O)=O GHYOCDFICYLMRF-UTIIJYGPSA-N 0.000 description 2
- UNILWMWFPHPYOR-KXEYIPSPSA-M 1-[6-[2-[3-[3-[3-[2-[2-[3-[[2-[2-[[(2r)-1-[[2-[[(2r)-1-[3-[2-[2-[3-[[2-(2-amino-2-oxoethoxy)acetyl]amino]propoxy]ethoxy]ethoxy]propylamino]-3-hydroxy-1-oxopropan-2-yl]amino]-2-oxoethyl]amino]-3-[(2r)-2,3-di(hexadecanoyloxy)propyl]sulfanyl-1-oxopropan-2-yl Chemical compound O=C1C(SCCC(=O)NCCCOCCOCCOCCCNC(=O)COCC(=O)N[C@@H](CSC[C@@H](COC(=O)CCCCCCCCCCCCCCC)OC(=O)CCCCCCCCCCCCCCC)C(=O)NCC(=O)N[C@H](CO)C(=O)NCCCOCCOCCOCCCNC(=O)COCC(N)=O)CC(=O)N1CCNC(=O)CCCCCN\1C2=CC=C(S([O-])(=O)=O)C=C2CC/1=C/C=C/C=C/C1=[N+](CC)C2=CC=C(S([O-])(=O)=O)C=C2C1 UNILWMWFPHPYOR-KXEYIPSPSA-M 0.000 description 2
- HZAXFHJVJLSVMW-UHFFFAOYSA-N 2-Aminoethan-1-ol Chemical compound NCCO HZAXFHJVJLSVMW-UHFFFAOYSA-N 0.000 description 2
- ASHGTJPOSUFTGB-UHFFFAOYSA-N 3-methoxyphenol Chemical compound COC1=CC=CC(O)=C1 ASHGTJPOSUFTGB-UHFFFAOYSA-N 0.000 description 2
- RBWNDBNSJFCLBZ-UHFFFAOYSA-N 7-methyl-5,6,7,8-tetrahydro-3h-[1]benzothiolo[2,3-d]pyrimidine-4-thione Chemical compound N1=CNC(=S)C2=C1SC1=C2CCC(C)C1 RBWNDBNSJFCLBZ-UHFFFAOYSA-N 0.000 description 2
- WHVNXSBKJGAXKU-UHFFFAOYSA-N Alexa Fluor 532 Chemical compound [H+].[H+].CC1(C)C(C)NC(C(=C2OC3=C(C=4C(C(C(C)N=4)(C)C)=CC3=3)S([O-])(=O)=O)S([O-])(=O)=O)=C1C=C2C=3C(C=C1)=CC=C1C(=O)ON1C(=O)CCC1=O WHVNXSBKJGAXKU-UHFFFAOYSA-N 0.000 description 2
- SHZGCJCMOBCMKK-UHFFFAOYSA-N D-mannomethylose Natural products CC1OC(O)C(O)C(O)C1O SHZGCJCMOBCMKK-UHFFFAOYSA-N 0.000 description 2
- PXGOKWXKJXAPGV-UHFFFAOYSA-N Fluorine Chemical compound FF PXGOKWXKJXAPGV-UHFFFAOYSA-N 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Chemical compound OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 2
- PNNNRSAQSRJVSB-UHFFFAOYSA-N L-rhamnose Natural products CC(O)C(O)C(O)C(O)C=O PNNNRSAQSRJVSB-UHFFFAOYSA-N 0.000 description 2
- 229910019142 PO4 Inorganic materials 0.000 description 2
- 241000607766 Shigella boydii Species 0.000 description 2
- 241000607762 Shigella flexneri Species 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 2
- WETWJCDKMRHUPV-UHFFFAOYSA-N acetyl chloride Chemical compound CC(Cl)=O WETWJCDKMRHUPV-UHFFFAOYSA-N 0.000 description 2
- 239000012346 acetyl chloride Substances 0.000 description 2
- PNNNRSAQSRJVSB-BXKVDMCESA-N aldehydo-L-rhamnose Chemical group C[C@H](O)[C@H](O)[C@@H](O)[C@@H](O)C=O PNNNRSAQSRJVSB-BXKVDMCESA-N 0.000 description 2
- 125000003277 amino group Chemical group 0.000 description 2
- 125000004429 atom Chemical group 0.000 description 2
- 238000005574 benzylation reaction Methods 0.000 description 2
- 125000002837 carbocyclic group Chemical group 0.000 description 2
- 235000014633 carbohydrates Nutrition 0.000 description 2
- 125000004432 carbon atom Chemical group C* 0.000 description 2
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 2
- 239000003054 catalyst Substances 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- 229940125797 compound 12 Drugs 0.000 description 2
- SQNZJJAZBFDUTD-UHFFFAOYSA-N durene Chemical compound CC1=CC(C)=C(C)C=C1C SQNZJJAZBFDUTD-UHFFFAOYSA-N 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 229910052731 fluorine Inorganic materials 0.000 description 2
- 239000011737 fluorine Substances 0.000 description 2
- 125000005842 heteroatom Chemical group 0.000 description 2
- 238000002329 infrared spectrum Methods 0.000 description 2
- 238000006386 neutralization reaction Methods 0.000 description 2
- QMMRZOWCJAIUJA-UHFFFAOYSA-L nickel dichloride Chemical compound Cl[Ni]Cl QMMRZOWCJAIUJA-UHFFFAOYSA-L 0.000 description 2
- FEMOMIGRRWSMCU-UHFFFAOYSA-N ninhydrin Chemical compound C1=CC=C2C(=O)C(O)(O)C(=O)C2=C1 FEMOMIGRRWSMCU-UHFFFAOYSA-N 0.000 description 2
- 230000003287 optical effect Effects 0.000 description 2
- 229910052760 oxygen Inorganic materials 0.000 description 2
- KDLHZDBZIXYQEI-UHFFFAOYSA-N palladium Substances [Pd] KDLHZDBZIXYQEI-UHFFFAOYSA-N 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 2
- 239000010452 phosphate Substances 0.000 description 2
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Chemical compound [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 description 2
- WGYKZJWCGVVSQN-UHFFFAOYSA-N propylamine Chemical group CCCN WGYKZJWCGVVSQN-UHFFFAOYSA-N 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 229920005989 resin Polymers 0.000 description 2
- 239000011347 resin Substances 0.000 description 2
- 208000026775 severe diarrhea Diseases 0.000 description 2
- AKHNMLFCWUSKQB-UHFFFAOYSA-L sodium thiosulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=S AKHNMLFCWUSKQB-UHFFFAOYSA-L 0.000 description 2
- 235000019345 sodium thiosulphate Nutrition 0.000 description 2
- 238000003756 stirring Methods 0.000 description 2
- 125000001424 substituent group Chemical group 0.000 description 2
- 229910052717 sulfur Inorganic materials 0.000 description 2
- 230000009897 systematic effect Effects 0.000 description 2
- 125000000999 tert-butyl group Chemical group [H]C([H])([H])C(*)(C([H])([H])[H])C([H])([H])[H] 0.000 description 2
- XROWMBWRMNHXMF-UHFFFAOYSA-J titanium tetrafluoride Chemical compound [F-].[F-].[F-].[F-].[Ti+4] XROWMBWRMNHXMF-UHFFFAOYSA-J 0.000 description 2
- YWWDBCBWQNCYNR-UHFFFAOYSA-N trimethylphosphine Chemical compound CP(C)C YWWDBCBWQNCYNR-UHFFFAOYSA-N 0.000 description 2
- RIOQSEWOXXDEQQ-UHFFFAOYSA-N triphenylphosphine Chemical compound C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1 RIOQSEWOXXDEQQ-UHFFFAOYSA-N 0.000 description 2
- QFLWZFQWSBQYPS-AWRAUJHKSA-N (3S)-3-[[(2S)-2-[[(2S)-2-[5-[(3aS,6aR)-2-oxo-1,3,3a,4,6,6a-hexahydrothieno[3,4-d]imidazol-4-yl]pentanoylamino]-3-methylbutanoyl]amino]-3-(4-hydroxyphenyl)propanoyl]amino]-4-[1-bis(4-chlorophenoxy)phosphorylbutylamino]-4-oxobutanoic acid Chemical compound CCCC(NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](Cc1ccc(O)cc1)NC(=O)[C@@H](NC(=O)CCCCC1SC[C@@H]2NC(=O)N[C@H]12)C(C)C)P(=O)(Oc1ccc(Cl)cc1)Oc1ccc(Cl)cc1 QFLWZFQWSBQYPS-AWRAUJHKSA-N 0.000 description 1
- OHWCAVRRXKJCRB-UISSTJQFSA-N (3r,4r,5r,6s)-2-methoxy-6-methyloxane-3,4,5-triol Chemical compound COC1O[C@@H](C)[C@H](O)[C@@H](O)[C@H]1O OHWCAVRRXKJCRB-UISSTJQFSA-N 0.000 description 1
- UPQQXPKAYZYUKO-UHFFFAOYSA-N 2,2,2-trichloroacetamide Chemical compound OC(=N)C(Cl)(Cl)Cl UPQQXPKAYZYUKO-UHFFFAOYSA-N 0.000 description 1
- SAPQIENQEZURNZ-UHFFFAOYSA-N 2,2,2-trifluoro-n-phenylacetamide Chemical compound FC(F)(F)C(=O)NC1=CC=CC=C1 SAPQIENQEZURNZ-UHFFFAOYSA-N 0.000 description 1
- MSWZFWKMSRAUBD-CBPJZXOFSA-N 2-amino-2-deoxy-D-mannopyranose Chemical compound N[C@@H]1C(O)O[C@H](CO)[C@@H](O)[C@@H]1O MSWZFWKMSRAUBD-CBPJZXOFSA-N 0.000 description 1
- 125000003903 2-propenyl group Chemical group [H]C([*])([H])C([H])=C([H])[H] 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- WKBOTKDWSSQWDR-UHFFFAOYSA-N Bromine atom Chemical compound [Br] WKBOTKDWSSQWDR-UHFFFAOYSA-N 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-M Chloride anion Chemical compound [Cl-] VEXZGXHMUGYJMC-UHFFFAOYSA-M 0.000 description 1
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- KIWBPDUYBMNFTB-UHFFFAOYSA-N Ethyl hydrogen sulfate Chemical compound CCOS(O)(=O)=O KIWBPDUYBMNFTB-UHFFFAOYSA-N 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 101150084935 PTER gene Proteins 0.000 description 1
- PEGDPHGOIZTRIT-UHFFFAOYSA-L S(=O)(=O)([O-])[O-].[Ce+2] Chemical compound S(=O)(=O)([O-])[O-].[Ce+2] PEGDPHGOIZTRIT-UHFFFAOYSA-L 0.000 description 1
- 101100219435 Shigella flexneri can gene Proteins 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 239000004098 Tetracycline Substances 0.000 description 1
- 229910021626 Tin(II) chloride Inorganic materials 0.000 description 1
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 1
- UTPYQWVXECMZMA-UHFFFAOYSA-N [dimethyl(sulfo)silyl]methane Chemical compound C[Si](C)(C)S(O)(=O)=O UTPYQWVXECMZMA-UHFFFAOYSA-N 0.000 description 1
- 150000001241 acetals Chemical group 0.000 description 1
- 125000000738 acetamido group Chemical group [H]C([H])([H])C(=O)N([H])[*] 0.000 description 1
- GZCGUPFRVQAUEE-KVTDHHQDSA-N aldehydo-D-mannose Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)C=O GZCGUPFRVQAUEE-KVTDHHQDSA-N 0.000 description 1
- PMMURAAUARKVCB-UHFFFAOYSA-N alpha-D-ara-dHexp Natural products OCC1OC(O)CC(O)C1O PMMURAAUARKVCB-UHFFFAOYSA-N 0.000 description 1
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 1
- 229910052782 aluminium Inorganic materials 0.000 description 1
- 239000011609 ammonium molybdate Substances 0.000 description 1
- APUPEJJSWDHEBO-UHFFFAOYSA-P ammonium molybdate Chemical compound [NH4+].[NH4+].[O-][Mo]([O-])(=O)=O APUPEJJSWDHEBO-UHFFFAOYSA-P 0.000 description 1
- 235000018660 ammonium molybdate Nutrition 0.000 description 1
- 229940010552 ammonium molybdate Drugs 0.000 description 1
- 229960000723 ampicillin Drugs 0.000 description 1
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 239000008346 aqueous phase Substances 0.000 description 1
- 125000003118 aryl group Chemical group 0.000 description 1
- IVRMZWNICZWHMI-UHFFFAOYSA-N azide group Chemical group [N-]=[N+]=[N-] IVRMZWNICZWHMI-UHFFFAOYSA-N 0.000 description 1
- 229960004099 azithromycin Drugs 0.000 description 1
- MQTOSJVFKKJCRP-BICOPXKESA-N azithromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)N(C)C[C@H](C)C[C@@](C)(O)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 MQTOSJVFKKJCRP-BICOPXKESA-N 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- GDTBXPJZTBHREO-UHFFFAOYSA-N bromine Substances BrBr GDTBXPJZTBHREO-UHFFFAOYSA-N 0.000 description 1
- 229910052794 bromium Inorganic materials 0.000 description 1
- 125000006278 bromobenzyl group Chemical group 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- DKPFZGUDAPQIHT-UHFFFAOYSA-N butyl acetate Chemical compound CCCCOC(C)=O DKPFZGUDAPQIHT-UHFFFAOYSA-N 0.000 description 1
- 239000000969 carrier Substances 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 229960005091 chloramphenicol Drugs 0.000 description 1
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 1
- CZKMPDNXOGQMFW-UHFFFAOYSA-N chloro(triethyl)germane Chemical compound CC[Ge](Cl)(CC)CC CZKMPDNXOGQMFW-UHFFFAOYSA-N 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 229940125904 compound 1 Drugs 0.000 description 1
- 229940125782 compound 2 Drugs 0.000 description 1
- 229940126214 compound 3 Drugs 0.000 description 1
- 229910052802 copper Inorganic materials 0.000 description 1
- 239000010949 copper Substances 0.000 description 1
- 238000012217 deletion Methods 0.000 description 1
- 230000037430 deletion Effects 0.000 description 1
- 108010001092 disaccharide receptor Proteins 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- KPUWHANPEXNPJT-UHFFFAOYSA-N disiloxane Chemical group [SiH3]O[SiH3] KPUWHANPEXNPJT-UHFFFAOYSA-N 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 239000002158 endotoxin Substances 0.000 description 1
- 125000001033 ether group Chemical group 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000011888 foil Substances 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 150000002431 hydrogen Chemical class 0.000 description 1
- OUUQCZGPVNCOIJ-UHFFFAOYSA-N hydroperoxyl Chemical compound O[O] OUUQCZGPVNCOIJ-UHFFFAOYSA-N 0.000 description 1
- 230000003100 immobilizing effect Effects 0.000 description 1
- 230000005965 immune activity Effects 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 239000011981 lindlar catalyst Substances 0.000 description 1
- 229920006008 lipopolysaccharide Polymers 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- UKVIEHSSVKSQBA-UHFFFAOYSA-N methane;palladium Chemical compound C.[Pd] UKVIEHSSVKSQBA-UHFFFAOYSA-N 0.000 description 1
- 239000002480 mineral oil Substances 0.000 description 1
- 235000010446 mineral oil Nutrition 0.000 description 1
- 230000003472 neutralizing effect Effects 0.000 description 1
- 239000012074 organic phase Substances 0.000 description 1
- NXJCBFBQEVOTOW-UHFFFAOYSA-L palladium(2+);dihydroxide Chemical compound O[Pd]O NXJCBFBQEVOTOW-UHFFFAOYSA-L 0.000 description 1
- PIBWKRNGBLPSSY-UHFFFAOYSA-L palladium(II) chloride Chemical compound Cl[Pd]Cl PIBWKRNGBLPSSY-UHFFFAOYSA-L 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 239000008055 phosphate buffer solution Substances 0.000 description 1
- 229960003342 pivampicillin Drugs 0.000 description 1
- ZEMIJUDPLILVNQ-ZXFNITATSA-N pivampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@@H]3N(C2=O)[C@H](C(S3)(C)C)C(=O)OCOC(=O)C(C)(C)C)=CC=CC=C1 ZEMIJUDPLILVNQ-ZXFNITATSA-N 0.000 description 1
- 150000004804 polysaccharides Polymers 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 229910000027 potassium carbonate Inorganic materials 0.000 description 1
- ZJLMKPKYJBQJNH-UHFFFAOYSA-N propane-1,3-dithiol Chemical compound SCCCS ZJLMKPKYJBQJNH-UHFFFAOYSA-N 0.000 description 1
- KOUKXHPPRFNWPP-UHFFFAOYSA-N pyrazine-2,5-dicarboxylic acid;hydrate Chemical compound O.OC(=O)C1=CN=C(C(O)=O)C=N1 KOUKXHPPRFNWPP-UHFFFAOYSA-N 0.000 description 1
- 230000009257 reactivity Effects 0.000 description 1
- 238000012772 sequence design Methods 0.000 description 1
- 239000001632 sodium acetate Substances 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical class O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 1
- 125000006850 spacer group Chemical group 0.000 description 1
- 235000011150 stannous chloride Nutrition 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- 238000005556 structure-activity relationship Methods 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- FDDDEECHVMSUSB-UHFFFAOYSA-N sulfanilamide Chemical compound NC1=CC=C(S(N)(=O)=O)C=C1 FDDDEECHVMSUSB-UHFFFAOYSA-N 0.000 description 1
- 229940124530 sulfonamide Drugs 0.000 description 1
- 229960002180 tetracycline Drugs 0.000 description 1
- 229930101283 tetracycline Natural products 0.000 description 1
- 235000019364 tetracycline Nutrition 0.000 description 1
- 150000003522 tetracyclines Chemical class 0.000 description 1
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- AXZWODMDQAVCJE-UHFFFAOYSA-L tin(II) chloride (anhydrous) Chemical compound [Cl-].[Cl-].[Sn+2] AXZWODMDQAVCJE-UHFFFAOYSA-L 0.000 description 1
- BDZBKCUKTQZUTL-UHFFFAOYSA-N triethyl phosphite Chemical compound CCOP(OCC)OCC BDZBKCUKTQZUTL-UHFFFAOYSA-N 0.000 description 1
- XSQUKJJJFZCRTK-UHFFFAOYSA-N urea group Chemical group NC(=O)N XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 1
- 229940125575 vaccine candidate Drugs 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
-
- 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
- C07H15/02—Acyclic radicals, not substituted by cyclic structures
- C07H15/04—Acyclic radicals, not substituted by cyclic structures attached to an oxygen atom of the saccharide radical
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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)
- Crystallography & Structural Chemistry (AREA)
- Saccharide Compounds (AREA)
Abstract
Description
技术领域
本发明涉及一种痢疾志贺氏菌10型O-抗原寡糖的化学合成方法,属于化学技术领域。
背景技术
重症腹泻是五岁以下儿童的第二大致死原因。志贺氏菌是导致儿童重症腹泻的第二大致病菌,可分为痢疾志贺氏菌,福氏志贺氏菌,鲍氏志贺氏菌和宋内氏志贺氏菌。基于细胞表面脂多糖O-抗原结构,痢疾志贺氏菌可进一步分为15种血清型,福氏志贺氏菌可分为6种血清型和12个亚型,鲍氏志贺氏菌可分为20种血清型,宋内氏志贺氏菌则只有一种血清型。研究表明,志贺氏菌对于磺胺,四环素,氯霉素,氨苄西林,皮夫梅西林和阿奇霉素等各种治疗用抗生素具有耐药性。
志贺氏菌O-抗原的蛋白缀合物是最为领先的疫苗候选物。已证实痢疾志贺氏菌1型O-抗原,福氏志贺氏菌2a和6型O-抗原,以及宋内氏志贺氏菌O-抗原是志贺氏菌疫苗开发的有效免疫原。
由于多糖提取过程中存在关键表位改变和不稳定性取代基缺失的问题,结构明确的合成寡糖已在糖缀合疫苗开发领域发挥重要作用。痢疾志贺氏菌10血清型O-抗原由四糖重复单元[→2)-β-D-Manp4,6(S)Pyr-(1→3)-α-D-ManpNAc-(1→3)-β-L-Rhap-(1→4)-α-D-GlcpNAc-(1→](如图1所示)组成,其中D-甘露糖修饰有(S)-4,6-O-丙酮酸缩酮基。痢疾志贺氏菌10血清型O-抗原四糖重复单元含有三个1,2-顺式-糖苷键,包括一个β-D-甘露糖苷键、一个β-L-鼠李糖苷键和一个α-D-氨基葡萄糖苷键。四糖的立体专一性组装需要对不同糖基化策略进行合理组合。所以,痢疾志贺氏菌10血清型O-抗原四糖的化学合成仍具有很大挑战性。首先,在一个四糖分子中完成两个1,2-顺式-β-糖苷键和一个1,2-顺式-α-糖苷键的构建是一个系统性问题,而非单一的糖苷键构建。此外,(S)-4,6-O-丙酮酸缩酮基对于糖基化反应的立体选择性和糖基连接效率存在潜在影响,需对四糖组装序列开展合理设计。
发明内容
技术问题:
本发明涉及痢疾志贺氏菌10血清型O-抗原的组装有连接臂的寡糖片段的化学合成方法。该多糖的四糖重复单元含有三个具有很大合成难度的1,2-顺式-糖苷键,需要解决各个糖苷键的立体专一性构建的技术问题。尤其在一个四糖分子中完成两个1,2-顺式-β-糖苷键和一个1,2-顺式-α-糖苷键的构建是一个系统性问题,需要综合运用各种糖基化策略,并合理设计寡糖组装序列,避免不同糖苷键构建过程中相互制约的问题。多糖结构中(S)-4,6-O-丙酮酸缩酮基对于糖基化反应的立体选择性和糖基连接效率存在潜在影响,进一步提高了四糖组装序列设计的难度。本发明要达到的目标为综合运用各种糖基化策略,实现糖链的立体专一性组装;探究(S)-4,6-O-丙酮酸缩酮基对于糖基化反应的影响,合理设计寡糖组装序列;开展天然结构及其衍生物制备,利用糖芯片分析解析合成寡糖及其(S)-4,6-O-丙酮酸缩酮基修饰的免疫活性。
技术方案:
本发明首次完成了痢疾志贺氏菌10血清型O-抗原寡糖的化学合成。此外通过在糖类结构还原端引入相应连接臂,为合成寡糖结构偶联载体分子或固定化于相应基质提供基础。通过糖芯片实验探究了合成寡糖的抗原性,将有助于痢疾志贺氏菌诊断技术和糖类疫苗的开发。
本发明提供一种痢疾志贺氏菌10血清型O-抗原的组装有连接臂的寡糖片段的合成方法,所述寡糖片段的结构如通式I所示:
V*-[Ux+3-Ux+2-Ux+1-Ux]n-V-O-L-NH2通式I;
其中,x为1,2,3;n为1,2,3;-V-表示:化学键,-Ux+3-,-Ux+3-Ux+2-,或-Ux+3-Ux+2-Ux+1-;V*-表示:H-,H-Ux-,H-Ux+1-Ux-,或H-Ux+2-Ux+1-Ux-;L表示连接臂;Ux,Ux+1,Ux+2,Ux+3的结构如下所示:
所述合成方法以四个单糖砌块1,2,3,4和连接臂5为原料,进行多步糖苷化反应,最后脱保护;
其中:
PG1,PG6,PG8,PG9,PG11为苄基;PG2,PG5为2-萘甲基,甲氧苄基,叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基;PG3为乙酰基,苯甲酰基,氯乙酰基,二氯乙酰基,三氯乙酰基,新戊酰基,烯丙氧羰酰基;PG4为乙酰丙酰基,芴甲氧羰基;PG7为2-萘甲基,甲氧苄基,叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基,乙酰基,苯甲酰基,乙酰丙酰基,氯乙酰基,二氯乙酰基,三氯乙酰基,新戊酰基,烯丙氧羰酰基;PG12为苯基,萘基;PG10为苄基,2-萘甲基,对甲氧苄基;PG13,PG14为苄基,苄氧羰基;LG1为乙硫基,对甲苯硫基,苯硫基,硒苯基,溴,氟,三氯乙酰亚氨酯,N-苯基三氟乙酰亚胺酯,二丁基磷酸酯;LG2为乙硫基,对甲苯硫基,苯硫基。
本发明中连接臂L可以是2-45碳原子数(包括侧链的碳原子数)的链式结构。
本发明中连接臂的主链长为4-9原子数时,链中可以包含1、2或3个杂原子(O,N和S)。当连接臂的主链长为10-14原子数时,链中可以包含1、2、3、4、5或6个杂原子(O,N和S)。
本发明中连接臂-L-可以全部或部分氟取代。连接臂-L-可以包含一个三、四、五或六元饱和碳环;也可以包含一个五元不饱和碳环(非芳香环);也可以包含四、五或六元饱和氧杂环;也可以包含一个四、五或六元饱和氮杂环;也可以包含一个六元芳香碳环。
本发明中连接臂-L-也可以包含酰胺键和/或脲基。
本发明中连接臂-L-可以含有一个或多个取代基团,这些取代基可以包括:-F,-Cl,-CH3,-C2H5,-NHC(O)CH3,-N(CH3)2,-C3H7,-CH2F,-CHF2,-CF3,-C5H9,-C6H13,-OCH3,-OC2H5,-C(O)-NH2,-SCH3,-SC2H5和-N(C2H5)2。
在本发明的一种实施方式中,单糖砌块(Ux,Ux+1,Ux+2,Ux+3)之间的连接方式均为一单糖砌块通过端基位碳(1号位碳)与另一单糖砌块的相应羟基氧所形成的糖苷键。
在本发明的一种实施方式中,本发明中涉及的痢疾志贺氏菌10血清型O-抗原糖链可以表示为通式II:V*-[Ux+3-Ux+2-Ux+1-Ux]n-O-L-NH2,其中x,n,L,Ux,Ux+1,Ux+2,Ux+3和V*与通式I一致;通式II可以具体表示为通式II-a,II-b,II-c和II-d:
在本发明的一种实施方式中,本发明中涉及的痢疾志贺氏菌10血清型O-抗原糖链可以表示为通式III:V*-[Ux+3-Ux+2-Ux+1-Ux]n-Ux+3-O-L-NH2,其中x,n,L,Ux,Ux+1,Ux+2,Ux+3和V*与通式I一致;通式III可以具体表示为通式III-a,III-b,III-c和III-d:
在本发明的一种实施方式中,本发明中涉及的痢疾志贺氏菌10血清型O-抗原糖链可以表示为通式IV:V*-[Ux+3-Ux+2-Ux+1-Ux]n-Ux+3-Ux+2-O-L-NH2,其中x,n,L,Ux,Ux+1,Ux+2,Ux+3和V*与通式I一致;通式IV可以具体表示为通式IV-a,IV-b,IV-c和IV-d:
在本发明的一种实施方式中,本发明中涉及的痢疾志贺氏菌10血清型O-抗原糖链可以表示为通式V:V*-[Ux+3-Ux+2-Ux+1-Ux]n-Ux+3-Ux+2-Ux+1-O-L-NH2,其中x,n,L,Ux,Ux+1,Ux+2,Ux+3和V*与通式I一致;通式V可以具体表示为通式V-a,V-b,V-c和V-d:
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原的连接臂修饰糖链(通式I)的化学合成方法,其特征在于,以四个单糖砌块1,2,3,4和连接臂5为原料:
其中:
PG1,PG6,PG8,PG9,PG11为羟基保护基团,可以是苄基。
PG2,PG5为羟基保护基团,可以是2-萘甲基,甲氧苄基,叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基。
PG3为羟基保护基团,可以是乙酰基,苯甲酰基,氯乙酰基,二氯乙酰基,三氯乙酰基,新戊酰基,烯丙氧羰酰基。
PG4为羟基保护基团,可以是乙酰丙酰基,芴甲氧羰基。
PG7为羟基保护基团,可以是2-萘甲基,甲氧苄基,叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基,乙酰基,苯甲酰基,乙酰丙酰基,氯乙酰基,二氯乙酰基,三氯乙酰基,新戊酰基,烯丙氧羰酰基。
PG12为羟基保护基团,可以是苯基,萘基。
PG10为羟基保护基团,可以是苄基,2-萘甲基,对甲氧苄基。
PG13,PG14为氨基保护基团,可以是苄基,苄氧羰基。
LG1为用于常规糖基化反应的离去基团,可以是乙硫基,对甲苯硫基,苯硫基,硒苯基,溴,氟,三氯乙酰亚氨酯,N-苯基三氟乙酰亚胺酯,二丁基磷酸酯。
LG2为用于预活化糖基化反应的离去基团,可以是乙硫基,对甲苯硫基,苯硫基。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,其特征包括如下反应模块:
1)反应模块A:常规糖苷化反应
当糖苷化反应中所用离去基团为乙硫基,对甲苯硫基,苯硫基时,糖苷化反应中活化剂可以选自三氟甲磺酸甲酯(TfOMe),二甲基甲硫基硫鎓三氟甲磺酸盐(DMTST),三氟甲磺酸(TfOH)/N-碘代丁二酰亚胺(NIS),三氟甲磺酸三甲基硅酯(TMSOTf)/N-碘代丁二酰亚胺(NIS),反应温度可以是-40℃至室温之间;
当离去基是氟时,糖苷化反应中活化剂可以是高氯酸银(AgClO4),四氟化钛(TiF4),三氟甲磺酸酐(Tf2O)等,反应温度可以是-40℃至室温之间;
当离去基是溴时,糖苷化反应中活化剂可以是高氯酸银(AgClO4),三氟甲磺酸银(AgOTf),反应温度可以是-40℃至室温之间;
当离去基团是三氯乙酰亚胺酯,N-苯基三氟乙酰亚胺酯,二丁基磷酸酯时,糖苷化反应中活化剂可以是三氟化硼乙醚络合物(BF3·OEt2),三氟甲磺酸三甲基硅酯(TMSOTf),三氟甲磺酸银(AgOTf),反应温度可以是-40℃至室温之间。
2)反应模块B:预活化糖苷化反应
供体活化剂可以是1-(苯基亚硫酰基)哌啶(BSP)/2,4,6-三叔丁基嘧啶(TTBP)/三氟甲磺酸酐(Tf2O),活化温度为-80至-40℃,活化时间为20至60分钟。
活化剂的淬灭剂可以是1-戊烯,1-己烯,1-庚烯,1-辛烯,1-壬烯,1-癸烯,1-十一碳烯,反应温度为-80至-40℃,处理时间为5至40分钟。
糖基受体在温度-100至-60℃下加入预活化的供体溶液中进行糖苷化反应。
3)反应模块C:叠氮基还原乙酰化
叠氮基还原乙酰化可以通过硫代乙酸(AcSH)/吡啶方法直接完成,也可以经叠氮基还原为氨基,再乙酰化完成。叠氮基还原为氨基的方法可以是锌/铜/醋酸,三甲基膦(PMe3)/水,1,3-丙二硫醇/三乙胺,三苯基膦(PPh3)/水,Lindlar催化剂/氢气,二氯化锡(SnCl2)/苯硫醇(PhSH)/三乙胺,硼氢化钠(NaBH4)/二氯化镍(NiCl2)等;
氨基乙酰化方法可以是醋酸酐(Ac2O)/甲醇,乙酰氯(AcCl)/三乙胺,醋酸酐/吡啶等。
4)反应模块D:单糖砌块1的4号位脱保护
当羟基保护基团为2-萘甲基,对甲氧苄基时,所用脱保护条件可以是2,3-二氯-5,6-二氰基-1,4-对苯醌(DDQ)/水,硝酸铈铵(CAN)/水等;
当羟基保护基团为叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基时,所用脱保护条件可以是四丁基氟化铵(TBAF),氢氟酸等。
5)反应模块E:单糖砌块2的2号位构型翻转
当L-奎诺糖2号位羟基的保护基为乙酰丙酰基时,脱保护的方法可以是醋酸肼/吡啶等;
当L-奎诺糖2号位羟基的保护基为芴甲氧羰基时,脱保护的方法可以是三乙胺等;
构型翻转可以通过三氟甲磺酸酯的Lattrell-Dax反应完成,三氟甲磺酸酯化反应所用试剂可以是三氟甲磺酸酐(Tf2O)/吡啶等,反应温度可以是-40至40℃之间;Lattrell-Dax反应所用试剂可以是亚硝酸钾(KNO2),四丁基亚硝酸铵(TBANO2),亚硝酸钠(NaNO2)等,反应温度可以是0至80℃之间;
构型翻转可以通过Swern氧化&羰基还原反应完成,Swern氧化所用试剂可以是二甲基亚砜(DMSO)/草酰氯((COCl)2)/三乙胺等,反应温度可以是-100℃至室温之间;羰基还原反应所用试剂可以是硼氢化钠(NaBH4),氢化铝锂(LiAlH4)等,反应温度可以是-20至40℃之间。
6)反应模块F:单糖砌块1的2、6号位、单糖砌块2的2号位苄基化
当D-葡萄糖胺6号位羟基保护基团为乙酰基,三氯乙酰基,苯甲酰基,二氯乙酰基,氯乙酰基,新戊酰基时,所用脱保护条件可以是甲醇钠/甲醇,氢氧化钠/甲醇,氢氧化钾/甲醇等;
当D-葡萄糖胺6号位羟基保护基团为烯丙氧羰酰基时,所用脱保护条件可以是二乙酸钯(Pd(OAc)2)/二乙胺等;
苄基化方法可以是溴化苄(BnBr)/钠氢(NaH),氯化苄(BnCl)/钠氢(NaH),碘化苄(BnI)/钠氢(NaH)等,反应温度可以是-20至40℃之间。
7)反应模块G:单糖砌块2的3号位脱保护
当羟基保护基团为2-萘甲基,对甲氧苄基时,所用脱保护条件可以是2,3-二氯-5,6-二氰基-1,4-对苯醌(DDQ)/水,硝酸铈铵(CAN)/水等;
当羟基保护基团为叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基时,所用脱保护条件可以是四丁基氟化铵(TBAF),氢氟酸等。
8)反应模块H:单糖砌块3的3号位脱保护
当羟基保护基团为2-萘甲基,甲氧苄基时,所用脱保护条件可以是2,3-二氯-5,6-二氰基-1,4-对苯醌(DDQ)/水,硝酸铈铵(CAN)/水等;
当羟基保护基团为叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基时,所用脱保护条件可以是四丁基氟化铵(TBAF),氢氟酸等;
当羟基保护基团为乙酰基,苯甲酰基,三氯乙酰基,氯乙酰基,乙酰丙酰基,二氯乙酰基,新戊酰基时,所用脱保护条件可以是甲醇钠/甲醇,氢氧化钾/甲醇,氢氧化钠/甲醇等;
当羟基保护基团为烯丙氧羰酰基时,所用脱保护条件可以是二乙酸钯(Pd(OAc)2)/二乙胺等。
9)反应模块I:单糖砌块4的2号位脱保护
当羟基保护基团为2-萘甲基,对甲氧苄基时,所用脱保护条件可以是2,3-二氯-5,6-二氰基-1,4-对苯醌(DDQ)/水,硝酸铈铵(CAN)/水等。
10)反应模块J:D-甘露糖4,6号位丙酮酸组装
4,6号位苯甲缩醛或萘甲缩醛保护基的脱去方法可以是40-90%醋酸水溶液方法,反应温度可以是40至80℃之间,也可以是三氟乙酸/水/有机溶剂方法,三氟乙酸的当量可以是50至500之间,水的当量可以是5-50之间,反应温度可以是-20至室温之间;
4,6号位丙酮酸组装方法可以是2,2-二(乙硫基)丙酸酯方法,2,2-二(乙硫基)丙酸酯可以选自2,2-二(乙硫基)丙酸甲酯,2,2-二(乙硫基)丙酸苄酯,2,2-二(乙硫基)丙酸萘甲酯,2,2-二(乙硫基)丙酸乙酯,2,2-二(乙硫基)丙酸丙酯,2,2-二(乙硫基)丙酸丁酯等,即羧基保护基PG15可以是甲基,苄基,萘甲基,乙基,丙基,丁基;活化剂可以选自三氟甲磺酸甲酯(TfOMe),二甲基甲硫基硫鎓三氟甲磺酸盐(DMTST),三氟甲磺酸(TfOH)/N-碘代丁二酰亚胺(NIS),三氟甲磺酸三甲基硅酯(TMSOTf)/N-碘代丁二酰亚胺(NIS),反应温度可以是-40℃至室温之间,用于除水的分子筛可以是分子筛或分子筛;
4,6号位丙酮酸组装方法可以是丙酮酸酯方法,丙酮酸酯可以选自丙酮酸甲酯,丙酮酸苄酯,丙酮酸萘甲酯,丙酮酸乙酯,丙酮酸丙酯,丙酮酸丁酯等,即羧基保护基PG15可以是甲基,苄基,萘甲基,乙基,丙基,丁基;活化剂可以是三氟化硼乙醚络合物(BF3·OEt2),反应温度可以是-40℃至室温之间,用于除水的分子筛可以是分子筛或分子筛。
11)反应模块K:全脱保护。
羧基和羟基的酯类保护基的脱保护方法可以是氢氧化钾/甲醇/水,氢氧化钠/乙醇/水,氢氧化锂/甲醇/水,氢氧化钠/甲醇/水,氢氧化钾/乙醇/水,氢氧化锂/乙醇/水等;
羟基的硅醚类保护基的脱保护方法可以是四丁基氟化铵,氢氟酸等;
羟基和氨基的碳醚类保护基的脱保护方法可以是催化氢化,通过催化条件下通入氢气进行反应,催化氢化所用催化剂可以是10%钯碳催化剂或氢氧化钯等,反应所用溶剂可以是水/甲醇/二氯甲烷/醋酸混合液,水/叔丁醇/二氯甲烷混合液,水/叔丁醇/四氢呋喃混合液等,反应温度可以是0至40℃之间。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,II-a的合成步骤包括:单糖砌块1经糖苷化反应A组装连接臂,得到化合物6,经反应模块C,D得到单糖受体7,与单糖砌块2经反应模块A得到二糖8,经反应模块E得到9,经反应模块F,G得到二糖受体11;单糖砌块3经反应模块H制得单糖受体12,与单糖砌块4经反应模块B制得二糖13;二糖受体11与二糖供体13经反应模块A制得四糖14,经反应模块J,K制得四糖II-a。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,II-b的合成步骤包括:单糖砌块2经反应模块A制得组装有连接臂的16,经反应模块E制得构型翻转产物17,经反应模块F,G制得受体19,进一步与单糖砌块3进行糖苷化反应,制得二糖20;经反应模块H制得二糖受体21,与单糖砌块4糖苷化反应得到三糖22,经反应模块I制得23,进一步与单糖砌块1糖苷化得到四糖24,经过反应模块C,D,J制得丙酮酸四糖26,经过全脱保护模块K制得四糖II-b。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,II-c的合成步骤包括:单糖砌块3与连接臂5经反应模块A制得27,经反应模块H脱保护基得受体28,与单糖砌块4糖苷化反应得二糖29,经反应模块I制得二糖受体30,与单糖砌块1糖苷化反应制得三糖31,经反应模块C,D制得三糖受体32,与单糖砌块2经反应模块A制得四糖33,经反应模块E得到构型翻转的34,经反应模块J,K制得四糖II-c。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,II-d的合成步骤包括:单糖砌块4与连接臂5通过糖苷化反应制备35,经反应模块I,得到受体36,与单糖砌块1进行糖苷化反应得到二糖37,经反应模块C,D得到二糖受体38,与单糖砌块2反应得到三糖39,经反应模块E得到三糖40,经过反应模块F,G得到三糖受体42,进一步与单糖砌块3进行糖苷化反应,得到四糖43,经反应模块J,K,制得四糖II-d。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,III-a的合成步骤包括:四糖43经反应模块H制得四糖受体44,与单糖砌块4经反应模块B制得五糖45,经反应模块J,K制得五糖III-a。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,III-b的合成步骤包括:四糖14经反应模块I制得四糖受体46,与单糖砌块1经反应模块A制得五糖47,经反应模块C,D制得五糖48,进一步经反应模块J,K制得III-b。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,III-c的合成步骤包括:四糖受体25与单糖砌块2经反应模块A制得五糖49,经反应模块E制得50,经反应模块J,K制得III-c。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,III-d的合成步骤包括:四糖34经反应模块F制得51,经反应模块G制得四糖受体52,与单糖砌块3经反应模块A制得五糖53,经反应模块J,K制得III-d。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,IV-a的合成步骤包括:五糖53经反应模块H制得五糖受体54,与单糖砌块4经反应模块B制得六糖55,经反应模块J,K制得六糖IV-a。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,IV-b的合成步骤包括:五糖45经反应模块I制得五糖受体56,与单糖砌块1经反应模块A制得六糖57,经反应模块C,D,J,K制得IV-b。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,IV-c的合成步骤包括:五糖48与单糖砌块2经反应模块A制得六糖58,经反应模块E制得59,经反应模块J,K制得IV-c。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,IV-d的合成步骤包括:五糖50经反应模块F制得60,经反应模块G制得五糖受体61,与单糖砌块3经反应模块A制得六糖62,经反应模块J,K制得IV-d。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,V-a的合成步骤包括:六糖62经反应模块H制得受体63,与单糖砌块4经反应模块B制得七糖64,经反应模块J,K制得V-a。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,V-b的合成步骤包括:六糖55经反应模块I制得受体65,与单糖砌块1经反应模块A制得66,经反应模块C,D,J,K制得V-b。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,V-c的合成步骤包括:六糖57经反应模块C,D制得受体67,与单糖砌块2经反应模块A制得七糖68,经反应模块E制得69,经反应模块J,K制得V-c。
在本发明的一种实施方式中,痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,如下反应步骤:
当n为1,V*-为H-时,V-d的合成步骤包括:六糖59经反应模块F制得70,经反应模块G制得受体71,与单糖砌块3经反应模块A制得七糖72,经反应模块J,K制得V-d。
本发明提供的痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,当糖砌块1作为糖基供体时,其2号位氨基可以由叠氮基进行保护,以利于1,2-顺式-α-糖苷键的生成。
本发明提供的痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,当糖砌块2(L-奎诺糖)作为糖基供体时,其2号位羟基可以由酯类保护基进行保护,以利于1,2-反式-β-糖苷键的生成。
本发明提供的痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,当L-鼠李糖的3号位羟基作为糖基化反应受体时,为了提高3号位羟基的亲核性,该L-鼠李糖的2号位羟基,D-氨基葡萄糖的2号位氨基和6号位羟基可以由苄基(Bn)保护。
本发明提供的痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,当糖砌块3(D-甘露糖胺)作为糖基供体时,其结构中2号位氨基可以由三氯乙酰基保护,以利于α-构型糖苷键的生成,且可通过催化氢化转化为乙酰氨基。
本发明提供的痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,当糖砌块4(D-甘露糖)作为糖基供体时,其2号位羟基可以由醚类保护基进行保护,4,6号位羟基由缩醛类保护基进行保护,采用反应模块B(预活化糖苷化反应)作为糖苷化方法,以利于1,2-顺式-β-糖苷键的生成。
本发明提供的痢疾志贺氏菌10血清型O-抗原寡糖可被制备为糖芯片,并应用于痢疾志贺氏菌抗血清中特异性抗体的检测。
本发明提供的痢疾志贺氏菌10血清型O-抗原寡糖的糖芯片,可应用于合成寡糖抗原表位的解析。
有益效果:
本发明提供的痢疾志贺氏菌10血清型O-抗原寡糖的化学合成方法,实现了1,2-顺式-α-葡萄糖苷键,1,2-顺式-β-鼠李糖苷键,1,2-反式-α-甘露糖苷键和1,2-顺式-β-甘露糖苷键的立体专一性合成。本发明提供的寡糖合成方法中,通过糖环羟基和氨基保护基的调整提高了糖基受体和糖基供体的反应活性,使各个糖苷键的合成产率均达到65%以上。本方法制备的寡糖组装有与天然糖链一致的(S)-4,6-O-丙酮酸,且还原端组装的氨基连接臂可实现寡糖与蛋白、芯片等载体的结合,为生物活性评价实验的开展奠定基础。同时,利用本方法也可以便利地制备无丙酮酸寡糖衍生物和(R)-4,6-O-丙酮酸寡糖衍生物,为开展痢疾志贺氏菌10血清型O-抗原寡糖的免疫活性研究及其构效关系探究提供了物质基础。本发明制备的合成寡糖已被制备为糖芯片,并成功应用于诊断痢疾志贺氏菌10血清型菌株的感染,以及合成寡糖抗原表位的解析。
附图说明
图1:痢疾志贺氏菌10血清型O-抗原四糖重复单元。
图2:通式I中Ux,Ux+1,Ux+2,Ux+3所示化合物。
图3:通式II-a,II-b,II-c和II-d所示化合物。
图4:通式III-a,III-b,III-c和III-d所示化合物。
图5:通式IV-a,IV-b,IV-c和IV-d所示化合物。
图6:通式V-a,V-b,V-c和V-d所示化合物。
图7:单糖砌块1,2,3,4和连接臂5所示化合物。
图8:化合物4*的合成反应式。
图9:化合物9*的合成反应式。
图10:化合物14*和15*的合成反应式。
图11:合成寡糖芯片的制备及其应用。
图12:四糖合成的对比例。
具体实施方式
实验中所用商品化试剂均未经处理直接使用。反应所用无水溶剂由MBraun MB-SPS 800型溶剂干燥系统制备。硅胶柱层析所用溶剂均为分析纯且经减压蒸馏后使用。薄层层析(TLC)所用硅胶板为60-F254硅胶制备的玻璃基或铝箔基硅胶板。薄层层析显色试剂为糖显色剂(0.1%(v/v)3-甲氧基苯酚,2.5%(v/v)硫酸乙醇溶液),或茚三酮显色剂(1.5%(w/v)茚三酮和3%(v/v)醋酸正丁醇溶液),或CAM显色剂(5%(w/v)钼酸铵,1%(w/v)硫酸铈(II)和10%(v/v)硫酸水溶液)。正相硅胶柱层析所用硅胶为200-300目硅胶。
分别对各反应步骤产率进行计算,产率计算方式为:(目标产物物质的量/原料物质的量)×100%。利用核磁图谱,红外图谱,旋光度,高分辨质谱对于产品进行结构鉴定,利用核磁图谱对于产品进行纯度分析。氢谱、碳谱以及二维核磁谱由Bruker Ascend 600兆核磁共振仪,或Bruker Ultrashield Plus 400兆核磁共振仪在25℃下测得。高分辨质谱由Agilent 6220电喷雾离子源-飞行时间质谱仪测得。红外图谱由Thermo FisherScientific Nicolet iS5红外仪测得。旋光度由Schmidt&Haensch UniPol L 1000全自动旋光仪在589nm下测得,测定浓度(c)单位为g/100mL。
实施例1:
N-苄基-N-苄氧羰基-3-氨基丙基2-乙酰氨基-3-O-苄基-4-O-(4-O-苄基-3-O-(2-萘)甲基-β-L-吡喃鼠李糖)-2-脱氧-α-D-吡喃葡萄糖(2*)的合成
反应式如图8所示;
化合物1*(宋武琼,江南大学硕士学位论文,2018)(15mg,0.014mmol)溶解于甲醇(0.5mL),加入甲醇钠(0.4mg,0.007mmol),在室温下搅拌4h。以Amberlite IR 120(H+)离子交换树脂中和,过滤和减压蒸馏后,粗品经硅胶柱层析纯化(石油醚:乙酸乙酯1:1v/v)制得2*(12mg,0.012mmol,86%)。[α]D 20=+37.0°(c=0.20,CHCl3);1H NMR(400MHz,CDCl3)δ=7.86-7.66(m,4H,Ar),7.51-7.28(m,17H,Ar),7.21(m,7H,Ar),6.82(d,J=9.4Hz,1H,2-NH),5.17(m,2H,ArCH2),4.88(d,J=10.8Hz,1H,ArCH2),4.81-4.51(m,7H,1-H,1’-H,ArCH2),4.46(d,J=10.4Hz,1H,ArCH2),4.37(d,J=15.4Hz,1H,ArCH2),4.27(br,1H,2-H),3.98-3.75(m,4H,2’-H,3-H,linker-CH2,6-CH2),3.68(m,4H,linker-CH2,4-H,5-H,6-CH2),3.45(t,J=9.1Hz,1H,4’-H),3.40-3.08(m,4H,3’-H,5’-H,linker-CH2),2.40(br,2H,6-OH,2’-OH),2.00(s,3H,CH3),1.79-1.64(m,2H,linker-CH2),1.32(d,J=6.0Hz,3H,6’-CH3);13CNMR(100MHz,CDCl3)δ=170.8,156.5,138.3,138.0,137.4,136.4,135.2,133.2,133.1,128.7,128.54,128.50,128.46,128.4,128.1,128.0,127.90,127.8,127.7,127.6,127.3,126.6,126.3,126.1,125.7,100.2(1-C),97.9(1’-C),81.7,81.6,79.2,77.2,76.4,75.6,75.5,72.0,71.6,70.9,68.4,67.5,63.5,62.2,53.0,49.6,42.7,27.2,23.2,17.6,14.1,11.0;HR-ESI-MS(m/z):calcd for C57H64N2NaO12 +(M+Na+):991.4351,found:991.4354.
实施例2:
N-苄基-N-苄氧羰基-3-氨基丙基2-N-乙酰基-2-N-苄基-3,6-二-O-苄基-4-O-(2,4-二-O-苄基-3-O-(2-萘)甲基-β-L-吡喃鼠李糖苷)-2-脱氧-α-D-吡喃葡萄糖(3*)的合成
反应式如图8所示;
在氮气保护下,化合物2*(20mg,0.021mmol)溶解于二甲基甲酰胺(0.5mL),在-10℃下,加入钠氢(60%分散于矿物油中)(4mg,0.100mmol),溴苄(10μL,0.084mmol),反应液在0℃下搅拌3h。加入水(0.3mL)淬灭反应后,以二氯甲烷萃取(20mL),经饱和碳酸氢钠水溶液(20mL)萃洗,硫酸钠除水后减压蒸馏除去溶剂。粗品经硅胶柱层析纯化(石油醚:乙酸乙酯3:1v/v)得到3*(17.4mg,0.014mmol,67%)。[α]D 20=+65.2°(c=0.25,CHCl3);IRνmax(film)3017,1693,1496,1453,1363,1215,1068,1027,750,697cm-1;1H NMR(400MHz,CDCl3)δ=7.87-7.64(m,4H,Ar),7.55-7.27(m,23H,Ar),7.25-6.82(m,15H,Ar),5.13(m,3H,ArCH2),4.92(dd,J=10.9,2.8Hz,1H,ArCH2),4.85(m,2H,ArCH2),4.77(d,J=17.5Hz,1H,ArCH2),4.65-4.55(m,3H,ArCH2),4.51(m,2H,ArCH2),4.47-4.37(m,2H,1’-H,ArCH2),4.31(m,3H,ArCH2),4.08(m,1H,2’-H),3.94(m,2H,1-H,3’-H),3.86(d,J=8.0Hz,1H,5-H),3.81(m,2H,4-H,6-CH2),3.69(m,2H,2-H,6-CH2),3.55(m,1H,4’-H),3.48(m,1H,Linker-CH2),3.19(m,2H,3-H,5’-H),3.06-2.76(m,2H,5’-H,Linker-CH2),2.74-2.41(m,1H,Linker-CH2),2.28(s,1.2H,CH3CO),1.81(s,1.8H,CH3CO),1.28(m,5H,6’-CH3,Linker-CH2);13C NMR(100MHz,CDCl3)δ=139.0,138.5,137.8,135.8,133.3,128.6,128.4,128.2,128.1,127.8,127.7,127.5,127.4,127.0,126.1,16.0,125.9,125.6,125.5,125.3,125.0,102.0,98.4,82.9,82.6,80.1,75.4,74.7,74.0,73.3,71.9,70.7,69.5,67.2,61.5,54.9,50.1,27.2,22.2,17.9;HR-ESI-MS(m/z):calcd for C78H82N2NaO12 +(M+Na+):1261.5760,found:1261.5762.
实施例3:
N-苄基-N-苄氧羰基-3-氨基丙基2-N-乙酰基-2-N-苄基-3,6-二-O-苄基-4-O-(2,4-二-O-苄基-β-L-吡喃鼠李糖)-2-脱氧-α-D-吡喃葡萄糖(4*)的合成
反应式如图8所示;
化合物3*(35mg,0.028mmol)溶解于二氯甲烷(6mL)和水(3mL),加入2,3-二氯-5,6-二氰基苯醌(DDQ)(10mg,0.045mmol),反应液在室温下搅拌5h。经5%硫代硫酸钠溶液萃洗(3×25mL),减压蒸馏除去溶剂,粗品经硅胶柱层析纯化(石油醚:乙酸乙酯3:1v/v)制得4*(24mg,0.022mmol,79%)。[α]D 20=+64.5°(c=0.50,CHCl3);IRνmax(film)3031,2905,1696,1654,1496,1452,1420,1363,1218,1068,1027,914,751,733,697,553cm-1;1H NMR(600MHz,CDCl3)δ=7.79-7.26(m,26H,Ar),7.24-6.72(m,9H,Ar),5.11(m,2H,ArCH2),4.96(m,2H,ArCH2),4.85(m,1H,ArCH2),4.79(s,1H,1-H),4.69(s,1H,1’-H),4.68-4.48(m,6H,ArCH2),4.47-4.33(m,1H,ArCH2),4.26(m,2H,ArCH2),4.13(m,1H,ArCH2),3.99(s,1H,2-H),3.94(m,2H,4-H,6-CH2),3.86-3.69(m,3H,3-H,5-H,6-CH2),3.64(m,2H,6-CH2,2'-H),3.53-3.36(m,1H,Linker-CH2),3.33(m,1H,3’-H),3.25-3.07(m,2H,4’-H,5’-H),3.07-2.74(m,2H,Linker-CH2),2.72-2.42(m,1H,Linker-CH2),2.32(s,1H,CH3CO),2.29(d,J=9.6Hz,1H,3-OH),1.84(s,2H,CH3CO),1.27(m,5H,Linker-CH2,6’-CH3);13C NMR(150MHz,CDCl3)δ=173.7,172.3,156.4,155.9,139.0,138.6,138.4,138.2,137.7,136.7,128.8,128.7,128.5,128.4,128.3,128.2,128.0,127.8,127.5,127.4,127.1,125.2,125.0,101.6(1’-C),98.4(1-C),82.1,78.7,78.2,75.3,75.1,75.0,74.1,73.5,73.3,71.4,70.7,70.4,69.7,68.9,67.2,65.9,65.5,61.5,54.7,50.0,48.3,46.3,44.1,43.3,29.7,27.2,22.6,22.2,17.9,17.9;HR-ESI-MS(m/z):calcd for C67H74N2NaO12 +(M+Na+):1121.5134,found:1121.5136.
实施例4:
烯丙基4,6-二-O-苄基-3-O-(2,3-二-O-苄基-4,6-O-苯甲缩醛-β-D-吡喃甘露糖)-2-脱氧-2-三氯乙酰氨基-α-D-吡喃甘露糖(7*)的合成
反应式如图9所示;
在氩气保护下,糖基供体5*(J.Carbohyd.Chem.1993,12(7),933)(860mg,1.75mmol)溶解于无水二氯甲烷(16mL),在-60℃下加入分子筛,1-(苯基亚硫酰基)哌啶(BSP)(440mg,2.10mmol),2,4,6-三叔丁基嘧啶(TTBP)(653mg,2.63mmol)和三氟甲磺酸酐(Tf2O)(383μL,2.28mmol)。反应液在-60℃下搅拌30min,加入1-辛烯(275μL,1.76mmol),继续搅拌15min。反应液降温至-78℃后,受体6*(宋武琼,江南大学硕士学位论文,2018)(1.1g,2.02mmol)的无水二氯甲烷溶液(9mL)加入反应液中,继续在-78℃下搅拌。当原料反应完,加入亚磷酸三乙酯(900μL,5.25mmol),反应液继续搅拌1h。反应液回温至室温后,加入二氯甲烷(30mL),经饱和碳酸氢钠水溶液(30mL)萃洗。以二氯甲烷萃取水相(2×20mL)后,所得有机相依次以饱和食盐水(30mL)萃洗,经硫酸钠除水,减压蒸馏除去溶剂。粗品经硅胶柱层析纯化(石油醚:乙酸乙酯8:1v/v)制得7*(1.28g,1.31mmol,75%,β-only)。[α]D 20=-22.2°(c=0.80,CHCl3);IRνmax(film)3410,3032,2864,1719,1512,1453,1369,1278,1214,1086,1028,992,918,819,790,736,697,618,600cm-1;1HNMR(400MHz,CDCl3)δ=7.52-7.47(m,2H,Ar),7.42-7.27(m,16H,Ar),7.25-7.15(m,7H,Ar),6.99(d,J=9.3Hz,1H,2-NH),5.89(dddd,J=17.0,10.3,6.6,5.3Hz,1H,CH=C),5.62(s,1H,PhCH),5.34-5.22(m,2H,C=CH2),5.02(d,J=10.4Hz,1H,PhCH2),4.93(d,J=1.6Hz,1H,1-H),4.84-4.81(m,2H,PhCH2),4.79(m,2H,1’-H,PhCH2),4.67(d,J=12.3Hz,1H,PhCH2),4.62(d,J=11.7Hz,1H,PhCH2),4.54-4.47(m,2H,2-H,4-H),4.45(d,J=11.7Hz,1H,PhCH2),4.33(dd,J=10.4,4.8Hz,1H,6’-CH2),4.24(d,J=10.4Hz,1H,PhCH2),4.21(d,J=9.5Hz,1H,4’-H),4.18-4.16(m,1H,OCHa),4.08(d,J=3.2Hz,1H,2’-H),4.02(ddt,J=12.6,6.6,1.2Hz,1H,OCHb),3.86-3.72(m,5H,3-H,5-H,3'-H,6'-CH2,6-CH2),3.70(m,1H,6-CH2),3.41(td,J=9.7,4.8Hz,1H,5’-H);13C NMR(100MHz,CDCl3)δ=162.1,139.1,138.6,138.3,137.9,137.7,132.9,128.8,128.5,128.4,128.3,128.2,128.1,128.0,127.9,127.8,127.7,127.52,127.46,127.4,127.2,126.1,118.9,101.5,98.8(1’-C),97.1(1-C),92.6,78.7,78.4,77.8,77.2,76.1,75.7,75.0,73.5,72.7,72.4,71.1,68.8,68.7,68.3,67.5,51.3;HR-ESI-MS(m/z):calcdfor C52H54Cl3NNaO11 +(M+Na+):996.2655,found:996.2653.
实施例5:
4,6-二-O-苄基-3-O-(2,3-二-O-苄基-4,6-O-苯甲缩醛-β-D-吡喃甘露糖)-2-脱氧-2-三氯乙酰氨基-D-吡喃甘露糖(8*)的合成
反应式如图9所示;
在氩气保护下,化合物7*(380mg,0.39mmol)溶解于乙酸乙酯(98mL),加入90%醋酸水溶液(10mL),醋酸钠(320mg,3.90mmol)和氯化钯(208mg,1.17mmol),反应液在室温下搅拌过夜。反应结束后,硅藻土过滤,滤液以乙酸乙酯稀释(100mL),进一步以水(2×100mL),饱和碳酸氢钠(2×100mL)萃洗,经硫酸钠处理除水,减压蒸馏除去溶剂。粗品经硅胶柱层析纯化(石油醚:乙酸乙酯4:1v/v)制得8*(346mg,0.37mmol,95%)。IRνmax(film)3396,2929,1719,1513,1454,1364,1269,1087,819,738,697,600cm-1;1H NMR(600MHz,CDCl3)δ=7.54-7.27(m,18H,Ar),7.25-7.16(m,7H,Ar),6.95(d,J=9.4Hz,1H,2-NH),5.61(s,1H,PhCH),5.23(t,J=2.5Hz,1H,1-H),5.02(d,J=10.6Hz,1H,PhCH2),4.85-4.76(m,4H,1’-H,PhCH2),4.67(d,J=12.1Hz,1H,PhCH2),4.60-4.55(m,1H,PhCH2),4.52(dd,J=9.1,4.0Hz,1H,3-H),4.49-4.43(m,2H,2-H,PhCH2),4.31(dd,J=10.5,5.0Hz,1H,6’-CH2),4.25(dd,J=10.5,7.3Hz,1H,PhCH2),4.19(t,J=9.6Hz,1H,4’-H),4.08(d,J=3.2Hz,1H,2’-H),4.04(m,1H,5-H),3.83-3.70(m,3H,3’-H,6’-CH2,6-CH2),3.70-3.61(m,2H,4-H,6-CH2),3.40(m,1H,5’-H),3.33(d,J=3.7Hz,1H,1-OH);13C NMR(150MHz,CDCl3)δ=162.2,139.0,138.5,138.2,137.8,137.6,128.9,128.5,128.4,128.3,128.2,128.13,128.05,127.9,127.8,127.7,127.54,127.48,127.3,127.2,126.1,101.5,98.8(1’-C),92.8(1-C),92.6,78.7,78.4,77.7,75.7,75.6,75.1,74.9,73.5,72.9,72.6,72.5,72.4,71.7,71.4,70.9,68.7,68.2,67.5,51.5,26.3;HR-ESI-MS(m/z):calcd for C49H50Cl3NNaO11 +(M+Na+):956.2342,found:956.2336.
实施例6:
4,6-二-O-苄基-3-O-(2,3-二-O-苄基-4,6-O-苯甲缩醛-β-D-吡喃甘露糖)-2-脱氧-2-三氯乙酰氨基-D-吡喃甘露糖苷1-(N-苯基)-2,2,2-三氟乙酰亚胺酯(9*)的合成
反应式如图9所示;
在氮气保护下,化合物8*(241mg,0.258mmol),碳酸钾(356mg,2.576mmol)和N-苯基三氟乙酰亚胺氯(385μL,2.578mmol)溶解于无水二氯甲烷(2.5mL),反应液在室温下搅拌过夜。过滤后,滤液经减压蒸馏除去溶剂,粗品经硅胶柱层析纯化(石油醚:乙酸乙酯10:1v/v)制得9*(214mg,0.193mmol,75%)。1H NMR(600MHz,CDCl3)δ=7.53-7.08(m,30H),6.82(d,J=7.8Hz,1H),5.61(s,1H),4.98(d,J=10.5Hz,1H),4.89-4.80(m,4H),4.79(d,J=12.3Hz,1H),4.66(d,J=12.3Hz,1H),4.61(d,J=11.8Hz,1H),4.49(d,J=11.8Hz,1H),4.30-4.24(m,2H),4.18(t,J=9.6Hz,1H),4.09-4.05(m,1H),3.83-3.70(m,5H),3.37(td,J=9.7,4.9Hz,1H);13C NMR(150MHz,CDCl3)δ=162.9,138.8,138.4,138.0,137.8,137.5,128.91,128.88,128.5,128.4,128.3,128.19,128.17,128.1,127.9 127.8,127.71,127.70,127.5,127.3,126.1,119.3,101.4,98.7,93.5,92.4,78.6,78.2,77.7,77.5,76.0,75.7,74.8,73.4,72.5,72.3,68.6,67.6,60.4,50.7,21.1,14.2.
实施例7:
N-苄基-N-苄氧羰基-3-氨基丙基2,3-二-O-苄基-4,6-O-苯甲缩醛-β-D-吡喃甘露糖-(1→3)-4,6-二-O-苄基-2-脱氧-2-三氯乙酰氨基-α-D-吡喃甘露糖-(1→3)-2,4-二-O-苄基-β-L-吡喃鼠李糖-(1→4)-2-N-乙酰基-2-N-苄基-3,6-二-O-苄基-2-脱氧-α-D-吡喃葡萄糖(10*)的合成
反应式如图10所示;
在氩气保护下,供体9*(130mg,0.117mmol)和受体4*(42mg,0.038mmol)溶解于无水二氯甲烷(6mL),加入分子筛,在室温下搅拌30min。降温至0℃后,加入TMSOTf(5μL,0.028mmol),反应液在室温下搅拌6h。加入吡啶(0.2mL)淬灭反应后,硅藻土过滤,滤液以饱和碳酸氢钠水溶液萃洗(2×10mL),减压蒸馏除去溶剂。粗品经硅胶柱层析纯化(石油醚:乙酸乙酯3:1v/v)制得10*(50mg,0.025mmol,66%,αonly)。[α]D 20=+32.2°(c=0.40,CHCl3);IRνmax(film)3030,2867,1698,1659,1516,1453,1363,1216,1088,1027,915,819,735,697,612,594cm-1;1H NMR(400MHz,CDCl3)δ=7.48-7.27(m,30H,Ar),7.25-6.94(m,30H,Ar),6.85(d,J=10.0Hz,1H,2”-NH),5.54(s,1H,PhCH),5.15-5.01(m,3H,PhCH2),4.90-4.71(m,9H,1-H,1”’-H,PhCH2),4.68(d,J=12.2Hz,1H,PhCH2),4.65-4.56(m,4H,1’-H,3”-H,PhCH2),4.56-4.38(m,5H,PhCH2),4.34(s,1H,2”-H),4.32-4.19(m,5H,1”-H,PhCH2),4.19-4.05(m,3H,5”-H,2”’-H,4”’-H),4.04-3.91(m,3H,2-H,4-H,6”’-CH2),3.90-3.72(m,5H,3-H,5-H,6-CH2,3’-H,3”’-H),3.71-3.52(m,4H,6-CH2,2’-H,6”-CH2,6”’-CH2),3.54-3.32(m,5H,4’-H,4”-H,6”-CH2,5”’-H,Linker-OCH2),3.16(ddd,J=8.6,6.1,2.4Hz,1H,5’-H),3.08-2.74(m,2H,Linker-NCH2),2.59(m,1H,Linker-OCH2),2.30(s,1H,CH3CO),1.82(s,2H,CH3CO),1.33(d,J=4.2Hz,3H,6’-CH3),1.29-1.19(m,2H,Linker-CH2);13C NMR(100MHz,CDCl3)δ=161.8,139.0,138.6,137.9,137.8,137.6,137.0,129.1,128.6,128.4,128.3,128.2,128.1,127.9,127.8,127.74,127.65,127.5,127.4,126.1,125.3,125.0,102.1(1’-C),101.4,98.7(1”’-C),98.3(1-C),92.7(1”-C),79.4,78.7,75.7,75.0,73.4,73.2,72.4,71.9,70.9,69.7,68.6,67.5,67.2,51.0,22.2,17.8;HR-ESI-MS(m/z):calcd forC116H122Cl3N3NaO22 +(M+Na+):2036.7478,found:2036.7450.
对比例1
N-苄基-N-苄氧羰基-3-氨基丙基2-O-苯甲酰基-3-O-苄基-4,6-O-苯甲缩醛-β-D-吡喃甘露糖-(1→3)-4,6-二-O-苄基-2-脱氧-2-三氯乙酰氨基-α-D-吡喃甘露糖-(1→3)-4-O-苄基-2-O-苯甲酰基-β-L-吡喃鼠李糖-(1→4)-2-乙酰氨基-3-O-苄基-6-O-苯甲酰基-2-脱氧-α-D-吡喃葡萄糖(22*)的合成
反应式如图12所示;
在氩气保护下,受体20*(宋武琼,江南大学硕士学位论文,2018)(5mg,4.8μmol)和供体21*(宋武琼,江南大学硕士学位论文,2018)(15mg,13.4μmol)溶解于无水二氯甲烷(0.5mL),加入分子筛,反应液在室温下搅拌30min。降温至0℃后,加入TMSOTf(0.3μL,1.7μmol),反应液在室温下搅拌4h。加入吡啶(2滴)淬灭反应,以硅藻土过滤,滤液以饱和碳酸氢钠水溶液萃洗(2×5mL),减压蒸馏除去溶剂。虽然质谱检测到目标产物22*的信号峰,但并未成功得到目标化合物,表明反应仅生成痕量的目标产物。
实施例8:
N-苄基-N-苄氧羰基-3-氨基丙基2,3-二-O-苄基-β-D-吡喃甘露糖-(1→3)-4,6-二-O-苄基-2-脱氧-2-三氯乙酰氨基-α-D-吡喃甘露糖-(1→3)-2,4-二-O-苄基-β-L-吡喃鼠李糖-(1→4)-2-N-乙酰基-2-N-苄基-3,6-二-O-苄基-2-脱氧-α-D-吡喃葡萄糖(11*)的合成
反应式如图10所示;
在0℃下,三氟乙酸(152μL,2.046mmol)和水(4μL,0.222mmol)加入10*(32mg,0.016mmol)的二氯甲烷溶液中(1.5mL),反应液在0℃下搅拌8h。经饱和碳酸氢钠水溶液(5mL)中和,以二氯甲烷萃取(4×5mL),经硫酸钠除水,减压蒸馏除去溶剂。粗品经硅胶柱层析纯化(石油醚:乙酸乙酯3:1v/v)制得11*(29mg,0.015mmol,94%)。[α]D 20=+84.7°(c=0.20,CHCl3);IRνmax(film)3030,1698,1496,1453,1362,1215,1071,1026,820,736,697,602,576cm-1;1H NMR(400MHz,CDCl3)δ=7.52-7.27(m,27H,Ar),7.25-6.86(m,28H,Ar),6.82(d,J=9.5Hz,1H,2”-NH),5.11(m,2H,PhCH2),4.99(dd,J=10.6,7.6Hz,1H,PhCH2),4.88(dd,J=11.8,3.3Hz,1H,PhCH2),4.85-4.70(m,5H,1-H,1”’-H,PhCH2),4.69-4.58(m,5H,1’-H,PhCH2),4.57-4.38(m,7H,3”-H,PhCH2),4.33(d,J=5.8Hz,1H,2”-H),4.31-4.15(m,4H,1”-H,PhCH2),4.10(s,1H,2”’-H),3.98(m,4H,2’-H,5”-H,3”’-H,6-CH2),3.93-3.83(m,2H,2-H,4-H),3.82-3.70(m,3H,3-H,5-H,3’-H),3.70-3.58(m,2H,6-CH2,6”-CH2),3.55(dt,J=9.4,4.2Hz,2H,6”’-CH2),3.46(m,5H,4’-H,4”-H,6”-CH2,5”’-H,Linker-OCH2),3.32(dd,J=9.2,4.4Hz,1H,4”’-H),3.16(dt,J=8.2,5.5Hz,1H,5’-H),2.93(m,2H,Linker-NCH2),2.59(m,1H,Linker-OCH2),2.31(s,1.4H,CH3CO),1.82(s,1.6H,CH3CO),1.31(d,J=6.1Hz,3H,6’-CH3),1.26(m,2H,Linker-CH2);13C NMR(150MHz,Chloroform-d)δ161.8,138.9,138.5,137.8,137.1,128.7,128.6,128.4,128.3,128.14,128.06,127.9,127.8,127.72,127.66,127.4,127.1,125.3,125.0,102.1(1’-C),98.4(1-C,1”’-C),92.9(1”-C),81.7,79.4,77.8,76.4,75.7,75.1,75.0,74.8,73.5,73.4,73.3,72.7,72.0,71.1,70.8,69.7,68.1,67.1,62.6,53.4,51.1,29.7,22.6,22.2,17.8;HR-ESI-MS(m/z):calcdfor C109H118Cl3N3NaO22 +(M+Na+):1948.7165,found:1948.7135.
实施例9:
N-苄基-N-苄氧羰基-3-氨基丙基2,3-二-O-苄基-4,6-O-[(S)-1-甲氧基羰基亚乙基]-β-D-吡喃甘露糖-(1→3)-4,6-二-O-苄基-2-脱氧-2-三氯乙酰氨基-α-D-吡喃甘露糖-(1→3)-2,4-二-O-苄基-β-L-吡喃鼠李糖-(1→4)-2-N-乙酰基-2-N-苄基-3,6-二-O-苄基-2-脱氧-α-D-吡喃葡萄糖(12*)和N-苄基-N-苄氧羰基-3-氨基丙基2,3-二-O-苄基-4,6-O-[(R)-1-甲氧基羰基亚乙基]-β-D-吡喃甘露糖-(1→3)-4,6-二-O-苄基-2-脱氧-2-三氯乙酰氨基-α-D-吡喃甘露糖-(1→3)-2,4-二-O-苄基-β-L-吡喃鼠李糖-(1→4)-2-N-乙酰基-2-N-苄基-3,6-二-O-苄基-2-脱氧-α-D-吡喃葡萄糖(13*)的合成反应式如图10所示;
在氩气保护下,化合物11*(25mg,13.0μmol)溶解于无水乙腈(0.5mL),加入分子筛,甲基2,2-二(乙硫基)丙酸酯(4μL,25.3μmol),在室温下搅拌30min。反应液降温至0℃,加入N-碘代丁二酰亚胺(NIS)(11mg,48.9μmol),搅拌10min后,进一步加入三氟甲烷磺酸(TfOH)(0.35μL,4.0μmol),继续在0℃下反应5h。反应液以饱和硫代硫酸钠水溶液(3mL)淬灭,硅藻土过滤后,滤液中加入二氯甲烷(20mL),以饱和碳酸氢钠水溶液(2×5mL),水(2×5mL)萃洗,硫酸钠除水后减压蒸馏除去溶剂。粗品经硅胶柱层析纯化(石油醚:乙酸乙酯3:1v/v)制得12*(15.2mg,7.6μmol,58%)和13*(7.8mg,3.9μmol,30%)。
化合物12*:[α]D 20=+32.0°(c=0.36,CHCl3);IRνmax(film)3900,3886,3624,3034,2883,2616,2597,1751,1701,1654,1518,1452,1365,1074,1030,914,823,752,697,618cm-1;1H NMR(600MHz,CDCl3)δ=7.44-7.27(m,25H,Ar),7.25-6.84(m,30H,Ar),6.80(d,J=9.5Hz,1H,2”-NH),5.11(m,2H,PhCH2),4.93(m,2H,PhCH2),4.83-4.67(m,8H,1-H,1”’-H,PhCH2),4.65-4.56(m,3H,1’-H,PhCH2),4.56-4.38(m,6H,3”-H,PhCH2),4.36-4.13(m,7H,1”-H,2”-H,PhCH2),4.09-3.90(m,6H,2-H,4-H,2’-H,5”-H,6”’-CH2,5”’-H),3.89-3.76(m,5H,3’-H,3”’-H,COOCH3),3.76-3.65(m,4H,3-H,2”’-H,4”’-H,6”’-CH2),3.64-3.51(m,3H,6-CH2,6”-CH2),3.43(m,4H,4’-H,4”-H,6”-CH2,Linker-OCH2),3.29(dd,J=9.7,5.4Hz,1H,5-H),3.15(m,1H,5’-H),3.05-2.76(m,2H,Linker-NCH2),2.73-2.47(m,1H,Linker-OCH2),2.30(s,1H,CH3CO),1.82(s,2H,CH3CO),1.53(s,3H,(S)Pyr-CH3),1.32(d,J=6.2Hz,3H,6’-CH3),1.29(m,2H,Linker-CH2);13C NMR(150MHz,CDCl3)δ=170.4,161.8,139.3,139.0,138.6,137.94,137.86,137.0,128.9,128.8,128.64,128.57,128.55,128.52,128.45,128.40,128.38,128.36,128.31,128.26,128.21,128.18,128.0,127.9,127.86,127.8,127.75,127.7,127.6,127.51,127.47,127.31,127.29,127.26,127.0,125.3,125.0,102.1(1’-C),99.2,98.5(1-C),98.3(1”’-C),92.8(1”-C),92.7,78.3,78.1,77.8,76.5,75.5,74.9,73.4,73.2,72.3,71.9,70.8,69.7,68.1,67.2,67.0,65.1,52.6,51.0,31.9,29.7,25.6(S-Pyr-CH3),22.6,22.2,17.8;HR-ESI-MS(m/z):calcd for C113H122Cl3N3NaO24 +(M+Na+):2032.7376,found:2032.7351.
化合物13*:[α]D 20=+39.1°(c=0.25,CHCl3);IRνmax(film)3819,3734,3648,3566,3164,3028,2874,2810,2386,2342,1748,1731,1647,1507,1454,1361,1066,913,823,750,699,593,580cm-1;1H NMR(600MHz,CDCl3)δ=7.45-7.27(m,28H,Ar),7.24-6.84(m,27H,Ar),6.80(d,J=9.5Hz,1H,2”-NH),5.12(m,2H,PhCH2),5.01(t,J=10.1Hz,1H,PhCH2),4.89-4.70(m,8H,1-H,1”’-H,PhCH2),4.70-4.45(m,9H,1’-H,3”-H,PhCH2),4.44-4.15(m,9H,1”-H,2”-H,4”’-H,PhCH2),4.15-3.91(m,6H,2-H,4-H,2’-H,5”-H,5”’-H,6”’-CH2),3.90-3.76(m,5H,COOCH3,3’-H,3”’-H),3.75-3.54(m,6H,6-CH2,6”-CH2,2”’-H,3-H,6”’-CH2),3.43(m,4H,4’-H,4”-H,6”-CH2,Linker-OCH2),3.37-3.30(m,1H,5-H),3.15(m,1H,5’-H),3.05-2.77(m,2H,Linker-NCH2),2.60(m,1H,Linker-OCH2),2.29(s,1H,CH3CO),1.82(s,2H,CH3CO),1.65(s,3H,(R)Pyr-CH3),1.33(m,5H,6’-CH3,Linker-CH2);13C NMR(150MHz,CDCl3)δ=169.1,161.8,139.1,138.8,129.0,128.64,128.55,128.4,128.2,128.0,127.9,127.8,127.7,127.6,127.44,127.41,125.0,102.1(1’-C),98.8(1-C),98.4(1”’-C),98.1,92.7(1”-C),79.4,75.7,74.9,73.4,73.2,72.4,71.9,71.5,70.9,68.1,67.9,67.2,62.9,52.8,51.1,29.7,22.2,18.2(R-Pyr-CH3),17.8;HR-ESI-MS(m/z):calcdfor C113H122Cl3N3NaO24 +(M+Na+):2032.7376,found:2032.7349.
实施例10:
3-氨基丙基4,6-O-[(S)-1-羧基亚乙基]-β-D-吡喃甘露糖-(1→3)-2-乙酰氨基-2-脱氧-α-D-吡喃甘露糖-(1→3)-β-L-吡喃鼠李糖-(1→4)-2-乙酰氨基-2-脱氧-α-D-吡喃葡萄糖(14*)的合成反应式如图10所示;
化合物12*(5mg,2.5μmol)溶解于甲醇(1mL),加入氢氧化钠水溶液(1M)(100μL),反应液在室温下搅拌12h。以Amberlite IR 120(H+)离子交换树脂中和后,过滤除去树脂。浓缩后,样品溶解于叔丁醇/水/二氯甲烷混合液(5:2:1v/v/v,5mL),以氮气吹扫后,加入10%Pd/C,再以氢气吹扫反应液5min,将反应液置于氢气环境下搅拌24h。反应液经硅藻土过滤后,浓缩滤液,以Sep-Pak cartridge C18(Macherey-Nagel,Düren,Germany)小柱纯化制得14*(2mg,2.3μmol,92%)。1H NMR(600MHz,D2O)δ=5.05(d,J=5.0Hz,1H,1”-H),4.87(s,1H,1-H),4.85(s,2H,1’-H,1”’-H),4.56(d,J=4.7Hz,1H,2”-H),4.28(m,1H,2’-H),4.24(dd,J=9.6,4.3Hz,1H,3”-H),4.04(dd,J=10.6,5.0Hz,1H,6”’-CH2),3.98(d,J=9.4Hz,1H,5”-H),3.92(m,1H,2-H),3.87(m,5H,Linker-OCH2,6-CH2,6”-CH2,3-H),3.80(m,2H,6”’-CH2,5-H),3.72(m,4H,4-H,4”-H,3”’-H,4”’-H),3.65(m,2H,3’-H,2”’-H),3.55(dt,J=11.4,6.1Hz,1H,Linker-OCH2),3.45(t,J=9.5Hz,1H,4’-H),3.43-3.32(m,2H,5’-H,5”’-H),3.11(t,J=7.6Hz,1H,Linker-NCH2),2.03(s,6H,CH3CO),1.98(m,2H,Linker-CH2),1.46(s,3H,S-Pyr-CH3),1.33(d,J=5.7Hz,3H,6’-CH3);13C NMR(150MHz,D2O)δ=174.5,106.4,100.8(1”’-C),97.2(1’-C),97.0(1-C),95.4(1”-C),77.5,76.9,75.3,73.8,72.2,71.2,70.8,70.6,66.8,65.1,64.8,64.3,60.8,60.2,53.7,49.4,37.3,26.8,24.8(S-Pyr-CH3),21.9,16.9;HR-ESI-MS(m/z):calcd for C34H58N3O22 +(M+H+):860.3506,found:860.3530.
实施例11:
3-氨基丙基4,6-O-[(R)-1-羧基亚乙基]-β-D-吡喃甘露糖-(1→3)-2-乙酰氨基-2-脱氧-α-D-吡喃甘露糖-(1→3)-β-L-吡喃鼠李糖-(1→4)-2-乙酰氨基-2-脱氧-α-D-吡喃葡萄糖(15*)的合成反应式如图10所示;
化合物13*(4.5mg,2.2μmol)溶解于甲醇(1mL),加入氢氧化钠水溶液(1M)(100μL),反应液在室温下搅拌12h。以Amberlite IR 120(H+)离子交换树脂中和,过滤除去树脂,浓缩滤液。样品溶解于叔丁醇/水/二氯甲烷混合液(5:2:1v/v/v,5mL),以氮气吹扫反应液,加入10%Pd/C,进一步以氢气吹扫反应液5min,将反应液置于氢气环境下搅拌24h。硅藻土过滤后,减压蒸馏除去溶剂,粗品经Sep-Pak cartridge C18(Macherey-Nagel,Düren,Germany)纯化制得15*(1.8mg,2.1μmol,95%)。1H NMR(600MHz,D2O)δ=5.07(d,J=3.8Hz,1H,1”-H),4.87(m,3H,1-H,1’-H,1”’-H),4.58(d,J=4.7Hz,1H,2”-H),4.27(m,2H,3”-H,2”’-H),4.07(m,1H,6”’-CH2),4.04-3.96(m,2H,5”-H,6”’-CH2),3.95-3.91(m,2H,2-H,2’-H),3.90-3.80(m,7H,3-H,5-H,4”-H,6-CH2,6”-CH2,Linker-OCH2),3.74(m,4H,4-H,3’-H,6”-CH2,4”’-H),3.66(m,1H,3”’-H),3.56(m,1H,Linker-OCH2),3.45(m,1H,5’-H),3.42(m,2H,4’-H,5”’-H),3.13(t,J=7.4Hz,2H,Linker-NCH2),2.06(s,3H,CH3CO),2.05(s,3H,CH3CO),1.99(m,2H,Linker-CH2),1.64(s,3H,R-Pyr-CH3),1.34(d,J=5.7Hz,3H,6’-CH3);13C NMR(150MHz,D2O)δ=174.6,100.8(1”’-C),97.5(1’-C),97.0(1-C),95.5(1”-C),85.0,77.5,77.0,76.8,76.4,75.3,72.3,72.1,71.7,71.1,70.4,67.5,66.9,65.2,65.1,64.94,64.85,61.9,60.8,60.6,60.3,53.8,49.5,37.3,26.9,25.1,22.0,21.9,16.9(R-Pyr-CH3);HR-ESI-MS(m/z):calcd for C34H58N3O22 +(M+H+):860.3506,found:860.3541.
实施例12:
合成寡糖芯片的制备及其生物医药应用
如图11所示;
合成寡糖溶解于偶联用磷酸缓冲液(50mM,pH 8.5),通过芯片点样仪RMA-Arrayer96(瀚辰光翼,中国)点样于“CodeLink”玻片(SurModics)。玻片在26℃,55%湿度下孵育过夜后,置于芯片淬灭缓冲液(50nM Na2HPO4,100nM乙醇胺)中,在50℃下处理1h。玻片经水清洗后,离心除去玻片上残留液体。在4℃下,以3%牛血清白蛋白(w/v)磷酸缓冲液溶液处理玻片进行封闭过夜,进一步以PBST(0.1%吐温的磷酸缓冲液)清洗一次,以磷酸缓冲液清洗两次,离心除去玻片上残留液体。兔来源的痢疾志贺氏菌抗血清(1-10型)(SSIDiagnostica,丹麦)以1%磷酸缓冲液-牛血清白蛋白(w/v)稀释,稀释比例为1:50,加至糖芯片上,每个样品至少重复四次。将芯片置于孵育室(ProPlate)中,4℃孵育过夜。将孵育完的糖芯片除去抗血清溶液,以PBST清洗三次。羊抗兔IgG Alexa FluorTM 532(Invitrogen)用作二抗,以1%磷酸缓冲液-牛血清白蛋白(w/v)稀释,稀释比例为1:400。将二抗稀释液加入糖芯片孔板中,置于孵育室,在37℃下孵育45min。以PBST清洗后,将隔板除去,以水清洗糖芯片10min,离心除去玻片上残留液体。以LuxScan 10K芯片扫描仪(北京博奥生物,中国)对糖芯片进行扫描,所得图像以GenePix Pro 7软件(Molecular Devices)进行处理。
(S)-4,6-O-丙酮酸四糖14*,(R)-4,6-O-丙酮酸四糖15*,无丙酮酸四糖16*,二糖17*,(S)-4,6-O-丙酮酸D-甘露糖18*,D-甘露糖19*以及不相关二糖Cb(α-D-Manp-(1→4)-β-D-Rhap)(Angew.Chem.Int.Ed.,2020,59(46),20529)制备的糖芯片,与痢疾志贺氏菌抗血清结合实验如图11所示,抗血清中IgG和IgM均可特异性识别(S)-4,6-O-丙酮酸四糖14*,无法识别(R)-4,6-O-丙酮酸四糖15*,无丙酮酸四糖16*,二糖17*和D-甘露糖19*,表明痢疾志贺氏菌10血清型O-抗原四糖重复单元是关键抗原表位。而(S)-4,6-O-丙酮酸D-甘露糖18*可被抗血清中IgG和IgM识别,表明(S)-4,6-O-丙酮酸基团是该多糖免疫活性的重要结构因子。上述结果表明,含有(S)-4,6-O-丙酮酸的痢疾志贺氏菌10血清型O-抗原寡糖可有效用于痢疾志贺氏菌诊断技术和疫苗的开发。
Claims (10)
1.一种痢疾志贺氏菌10血清型O-抗原的组装有连接臂的寡糖片段的合成方法,其特征在于,所述寡糖片段的结构如通式I所示:
V*-[Ux+3-Ux+2-Ux+1-Ux]n-V-O-L-NH2 通式I;
其中,x为1,2,3;n为1,2,3;-V-表示:化学键,-Ux+3-,-Ux+3-Ux+2-,或-Ux+3-Ux+2-Ux+1-;V*-表示:H-,H-Ux-,H-Ux+1-Ux-,或H-Ux+2-Ux+1-Ux-;L表示连接臂;Ux,Ux+1,Ux+2,Ux+3的结构如下所示:
所述合成方法是以四个单糖砌块1,2,3,4和连接臂5为原料,进行多步糖苷化反应,最后脱保护;
其中:
PG1,PG6,PG8,PG9,PG11为苄基;PG2,PG5为2-萘甲基,甲氧苄基,叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基;PG3为乙酰基,苯甲酰基,氯乙酰基,二氯乙酰基,三氯乙酰基,新戊酰基,烯丙氧羰酰基;PG4为乙酰丙酰基,芴甲氧羰基;PG7为2-萘甲基,甲氧苄基,叔丁基二甲基硅烷基,叔丁基二苯基硅烷基,三乙基硅烷基,乙酰基,苯甲酰基,乙酰丙酰基,氯乙酰基,二氯乙酰基,三氯乙酰基,新戊酰基,烯丙氧羰酰基;PG12为苯基,萘基;PG10为苄基,2-萘甲基,对甲氧苄基;PG13,PG14为苄基,苄氧羰基;LG1为乙硫基,对甲苯硫基,苯硫基,硒苯基,溴,氟,三氯乙酰亚氨酯,N-苯基三氟乙酰亚胺酯,二丁基磷酸酯;LG2为乙硫基,对甲苯硫基,苯硫基。
6.根据权利要求1所述的方法,其特征在于,包括11种反应模块:A:常规糖苷化反应;B:预活化糖苷化反应;C:叠氮基还原乙酰化;D:单糖砌块1的4号位脱保护;E:单糖砌块2的2号位构型翻转;F:单糖砌块1的2、6号位、单糖砌块2的2号位苄基化;G:单糖砌块2的3号位脱保护;H:单糖砌块3的3号位脱保护;I:单糖砌块4的2号位脱保护;J:单糖砌块4的4,6号位丙酮酸组装;K:全脱保护。
8.根据权利要求6所述的方法,其特征在于,反应模块E:单糖砌块2的2号位构型翻转的过程中,
当单糖砌块2的2号位羟基的保护基为乙酰丙酰基时,脱保护的方法是醋酸肼/吡啶;当单糖砌块2的2号位羟基的保护基为芴甲氧羰基时,脱保护的方法是三乙胺;
构型翻转通过三氟甲磺酸酯化反应、Lattrell-Dax反应完成;三氟甲磺酸酯化反应所用试剂可以是三氟甲磺酸酐/吡啶,反应温度是-40至40℃之间;Lattrell-Dax反应所用试剂可以是亚硝酸钾、四丁基亚硝酸铵、或者亚硝酸钠,反应温度是0至80℃之间;
或者,构型翻转通过Swern氧化和羰基还原反应完成;Swern氧化所用试剂是二甲基亚砜/草酰氯/三乙胺,反应温度是-100℃至室温之间;羰基还原反应所用试剂是硼氢化钠或者氢化铝锂,反应温度是-20至40℃之间。
9.根据权利要求6所述的方法,其特征在于,反应模块F:单糖砌块1的2、6号位、单糖砌块2的2号位苄基化的过程中,当D-葡萄糖胺6号位羟基保护基团为乙酰基,三氯乙酰基,苯甲酰基,二氯乙酰基,氯乙酰基,新戊酰基时,所用脱保护条件可以是甲醇钠/甲醇,氢氧化钠/甲醇,氢氧化钾/甲醇;当D-葡萄糖胺6号位羟基保护基团为烯丙氧羰酰基时,所用脱保护条件可以是二乙酸钯/二乙胺;苄基化试剂是溴化苄/钠氢,氯化苄/钠氢,或者碘化苄/钠氢,反应温度可以是-20至40℃之间。
10.根据权利要求6所述的方法,其特征在于,反应模块J:单糖砌块4的4,6号位丙酮酸组装的过程中,
4,6号位苯甲缩醛或萘甲缩醛保护基的脱去方法是40-90%醋酸水溶液方法,反应温度可以是40至80℃之间;或者是三氟乙酸/水/有机溶剂方法,三氟乙酸的当量可以是50至500之间,水的当量可以是5-50之间,反应温度可以是-20至室温之间;
4,6号位丙酮酸组装方法是2,2-二(乙硫基)丙酸酯方法;2,2-二(乙硫基)丙酸酯选自2,2-二(乙硫基)丙酸甲酯,2,2-二(乙硫基)丙酸苄酯,2,2-二(乙硫基)丙酸萘甲酯,2,2-二(乙硫基)丙酸乙酯,2,2-二(乙硫基)丙酸丙酯,2,2-二(乙硫基)丙酸丁酯;活化剂选自三氟甲磺酸甲酯,二甲基甲硫基硫鎓三氟甲磺酸盐,三氟甲磺酸/N-碘代丁二酰亚胺,三氟甲磺酸三甲基硅酯/N-碘代丁二酰亚胺,反应温度是-40℃至室温之间,加入分子筛或分子筛;
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202210933673.7A CN115232177A (zh) | 2022-08-04 | 2022-08-04 | 一种痢疾志贺氏菌10型o-抗原寡糖的化学合成方法 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202210933673.7A CN115232177A (zh) | 2022-08-04 | 2022-08-04 | 一种痢疾志贺氏菌10型o-抗原寡糖的化学合成方法 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN115232177A true CN115232177A (zh) | 2022-10-25 |
Family
ID=83679350
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202210933673.7A Pending CN115232177A (zh) | 2022-08-04 | 2022-08-04 | 一种痢疾志贺氏菌10型o-抗原寡糖的化学合成方法 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN115232177A (zh) |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2018053468A1 (en) * | 2016-09-19 | 2018-03-22 | The University Of Toledo | Monoclonal igm antibodies from entirely carbohydrate constructs |
CN108558961A (zh) * | 2018-01-29 | 2018-09-21 | 江南大学 | 类志贺邻单胞菌o51血清型o抗原寡糖化学合成方法 |
-
2022
- 2022-08-04 CN CN202210933673.7A patent/CN115232177A/zh active Pending
Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2018053468A1 (en) * | 2016-09-19 | 2018-03-22 | The University Of Toledo | Monoclonal igm antibodies from entirely carbohydrate constructs |
US20190345255A1 (en) * | 2016-09-19 | 2019-11-14 | The University Of Toledo | Monoclonal igm antibodies from entirely carbohydrate constructs |
CN108558961A (zh) * | 2018-01-29 | 2018-09-21 | 江南大学 | 类志贺邻单胞菌o51血清型o抗原寡糖化学合成方法 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Huang et al. | Stereodirecting effect of C5-carboxylate substituents on the glycosylation stereochemistry of 3-deoxy-d-manno-oct-2-ulosonic acid (Kdo) thioglycoside donors: Stereoselective synthesis of α-and β-Kdo glycosides | |
CN108558961B (zh) | 类志贺邻单胞菌o51血清型o抗原寡糖化学合成方法 | |
JPH10279589A (ja) | シアル酸グリコシド、抗原、免疫吸着剤およびそれらの調製方法 | |
Walvoort et al. | Stereoselective Synthesis of 2, 3-Diamino-2, 3-dideoxy-β-D-mannopyranosyl Uronates | |
Cattaneo et al. | Synthesis of Group B Streptococcus type III polysaccharide fragments for evaluation of their interactions with monoclonal antibodies | |
Someya et al. | One-pot synthesis of cyclic oligosaccharides by the polyglycosylation of monothioglycosides | |
Abronina et al. | A Practical Silicon-Free Strategy for Differentiation of Hydroxy Groups in Arabinofuranose Derivatives | |
Karki et al. | An expeditious synthesis of blood-group antigens, ABO histo-blood group type II antigens and xenoantigen oligosaccharides with amino type spacer− arms | |
CN109627270B (zh) | 一种绿脓假单胞菌o11血清型o抗原寡糖的化学合成方法 | |
El Sayed et al. | Challenges in the stereocontrolled syntheses of β-rhamnosides | |
CN115232177A (zh) | 一种痢疾志贺氏菌10型o-抗原寡糖的化学合成方法 | |
CN114874345B (zh) | 一种幽门螺旋杆菌核心脂多糖寡糖抗原糖链的化学合成方法 | |
Pozsgay et al. | Synthesis of di-to penta-saccharides related to the O-specific polysaccharide of Shigella dysenteriae type 1, and their nuclear magnetic resonance study | |
Zhou et al. | Convergent synthesis of branched β-glucan tridecasaccharides ready for conjugation | |
Zhao et al. | Stereoselective synthesis of a branched α-decaglucan | |
CN114085255B (zh) | 一种苏黎世克罗诺杆菌5型脂多糖o-抗原寡糖片段及其制备方法与应用 | |
Boutet et al. | Synthesis of branched tri-to pentasaccharides representative of fragments of Shigella flexneri serotypes 3a and/or X O-antigens | |
Maity et al. | Convergent synthesis of the tetrasaccharide repeating unit related to the O-antigenic polysaccharide of Escherichia coli 78 | |
Liang et al. | One-pot synthesis of branched oligosaccharides by use of galacto-and mannopyranosyl thioglycoside diols as key glycosylating agents | |
Auzanneau et al. | Application of thioglycoside chemistry to the synthesis of trisaccharides and deoxy-trisaccharides related to the Shigella flexneri Y polysaccharide | |
US10851130B2 (en) | Chemical synthesis method of Plesiomonas shigelloides serotype O51 O-antigen oligosaccharide | |
Meng et al. | Stereoselective Synthesis of β-d-Manno-heptopyranoside via Cs2CO3-Mediated Anomeric O-Alkylation: Synthesis of a Tetrasaccharide Repeat Unit of Bacillus thermoaerophilus Surface-Layer Glycoprotein | |
Chen et al. | A concise and practical synthesis of antigenic globotriose, α-d-Gal-(1→ 4)-β-d-Gal-(1→ 4)-β-d-Glc | |
Ferrières et al. | A new approach to a disaccharidic hapten containing a galactofuranosyl entity | |
Pozsgay et al. | Synthesis of hexa-to tridecasaccharides related to Shigella dysenteriae type 1 for incorporation in experimental vaccines |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination |