CN114276222A - Diaryl benzyl alcohol compound as polypeptide liquid phase synthesis carrier and preparation method and application thereof - Google Patents
Diaryl benzyl alcohol compound as polypeptide liquid phase synthesis carrier and preparation method and application thereof Download PDFInfo
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- CN114276222A CN114276222A CN202111676811.XA CN202111676811A CN114276222A CN 114276222 A CN114276222 A CN 114276222A CN 202111676811 A CN202111676811 A CN 202111676811A CN 114276222 A CN114276222 A CN 114276222A
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- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 35
- 230000015572 biosynthetic process Effects 0.000 title claims abstract description 30
- 238000003786 synthesis reaction Methods 0.000 title claims abstract description 30
- -1 Diaryl benzyl alcohol compound Chemical class 0.000 title claims abstract description 24
- WVDDGKGOMKODPV-UHFFFAOYSA-N hydroxymethyl benzene Natural products OCC1=CC=CC=C1 WVDDGKGOMKODPV-UHFFFAOYSA-N 0.000 title claims abstract description 24
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 24
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 24
- 239000007791 liquid phase Substances 0.000 title claims abstract description 16
- 235000019445 benzyl alcohol Nutrition 0.000 title claims abstract description 12
- 238000002360 preparation method Methods 0.000 title claims abstract description 10
- 150000001875 compounds Chemical class 0.000 claims abstract description 56
- 150000001413 amino acids Chemical class 0.000 claims abstract description 35
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 19
- 238000000034 method Methods 0.000 claims abstract description 16
- 238000006467 substitution reaction Methods 0.000 claims abstract description 9
- 238000006243 chemical reaction Methods 0.000 claims description 111
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 108
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 102
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 claims description 34
- 239000000203 mixture Substances 0.000 claims description 33
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 claims description 30
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 21
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 claims description 18
- 239000002904 solvent Substances 0.000 claims description 17
- 238000007792 addition Methods 0.000 claims description 14
- 239000000126 substance Substances 0.000 claims description 13
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 claims description 13
- 238000010511 deprotection reaction Methods 0.000 claims description 12
- 239000007800 oxidant agent Substances 0.000 claims description 12
- 230000001590 oxidative effect Effects 0.000 claims description 12
- 238000005886 esterification reaction Methods 0.000 claims description 11
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Chemical compound [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 claims description 9
- WETWJCDKMRHUPV-UHFFFAOYSA-N acetyl chloride Chemical compound CC(Cl)=O WETWJCDKMRHUPV-UHFFFAOYSA-N 0.000 claims description 7
- 239000012346 acetyl chloride Substances 0.000 claims description 7
- 239000002253 acid Substances 0.000 claims description 7
- 229940125782 compound 2 Drugs 0.000 claims description 7
- 229940126214 compound 3 Drugs 0.000 claims description 7
- 229940125898 compound 5 Drugs 0.000 claims description 7
- FBPINGSGHKXIQA-UHFFFAOYSA-N 2-amino-3-(2-carboxyethylsulfanyl)propanoic acid Chemical compound OC(=O)C(N)CSCCC(O)=O FBPINGSGHKXIQA-UHFFFAOYSA-N 0.000 claims description 6
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 claims description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 6
- 239000011230 binding agent Substances 0.000 claims description 6
- 239000003638 chemical reducing agent Substances 0.000 claims description 6
- 229910000027 potassium carbonate Inorganic materials 0.000 claims description 6
- 239000002994 raw material Substances 0.000 claims description 6
- 230000032050 esterification Effects 0.000 claims description 5
- 238000006722 reduction reaction Methods 0.000 claims description 5
- JWUJQDFVADABEY-UHFFFAOYSA-N 2-methyltetrahydrofuran Chemical compound CC1CCCO1 JWUJQDFVADABEY-UHFFFAOYSA-N 0.000 claims description 4
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 claims description 4
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims description 4
- 229910052736 halogen Inorganic materials 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
- 238000007254 oxidation reaction Methods 0.000 claims description 4
- FYSNRJHAOHDILO-UHFFFAOYSA-N thionyl chloride Chemical compound ClS(Cl)=O FYSNRJHAOHDILO-UHFFFAOYSA-N 0.000 claims description 4
- 239000012051 hydrophobic carrier Substances 0.000 claims description 3
- 238000003747 Grignard reaction Methods 0.000 claims description 2
- 239000007818 Grignard reagent Substances 0.000 claims description 2
- 150000001335 aliphatic alkanes Chemical class 0.000 claims description 2
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 claims description 2
- 229910052782 aluminium Inorganic materials 0.000 claims description 2
- 150000004795 grignard reagents Chemical class 0.000 claims description 2
- 125000005843 halogen group Chemical group 0.000 claims description 2
- 150000002367 halogens Chemical class 0.000 claims description 2
- 229910052744 lithium Inorganic materials 0.000 claims description 2
- 239000012280 lithium aluminium hydride Substances 0.000 claims description 2
- 229910001623 magnesium bromide Inorganic materials 0.000 claims description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L magnesium chloride Substances [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 claims description 2
- 229910001629 magnesium chloride Inorganic materials 0.000 claims description 2
- GDOPTJXRTPNYNR-UHFFFAOYSA-N methyl-cyclopentane Natural products CC1CCCC1 GDOPTJXRTPNYNR-UHFFFAOYSA-N 0.000 claims description 2
- 230000003647 oxidation Effects 0.000 claims description 2
- 229910000029 sodium carbonate Inorganic materials 0.000 claims description 2
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 claims description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N sulfuric acid Substances OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 2
- 150000001298 alcohols Chemical class 0.000 claims 3
- 238000006482 condensation reaction Methods 0.000 claims 1
- 230000007062 hydrolysis Effects 0.000 claims 1
- 238000006460 hydrolysis reaction Methods 0.000 claims 1
- 238000004519 manufacturing process Methods 0.000 claims 1
- 238000001556 precipitation Methods 0.000 claims 1
- 230000008878 coupling Effects 0.000 abstract description 28
- 238000010168 coupling process Methods 0.000 abstract description 28
- 238000005859 coupling reaction Methods 0.000 abstract description 28
- 239000006166 lysate Substances 0.000 abstract description 6
- 238000001308 synthesis method Methods 0.000 abstract description 6
- 125000006239 protecting group Chemical group 0.000 abstract description 3
- 125000000217 alkyl group Chemical group 0.000 abstract 1
- 238000009776 industrial production Methods 0.000 abstract 1
- 238000003860 storage Methods 0.000 abstract 1
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 98
- 238000001914 filtration Methods 0.000 description 68
- 239000007787 solid Substances 0.000 description 63
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 54
- 229910052757 nitrogen Inorganic materials 0.000 description 49
- 238000001035 drying Methods 0.000 description 48
- 239000000047 product Substances 0.000 description 42
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 42
- 238000001816 cooling Methods 0.000 description 33
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 31
- IMNFDUFMRHMDMM-UHFFFAOYSA-N N-Heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 description 27
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 26
- 239000000243 solution Substances 0.000 description 22
- AYMLQYFMYHISQO-QMMMGPOBSA-N (2s)-3-(1h-imidazol-3-ium-5-yl)-2-[(2-methylpropan-2-yl)oxycarbonylamino]propanoate Chemical compound CC(C)(C)OC(=O)N[C@H](C(O)=O)CC1=CN=CN1 AYMLQYFMYHISQO-QMMMGPOBSA-N 0.000 description 21
- YQZVQKYXWPIKIX-UHFFFAOYSA-N 2-[2-[2-[[2-[2-(2-aminoethoxy)ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid Chemical compound NCCOCCOCC(=O)NCCOCCOCC(O)=O YQZVQKYXWPIKIX-UHFFFAOYSA-N 0.000 description 21
- 239000012452 mother liquor Substances 0.000 description 21
- VHYFNPMBLIVWCW-UHFFFAOYSA-N 4-Dimethylaminopyridine Chemical compound CN(C)C1=CC=NC=C1 VHYFNPMBLIVWCW-UHFFFAOYSA-N 0.000 description 18
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 18
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 18
- KDLHZDBZIXYQEI-UHFFFAOYSA-N Palladium Chemical compound [Pd] KDLHZDBZIXYQEI-UHFFFAOYSA-N 0.000 description 18
- 238000005984 hydrogenation reaction Methods 0.000 description 18
- 238000004321 preservation Methods 0.000 description 17
- 238000010438 heat treatment Methods 0.000 description 16
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 15
- 239000007864 aqueous solution Substances 0.000 description 15
- 238000010926 purge Methods 0.000 description 15
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical class O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 15
- NPZTUJOABDZTLV-UHFFFAOYSA-N hydroxybenzotriazole Substances O=C1C=CC=C2NNN=C12 NPZTUJOABDZTLV-UHFFFAOYSA-N 0.000 description 13
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 13
- 235000017557 sodium bicarbonate Nutrition 0.000 description 13
- 239000006104 solid solution Substances 0.000 description 12
- 238000003756 stirring Methods 0.000 description 11
- NQRYJNQNLNOLGT-UHFFFAOYSA-N Piperidine Chemical compound C1CCNCC1 NQRYJNQNLNOLGT-UHFFFAOYSA-N 0.000 description 10
- 239000012071 phase Substances 0.000 description 10
- HNICLNKVURBTKV-NDEPHWFRSA-N (2s)-5-[[amino-[(2,2,4,6,7-pentamethyl-3h-1-benzofuran-5-yl)sulfonylamino]methylidene]amino]-2-(9h-fluoren-9-ylmethoxycarbonylamino)pentanoic acid Chemical compound C12=CC=CC=C2C2=CC=CC=C2C1COC(=O)N[C@H](C(O)=O)CCCN=C(N)NS(=O)(=O)C1=C(C)C(C)=C2OC(C)(C)CC2=C1C HNICLNKVURBTKV-NDEPHWFRSA-N 0.000 description 9
- 229960000549 4-dimethylaminophenol Drugs 0.000 description 9
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 9
- 239000012267 brine Substances 0.000 description 9
- 239000003795 chemical substances by application Substances 0.000 description 9
- 229910052739 hydrogen Inorganic materials 0.000 description 9
- 239000001257 hydrogen Substances 0.000 description 9
- 239000012074 organic phase Substances 0.000 description 9
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 7
- 238000005406 washing Methods 0.000 description 7
- NDKDFTQNXLHCGO-UHFFFAOYSA-N 2-(9h-fluoren-9-ylmethoxycarbonylamino)acetic acid Chemical compound C1=CC=C2C(COC(=O)NCC(=O)O)C3=CC=CC=C3C2=C1 NDKDFTQNXLHCGO-UHFFFAOYSA-N 0.000 description 6
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical class [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 6
- 238000005336 cracking Methods 0.000 description 6
- 239000012043 crude product Substances 0.000 description 6
- 239000012634 fragment Substances 0.000 description 6
- 238000004128 high performance liquid chromatography Methods 0.000 description 6
- 238000010791 quenching Methods 0.000 description 6
- KKWVSPRXEUBOHT-UHFFFAOYSA-N CC1=CC=CC=C1.[AlH3] Chemical compound CC1=CC=CC=C1.[AlH3] KKWVSPRXEUBOHT-UHFFFAOYSA-N 0.000 description 5
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 5
- 239000008346 aqueous phase Substances 0.000 description 5
- 238000010532 solid phase synthesis reaction Methods 0.000 description 5
- 125000003088 (fluoren-9-ylmethoxy)carbonyl group Chemical group 0.000 description 4
- WSULSMOGMLRGKU-UHFFFAOYSA-N 1-bromooctadecane Chemical compound CCCCCCCCCCCCCCCCCCBr WSULSMOGMLRGKU-UHFFFAOYSA-N 0.000 description 4
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 4
- REITVGIIZHFVGU-IBGZPJMESA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-3-[(2-methylpropan-2-yl)oxy]propanoic acid Chemical compound C1=CC=C2C(COC(=O)N[C@@H](COC(C)(C)C)C(O)=O)C3=CC=CC=C3C2=C1 REITVGIIZHFVGU-IBGZPJMESA-N 0.000 description 3
- UGNIYGNGCNXHTR-SFHVURJKSA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-3-methylbutanoic acid Chemical compound C1=CC=C2C(COC(=O)N[C@@H](C(C)C)C(O)=O)C3=CC=CC=C3C2=C1 UGNIYGNGCNXHTR-SFHVURJKSA-N 0.000 description 3
- WDGICUODAOGOMO-DHUJRADRSA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-5-oxo-5-(tritylamino)pentanoic acid Chemical compound C([C@@H](C(=O)O)NC(=O)OCC1C2=CC=CC=C2C2=CC=CC=C21)CC(=O)NC(C=1C=CC=CC=1)(C=1C=CC=CC=1)C1=CC=CC=C1 WDGICUODAOGOMO-DHUJRADRSA-N 0.000 description 3
- OYXZPXVCRAAKCM-SANMLTNESA-N (2s)-2-[(2-methylpropan-2-yl)oxycarbonylamino]-3-(1-tritylimidazol-4-yl)propanoic acid Chemical compound C1=NC(C[C@H](NC(=O)OC(C)(C)C)C(O)=O)=CN1C(C=1C=CC=CC=1)(C=1C=CC=CC=1)C1=CC=CC=C1 OYXZPXVCRAAKCM-SANMLTNESA-N 0.000 description 3
- BRRSNXCXLSVPFC-UHFFFAOYSA-N 2,3,4-Trihydroxybenzoic acid Chemical compound OC(=O)C1=CC=C(O)C(O)=C1O BRRSNXCXLSVPFC-UHFFFAOYSA-N 0.000 description 3
- ZAFNJMIOTHYJRJ-UHFFFAOYSA-N Diisopropyl ether Chemical compound CC(C)OC(C)C ZAFNJMIOTHYJRJ-UHFFFAOYSA-N 0.000 description 3
- 238000004140 cleaning Methods 0.000 description 3
- 239000011248 coating agent Substances 0.000 description 3
- 238000000576 coating method Methods 0.000 description 3
- 230000007547 defect Effects 0.000 description 3
- 239000012065 filter cake Substances 0.000 description 3
- 239000000706 filtrate Substances 0.000 description 3
- IWCVDCOJSPWGRW-UHFFFAOYSA-M magnesium;benzene;chloride Chemical compound [Mg+2].[Cl-].C1=CC=[C-]C=C1 IWCVDCOJSPWGRW-UHFFFAOYSA-M 0.000 description 3
- 238000010907 mechanical stirring Methods 0.000 description 3
- 239000003208 petroleum Substances 0.000 description 3
- 239000013557 residual solvent Substances 0.000 description 3
- DTHNMHAUYICORS-KTKZVXAJSA-N Glucagon-like peptide 1 Chemical compound C([C@@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCCN)C(=O)NCC(=O)N[C@@H](CCCNC(N)=N)C(N)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@H](CCC(N)=O)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@@H](NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC=1N=CNC=1)[C@@H](C)O)[C@@H](C)O)C(C)C)C1=CC=CC=C1 DTHNMHAUYICORS-KTKZVXAJSA-N 0.000 description 2
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 239000012299 nitrogen atmosphere Substances 0.000 description 2
- 230000001376 precipitating effect Effects 0.000 description 2
- 238000007363 ring formation reaction Methods 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- 102400000322 Glucagon-like peptide 1 Human genes 0.000 description 1
- 101800000224 Glucagon-like peptide 1 Proteins 0.000 description 1
- 108010019598 Liraglutide Proteins 0.000 description 1
- YSDQQAXHVYUZIW-QCIJIYAXSA-N Liraglutide Chemical compound C([C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)NCC(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCNC(=O)CC[C@H](NC(=O)CCCCCCCCCCCCCCC)C(O)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(O)=O)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@@H](NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC=1NC=NC=1)[C@@H](C)O)[C@@H](C)O)C(C)C)C1=CC=C(O)C=C1 YSDQQAXHVYUZIW-QCIJIYAXSA-N 0.000 description 1
- DLSWIYLPEUIQAV-UHFFFAOYSA-N Semaglutide Chemical compound CCC(C)C(NC(=O)C(Cc1ccccc1)NC(=O)C(CCC(O)=O)NC(=O)C(CCCCNC(=O)COCCOCCNC(=O)COCCOCCNC(=O)CCC(NC(=O)CCCCCCCCCCCCCCCCC(O)=O)C(O)=O)NC(=O)C(C)NC(=O)C(C)NC(=O)C(CCC(N)=O)NC(=O)CNC(=O)C(CCC(O)=O)NC(=O)C(CC(C)C)NC(=O)C(Cc1ccc(O)cc1)NC(=O)C(CO)NC(=O)C(CO)NC(=O)C(NC(=O)C(CC(O)=O)NC(=O)C(CO)NC(=O)C(NC(=O)C(Cc1ccccc1)NC(=O)C(NC(=O)CNC(=O)C(CCC(O)=O)NC(=O)C(C)(C)NC(=O)C(N)Cc1cnc[nH]1)C(C)O)C(C)O)C(C)C)C(=O)NC(C)C(=O)NC(Cc1c[nH]c2ccccc12)C(=O)NC(CC(C)C)C(=O)NC(C(C)C)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CCCNC(N)=N)C(=O)NCC(O)=O DLSWIYLPEUIQAV-UHFFFAOYSA-N 0.000 description 1
- UYNAFHNUGOEVFE-UHFFFAOYSA-M [Cl-].C1CCOC1.[Mg+]C1=CC=CC=C1 Chemical compound [Cl-].C1CCOC1.[Mg+]C1=CC=CC=C1 UYNAFHNUGOEVFE-UHFFFAOYSA-M 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 230000001808 coupling effect Effects 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 229960002701 liraglutide Drugs 0.000 description 1
- 125000000956 methoxy group Chemical group [H]C([H])([H])O* 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000000425 proton nuclear magnetic resonance spectrum Methods 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- DLSWIYLPEUIQAV-CCUURXOWSA-N semaglutide Chemical compound C([C@@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCCNC(=O)COCCOCCNC(=O)COCCOCCNC(=O)CC[C@@H](NC(=O)CCCCCCCCCCCCCCCCC(O)=O)C(O)=O)NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@H](CCC(N)=O)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@@H](NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)C(C)(C)NC(=O)[C@@H](N)CC=1NC=NC=1)[C@@H](C)O)[C@@H](C)O)C(C)C)C1=CC=CC=C1 DLSWIYLPEUIQAV-CCUURXOWSA-N 0.000 description 1
- 239000007790 solid phase Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- 208000001072 type 2 diabetes mellitus Diseases 0.000 description 1
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- 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
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
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- Peptides Or Proteins (AREA)
Abstract
The invention provides a diaryl benzyl alcohol compound as a polypeptide liquid phase synthesis carrier, a preparation method and application thereof, which comprises a synthesis method of the compound, a method for constructing polypeptide chains by sequentially coupling amino acids as an initial template reagent, and a method for removing the template under mild conditions without removing other protecting groups on the peptide chains; a method for removing the template and protecting groups in a lysate. The compound containing long-chain alkyl substitution developed by the invention has the characteristics of simple synthesis method, convenient operation, low cost, easy storage and easy removal, is very suitable to be used as an initial template for liquid phase synthesis of polypeptide chains, and is expected to realize large-scale industrial production.
Description
Technical Field
The invention relates to the field of polypeptide synthesis, in particular to diaryl benzyl alcohol compounds, a preparation method thereof and application thereof in polypeptide liquid phase synthesis.
Background
The somaglutide is a new generation of GLP-1 (glucagon-like peptide-1) analogue developed by NovoNordisk, Denmark, and is a long-acting dosage form developed based on the basic structure of liraglutide, which has better effect in treating type 2 diabetes.
Structural information for somaglutide is as follows: chemical name: the structural formula of the somaglutide Sermaglutide is shown as a formula II:
the molecular formula is as follows: c187H291N45O59(ii) a Molecular weight: 4113.58, respectively; CAS accession number: 910463-68-2.
At present, the Fmoc strategy is mostly adopted for solid-phase synthesis of the somaglutide, and the solid-phase synthesis has certain advantages in the aspect of post-treatment of reaction, but the defects are obvious: 1. 2-5 times or even higher of the Fmoc-protected amino acid charge is required. 2. After coupling an amino acid, a large amount of solvent DMF is needed for washing (generally 4-6 times of washing). 3. The reaction is heterogeneous. For solid phase cyclization, coupling of hydrophobic amino acid and the like, the coupling effect is poor due to large steric hindrance, and secondary or even tertiary feeding is required. 4. The volume of the resin is large, the dosage of the solvent is large, so that the reaction kettle efficiency is low, and the batch yield is low.
Therefore, the liquid phase synthesis method of the somaglutide, which is simple and convenient to operate and high in yield, has important practical significance.
Furthermore, in most of the solid phase synthesis strategies reported so far, the last step of removing the solid phase synthesis template substance requires the use of stronger acids, such as: trifluoroacetic acid at higher concentrations. The template is removed simultaneously with the removal of the protecting groups on the polypeptide chain. The synthesis of fully protected polypeptide chains is relatively rare. Therefore, the method for synthesizing the fully-protected polypeptide chain, which is simple and convenient to operate and high in yield, has important practical significance.
Disclosure of Invention
The invention aims to solve the defects of the existing Fmoc strategy that the method for solid-phase synthesis of polypeptide has more defects, and provides a diaryl benzyl alcohol compound used as a polypeptide liquid-phase synthesis carrier, which is used as the starting end of the carrier to synthesize polypeptide chain, is beneficial to the coupling and cyclization of difficult amino acids, and has convenient deprotection and high yield.
The second purpose of the invention is to provide the preparation method of the diaryl benzyl alcohol compound, which has the advantages of simple and easily obtained raw materials, mild preparation conditions, simple steps, easy realization and low cost.
The third purpose of the invention is to provide the application of the diaryl benzyl alcohol compound, wherein the diaryl benzyl alcohol compound is used as a hydrophobic carrier to synthesize fully protected polypeptide or fully deprotected polypeptide in a liquid phase.
In order to achieve the purpose, the invention adopts the following technical scheme:
the invention provides a compound, which has a structure shown in a formula I:
r is selected from C12-C22 fatty chain, X is H, OCH3 and halogen.
In some embodiments of the invention, the compound has the structure of formula III:
the molecular formula of the carrier compound shown in formula III is as follows: c67H120O6Wt. mol: 989.69, tentatively designated HO-TAGa.
In some embodiments of the invention, the polypeptide is a somaglutide.
The invention also provides a preparation method of the compound, which takes the compound with the structure shown in the formula 1 as a raw material and carries out esterification reaction, substitution reaction, reduction reaction and other steps to prepare the compound.
A compound of formula 1 having the formula C7H6O5，Mol.wt：170.12。
The synthetic route for the compounds of formula I is shown in FIG. 5.
The synthesis method comprises the following steps:
a) dissolving the substance shown in the formula 1 in methanol, and adding an esterification reaction reagent to perform esterification reaction to generate a compound 2;
b) adding the compound 2, an acid-binding agent and a substitution reaction reagent into a solvent, and reacting to generate a compound 3 in a formula;
c) dissolving the compound 3, adding a reducing agent a, and carrying out reduction reaction to generate a compound 4;
d) adding an oxidant a and an oxidant b into dichloromethane to serve as a solution A for later use; dissolving the compound 4 in triethylamine and dichloromethane, dropwise adding the solution into the solution A system, and carrying out oxidation reaction to generate a compound 5;
e) dissolving the compound 5, adding a solution containing a Grignard reagent, and reacting to obtain the compound represented by the formula III.
In a preferred embodiment of the present invention, in step a), the esterification reagent comprises acetyl chloride, thionyl chloride or concentrated sulfuric acid, and the molar ratio of the substance represented by formula 1 to the esterification reagent is 1: 1.5 to 3.0.
As a preferable scheme of the invention, in the step b), the acid-binding agent comprises one or a mixture of more than two of potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, diisopropylethylamine and triethylamine, the substitution reaction reagent is halogenated alkane RX, R is a C12-C22 fatty chain, and X is halogen; the molar ratio of the compound 2, the substitution reaction reagent and the acid-binding agent is 1.0: 3.2-4.0: 8.0 to 10.0.
As a preferred embodiment of the present invention, in step c), the reducing agent a comprises red aluminum or lithium aluminum hydride, and the molar ratio of the compound 3 to the reducing agent a is 1: 2.5 to 4.0.
As a preferred embodiment of the present invention, in step d), the oxidant a comprises pyridine sulfur trioxide, oxalyl chloride or NaClO, the oxidant b comprises DMSO or TEMPO, and the molar ratio of the compound 4, the oxidant a and the oxidant b is 1: 3.0-5.0: 10.0 to 12.0;
in step e), the Grignard reaction reagent comprises aryl magnesium chloride, aryl magnesium bromide or aryl lithium, and one or more of tetrahydrofuran, methyltetrahydrofuran and toluene as a solvent; the molar ratio of compound 5 to the format reaction reagent is 1: 2.0 to 3.0.
The method comprises the following specific steps:
(a) dissolving a substance shown in a formula (1) in methanol, and dropwise adding acetyl chloride to perform an esterification reaction to generate a compound shown in a formula (2); wherein the temperature of dripping acetyl chloride is 15-35 ℃, then the compound 2, namely the compound represented by the formula (2), is obtained by heat preservation reaction at 25-35 ℃ and post treatment.
(b) Adding the substance shown in the formula (2) into DMF (dimethyl formamide), completely dissolving, adding potassium carbonate and 1-bromooctadecane, heating to 85-90 ℃, carrying out heat preservation reaction, and carrying out post-treatment to obtain a compound 3, namely the compound shown in the formula (3).
(c) Dissolving the substance shown in the formula (3) in toluene, dropwise adding 70% of red aluminum toluene solution, carrying out reduction reaction, wherein the temperature of dropwise adding the red aluminum toluene is 10-30 ℃, the reaction temperature is 20-30 ℃, carrying out heat preservation reaction, and carrying out post-treatment to obtain a compound 4, namely the compound shown in the formula (4).
(d) Adding DMSO and pyridine sulfur trioxide into dichloromethane, dissolving a substance shown in a formula (4) in triethylamine and dichloromethane, dropwise adding the substance into the system, wherein the dropwise adding temperature is 5-15 ℃, the reaction temperature is 20-30 ℃, performing oxidation and heat preservation reaction, and performing post-treatment to obtain a compound 5, namely the compound shown in the formula (5).
(e) Dissolving the substance shown in the formula (5) in toluene, dropwise adding a 2M phenylmagnesium chloride tetrahydrofuran solution, wherein the dropwise adding temperature is 5-15 ℃, the reaction temperature is 20-30 ℃, carrying out heat preservation reaction, and then carrying out post-treatment to obtain the compound shown in the formula III.
The invention also provides the application of the compound shown in the formula I or the compound shown in the formula I prepared by the preparation method in polypeptide synthesis.
The invention also provides a liquid-phase synthesis method of the Somaltulip, which comprises the steps of taking the formula I II as a raw material, sequentially coupling amino acids, carrying out deprotection to prepare the Somalux fully-protected peptide, and then carrying out cracking to obtain the Somalux.
The synthetic route is shown in figure 6.
The synthesis of the somaglutide comprises the following steps:
1) coupling amino acids: the molar ratio of the substance of formula I to the amino acid is 1: 1.1 to 1.5, and the coupling reagent is HOBt/DIC (1.1 to 1.5equiv), HOBt/DCC (1.1 to 1.5equiv) HOBt/EDC.HCl (1.1 to 1.5equiv), preferably HOBt/DIC. The solvent used is chloroform or dichloromethane or tetrahydrofuran, with dichloromethane being preferred.
2) Removing Fmoc: the Fmoc removing reagent is DBU or piperidine, wherein DBU is preferred; the solvent used is chloroform or dichloromethane or tetrahydrofuran, with dichloromethane being preferred.
Compared with the prior art, the invention has the following beneficial effects:
1) so far, the compound shown in the formula I is used as a raw material, the Somalutide is prepared by a liquid phase synthesis method, domestic and foreign data are not reported in detail, and the preparation of the compound is described in detail for the first time;
2) the raw materials used in the invention are simple and easy to obtain, and the cost is low;
3) the synthesis steps in the invention are simple to operate and easy to realize.
Drawings
FIG. 1 is a scheme for preparing a compound of formula III according to the method of example 11H NMR spectrum.
FIG. 2 is an HPLC chromatogram of a fully protected hexapeptide prepared according to the method of example 1.
FIG. 3 is an HPLC chromatogram of a fully protected decapeptide prepared according to the method of example 1.
Figure 4 is an HPLC profile of somaglutide prepared according to the method of example 1.
FIG. 5 is a scheme showing the synthesis of the compound of formula I according to the present invention.
Fig. 6 is a synthetic route for the present invention somaglutide.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1:
1264.0mL of MeOH was added to a 2L three-necked reaction flask under nitrogen protection, 216.0g of trihydroxybenzoic acid was added, and the mixture was stirred to dissolve and clear, and then cooled to below 15 ℃. 180.5mL of acetyl chloride was added dropwise, and the reaction was monitored after the addition. Keeping the temperature below 35 ℃ for reaction for 6 hours; and (4) removing the solvent after the heat preservation reaction is finished. Then 2.0L of EA was added to dissolve it, and 500.0mL of water was added to extract it once. 15% NaHCO for EA phase3Extracting with water solution for 2 times, mixing water phases, and extracting with 300mL EA for 3 times; and (3) combining the EA phases, washing the combined EA phases once by using 300mL of saturated sodium chloride aqueous solution, adding 100.0g of anhydrous sodium sulfate for drying, filtering, removing a solvent to obtain 230.0g of a crude product, heating to dissolve the crude product by using 240.0mL of ethyl acetate, dripping 840.0mL of petroleum ether, cooling to 10-15 ℃, filtering, and drying to obtain 212.8g of a product of the formula 2, wherein the yield is 91.0%, and the purity is 99.8%.
Adding DMF1056.0 mL and 49.0g of compound of formula (2) into a 3.0L three-neck flask under the protection of nitrogen, stirring to dissolve, adding K2CO3293.8g, white turbid, added with 310.5g of 1-bromooctadecane, heated to over 90 ℃ and reacted for 16 h. Adding 1300mL of toluene, heating to 70 ℃ for dissolving, filtering while hot, heating the filtrate to 70 ℃, slowly cooling under mechanical stirring, and separating out solids. Filtering and drying to obtain 204.8g of the product of the formula 3 with the yield of 81.8 percent.
Under the protection of nitrogen, 2500.0mL of toluene and 500.0g of the compound of formula 3 were added into a 3L three-neck reaction flask, the temperature was raised to 45 ℃ or higher, the solution was cooled to below 25 ℃, 306.8g of 70% red aluminum toluene solution was added in portions, and the reaction was carried out for 1 hour with heat preservation. After heat preservation is finished, 1000.0mL of water is dripped to quench, and then concentrated hydrochloric acid is used for adjusting the pH value to 4-5; the layers were separated and the organic phase was washed with 400.0mL of saturated brine. Drying 100.0g of anhydrous sodium sulfate, filtering, removing the solvent, adding 1500.0mL of acetonitrile when about 1000.0mL of the anhydrous sodium sulfate remains, separating out the solid, slowly cooling to about 5-10 ℃, stirring for 20min, filtering, and drying to obtain 422.0g of a product in the formula 4, wherein the yield is 87.0%.
To a 1L three-necked reaction flask, 35.0mL of DCM and 27.0g of DMSO were added under nitrogen atmosphere, and the temperature was controlled to 15 ℃ or lower. 17.8g of pyridine sulfur trioxide are added in portions. After the addition was complete, the mixture was stirred for 1 hour. 32.4g of the compound of formula 4 and 21.3g of TEA were dissolved in 320.0mL of DCM, which was added to the above reaction flask. The temperature is controlled below 15 ℃. Stirred for 1 hour. Then heating to room temperature of 25-30 ℃, and reacting for 12 hours under the condition of heat preservation. The reaction was quenched by the addition of 450.0mL of 12% aqueous citric acid. Layering; the aqueous phase was extracted once with 120.0mL of DCM. The organic phases were combined and washed 2 times with 120.0mL of 6% aqueous sodium bicarbonate; the column was washed once with 120.0mL of saturated saline. 30.0g of anhydrous sodium sulfate was dried and filtered. Desolventizing, and stopping desolventizing when about 60.0mL of residual solvent is left; raising the temperature to 40-45 ℃, and completely dissolving and cleaning the solid. 180.0mL of ethyl acetate are added dropwise. And then slowly cooling to 5-10 ℃. Stirred for 30 minutes. Filtering and drying to obtain 24.3g of a product shown in the formula 5, wherein the yield is 74.8%.
To a 1L three-necked reaction flask, 200.0mL of THF and 20.0g of the compound of formula 5 were added under nitrogen, and the temperature was raised to 35 ℃ or higher. The solid is dissolved and cleaned, and then the temperature is reduced to below 25 ℃. Dripping device28.0mL of phenylmagnesium chloride was added and the temperature was controlled below 25 ℃. Keeping the temperature and reacting for 1 hour; adding 20.0mL of 10% citric acid aqueous solution to quench the reaction; THF was removed under reduced pressure, 240.0mL of DCM and 80.0mL of 10% aqueous citric acid were added, the layers were separated, and the organic phase was washed with 120.0mL of water and 120.0mL of saturated brine. 30.0g of anhydrous sodium sulfate was dried and filtered. And stopping desolventizing when the desolventizing amount is about 70.0mL, heating to 40-45 ℃, and dropwise adding 200.0mL of methanol after the desolventizing is completed. Cooling to 20-25 ℃, and stirring for 30 minutes; filtering and drying to obtain 19.6g of a product shown in the formula III with the yield of 90.2%.1H-NMR(500MHz，CDCl3) δ (ppm): 7.35(5H), 6.56(2H), 5.74(1H), 3.92(6H), 1.75(6H), 1.43(6H), 1.30(84H), 0.87(9H), see FIG. 1.
Synthesis of somaglutide:
1) Fmoc-Gly-O-TAGa (coupling of first amino acid):
to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 5.4g Fmoc-Gly-OH were added and 4.56g DCC were added in portions. The temperature is controlled below 30 ℃. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); and (5) filtering. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. Then slowly cooling to 10-15 deg.C. And (3) filtering and drying to obtain 19.1g of Fmoc-Gly-O-TAGa product. The yield thereof was found to be 99.3%.
2) H-Gly-O-TAGa (DeFmoc):
to a 250mL three-necked reaction flask, under nitrogen, 148.0mL of dichloromethane and 19.1g of Fmoc-Gly-TAGa were added and the solid solution was stirred. 44.4mL of DMF was added, the system was cooled to below 15 ℃ and 2.56g of DBU was added. The reaction was carried out at 10-15 ℃ for 30 minutes, and monitored by TLC (EA: PE: 1: 4). 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. The column was washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine.
3) Fmoc-Arg (Pbf) -Gly-O-TAGa (coupling of second amino acid):
11.7g of Fmoc-Arg (Pbf) -OH and 2.46g of HOBt were added to the three-necked flask under a nitrogen atmosphere, and the solid was stirred to remove the supernatant, followed by addition of 2.7g of DIC. The temperature is controlled below 30 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. Filtering and drying to obtain 24.7g of Fmoc-Arg (Pbf) -Gly-O-TAGa product. The yield thereof was found to be 97.8%.
4) Repeating the steps 2-3 to carry out coupling and deprotection of 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 and 14 amino acids to 15 amino acid Fmoc-Gln (Trt) -OH to obtain Fmoc-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa).
5) Synthesis of fully protected hexapeptide fragment (Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-OH):
to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 5.4g Fmoc-Gly-OH were added and 4.56g DCC were added in portions. The temperature is controlled below 30 ℃. The reaction was incubated for 5 hours and filtered. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. Then slowly cooling to 10-15 deg.C. Filtering and drying to obtain 19.1g of product. The yield thereof was found to be 99.3%. Repeating the steps of 2-3 to carry out coupling and deprotection of 2, 3, 4 and 5 amino acids to the coupling of 6 amino acid Fmoc-Ser (tBu) -OH to obtain Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-O-TAGa; to a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above supported hexapeptide were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with nitrogen and then purging with hydrogen. The hydrogenation reaction is carried out under the conditions of below 35 ℃ and below 0.04 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering, drying to obtain Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-OH 7.65g, yield 98.8%, HPLC liquid phase purity 96.0% (peak time: 3.9min), as shown in figure 2.
6) Synthesis of a fully protected decapeptide fragment (Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-OH):
to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 6.2g Fmoc-Val-OH were added and 4.56g DCC were added in portions. The temperature is controlled below 30 ℃. The reaction was incubated for 5 hours and filtered. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. Then slowly cooling to 10-15 deg.C. Filtering and drying to obtain 19.6g of product. The yield thereof was found to be 98.6%. Repeating the steps 2-3 to carry out coupling of amino acids 2, 3, 4, 5, 6, 7, 8 and 9 and coupling of deprotection to amino acid 10 Boc-His (Trt) -OH to obtain Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-O-TAGa; to a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above-mentioned fully protected decapeptide containing a carrier were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with nitrogen and then purging with hydrogen. The hydrogenation reaction is carried out under the conditions of below 35 ℃ and below 0.04 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering, drying to obtain the product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-OH 9.14g, yield 98.0%, HPLC liquid phase purity (peak time: 3.4min) 84.8%, as shown in figure 3.
7) Synthesis of heneicosin, Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa:
148.0mL of methylene chloride and 18.5g of the fully protected pentadecapeptide with carrier were added to a 250mL three-necked reaction flask under nitrogen protection, and the solid was stirred to dissolve it. 44.4mL of DMF was added, the system was cooled to below 15 ℃ and 0.72g of DBU was added. The reaction was monitored by TLC (EA: PE ═ 1:4) at 15 ℃ or below for 30 minutes. 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. Then washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine.
To the three-necked reaction flask, 5.8g of the fully protected hexapeptide and 0.74g of HOBt were added, and the solid was stirred to dissolve and 0.84g of DIC was added. The temperature is controlled below 30 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. After filtration and drying, the product Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa is 20.8 g. The yield thereof was found to be 95.0%.
8) Synthesis of thirty-one peptide, Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg Pbf) -Gly-Arg (Pbf) -Gly-OH:
166.0mL of methylene chloride and 20.8g of the fully protected heneicosin containing carrier described above were added to a 250mL three-necked reaction flask under nitrogen, and the solid was stirred to dissolve it. 44.4mL of DMF was added, the system was cooled to below 15 ℃ and 0.85g of DBU was added. The reaction was monitored by TLC (EA: PE ═ 1:4) at 15 ℃ or below for 30 minutes. 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. The column was washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine.
To the three-necked reaction flask, 7.6g of the fully protected decapeptide and 0.61g of HOBt were added, and the solid was stirred to dissolve and 0.97g of DIC was added. The temperature is controlled below 30 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. Filtering and drying to obtain a product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg-Pbf) -Gly-O-TAGa 24.4 g. The yield thereof was found to be 92.9%. To a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above carrier-containing fully protected triundecapeptide were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with hydrogen. The hydrogenation reaction is carried out under the conditions of below 35 ℃ and below 0.04 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering and drying to obtain a fully protected thirty-one peptide product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-OH 12.39g with the yield of 98.0%.
9) Cracking: Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-OH 12.3g, into a 250.0mL three-port reaction flask, followed by 74.0g of TFA: EDT (electro-thermal transfer coating): the lysate of Tis 90:5:5 (vol/vol) was lysed at 30 ℃ or less for 2 hours, added dropwise to 370.0g of isopropyl ether, precipitated, and filtered. The filter cake was then washed with 120.0g of acetonitrile: water 2: 1 (volume ratio), dissolving, filtering, and purifying the crude peptide in the mother liquor by a column. 5.9g of Desomo-rubicin, the yield is 69.7 percent, the HPLC liquid phase purity (peak time: 21.1min) is 99.8 percent, and the attached figure 4 shows.
Example 2:
adding 1264.0mL of MeOH into a 2L three-mouth reaction bottle under the protection of nitrogen, adding 216.0g of trihydroxybenzoic acid, stirring to dissolve, and cooling to 5-10 ℃. 180.5mL of acetyl chloride was added dropwise, and the reaction was monitored after the addition. Keeping the temperature at 25-35 ℃ for reaction for 6 hours; and (4) removing the solvent after the heat preservation reaction is finished. Then 2.0L of EA was added to dissolve it, and 500.0mL of water was added to extract it once. The EA phase was extracted 2 times with 15% aqueous NaHCO3, the combined aqueous phases were extracted 3 times with 300mL EA; and (3) combining the EA phases, washing the combined EA phases once by using 300mL of saturated sodium chloride aqueous solution, adding 100.0g of anhydrous sodium sulfate for drying, filtering, removing a solvent to obtain 230.0g of a crude product, heating to dissolve the crude product by using 240.0mL of ethyl acetate, dripping 840.0mL of petroleum ether, cooling to 10-15 ℃, filtering, and drying to obtain 215.1g of a product in the formula 2 with the yield of 92.0%.
Adding DMF1056.0 mL and 49.0g of compound of formula (2) into a 3.0L three-neck flask under the protection of nitrogen, stirring to dissolve, adding K2CO3293.8g, white and turbid, and added with 310.5g of 1-bromooctadecane, heated to 90 ℃ for reaction for 16 h. Adding 1300mL of toluene, heating to 70 ℃ for dissolving, filtering while hot, heating the filtrate to 70 ℃, slowly cooling under mechanical stirring, and separating out solids. Filtering and drying to obtain 200.3g of the product in the formula 3 with the yield of 80.0 percent.
Under the protection of nitrogen, 2500.0mL of toluene and 500.0g of the compound of formula 3 are added into a 3L three-mouth reaction bottle, the temperature is raised to 45 ℃ for clearing, the temperature is reduced to 25 ℃, 306.8g of 70% red aluminum toluene solution is added in batches, and the reaction is carried out for 1h under the condition of heat preservation. After heat preservation is finished, 1000.0mL of water is dripped to quench, and then concentrated hydrochloric acid is used for adjusting the pH value to 4-5; the layers were separated and the organic phase was washed with 400.0mL of saturated brine. Drying 100.0g of anhydrous sodium sulfate, filtering, removing the solvent, adding 1500.0mL of acetonitrile when the residual 1000.0mL of the anhydrous sodium sulfate is about, separating out the solid, slowly cooling to about 5-10 ℃, stirring for 20min, filtering, and drying to obtain 417.1g of a product of the formula 4 with the yield of 86.0%.
Under the protection of nitrogen, 35.0mL of DCM and 27.0g of DMSO are added into a 1L three-port reaction bottle, and the temperature is controlled to be 5-10 ℃. 17.8g of pyridine sulfur trioxide are added in portions. After the addition was complete, the mixture was stirred for 1 hour. 32.4g of the compounds of formula 4 and 21.3g of TEA were dissolved in 320.0ml of CCM and added to the above reaction flask. Controlling the temperature to be 5-15 ℃. Stirred for 1 hour. Then heating to room temperature of 25-30 ℃, and reacting for 12 hours under the condition of heat preservation. The reaction was quenched by the addition of 450.0mL of 12% aqueous citric acid. Layering; the aqueous phase was extracted once with 120.0ml of EDC. The combined organic phases were washed 2 times with 120.0mL 6% aqueous sodium bicarbonate; the column was washed once with 120.0mL of saturated saline. 30.0g of anhydrous sodium sulfate was dried and filtered. Desolventizing, and stopping desolventizing when about 60.0mL of residual solvent is left; raising the temperature to 40-45 ℃, and completely dissolving and cleaning the solid. 180.0mL of ethyl acetate are added dropwise. And then slowly cooling to 5-10 ℃. Stirred for 30 minutes. Filtering and drying to obtain 24.4g of a product shown in the formula 5 with the yield of 75.0%.
Under the protection of nitrogen, THF 200.0mL and 20.0g of compound formula 5 are added to a 1L three-necked reaction flask and the temperature is raised to 30-35 ℃. The solid is dissolved clearly and then cooled to 20-25 ℃. 28.0mL of phenylmagnesium chloride is dripped, and the temperature is controlled to be 20-25 ℃. Keeping the temperature and reacting for 1 hour; adding 20.0mL of 10% citric acid aqueous solution to quench the reaction; THF was removed under reduced pressure, 240.0mL of LPCM and 80.0mL of 10% aqueous citric acid were added, the layers were separated, and the organic phase was washed with 120.0mL of water and 120.0mL of saturated brine. 30.0g of anhydrous sodium sulfate was dried and filtered. And stopping desolventizing when the desolventizing amount is about 70.0mL, heating to 40-45 ℃, and dropwise adding 200.0mL of methanol after the desolventizing is completed. Cooling to 20-25 ℃, and stirring for 30 minutes; filtering and drying to obtain 19.3g of a product in the formula III with the yield of 89.0%.
Synthesis of somaglutide:
1) Fmoc-Gly-O-TAGa (coupling of first amino acid):
to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP, and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 5.4g of Fmoc-Gly-OH were added and 2.76g of DIC were added in portions. Controlling the temperature to be 25-30 ℃. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); and (5) filtering. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. And then slowly cooling to 10-15 ℃. And (3) filtering and drying to obtain 19.0g of Fmoc-Gly-O-TAGa product. The yield thereof was found to be 98.8%.
2) H-Gly-O-TAGa (DeFmoc):
to a 250mL three-necked reaction flask, under nitrogen, 148.0mL of dichloromethane and 19.0g of Fmoc-Gly-TAGa were added and the solid solution was stirred. Adding 44.4mL of DMF, cooling the system to 10-15 ℃, and adding 14.8g of piperidine. The reaction was carried out at 10-15 ℃ for 30 minutes, and monitored by TLC (EA: PE: 1: 4). 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. The column was washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine.
3) Fmoc-Arg (Pbf) -Gly-O-TAGa (coupling of second amino acid):
under the protection of nitrogen, 11.5g of Fmoc-Arg (Pbf) -OH and 2.45g of HOBt were added to the three-necked reaction flask, and the solid was stirred to be clear, followed by addition of 4.4g of DCC. The temperature is controlled below 30 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. Filtering and drying to obtain 24.5g of Fmoc-Arg (Pbf) -Gly-O-TAGa product. The yield thereof was found to be 97.0%.
4) Repeating the steps 2-3 to carry out coupling and deprotection of 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 and 14 amino acids to 15 amino acid Fmoc-Gln (Trt) -OH to obtain Fmoc-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa).
5) Synthesis of fully protected hexapeptide fragment (Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-OH):
to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 5.4g Fmoc-Gly-OH was added and 2.76g DIC were added in portions. Controlling the temperature to be 25-30 ℃. The reaction was incubated for 5 hours and filtered. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. And then slowly cooling to 10-15 ℃. Filtering and drying to obtain 19.1g of product. The yield thereof was found to be 99.3%. Repeating 2-3 to carry out coupling of 2 nd, 3 th, 4 th and 5 th amino acids and coupling of deprotection to 6 th amino acid Fmoc-Ser (tBu) -OH. Obtaining Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-O-TAGa; to a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above supported hexapeptide were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with nitrogen and then purging with hydrogen. Carrying out hydrogenation reaction at 30-35 ℃ under the pressure of 0.02-0.04 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering and drying to obtain 7.66g of Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-OH of the fully-protected hexapeptide product with the yield of 99.0 percent.
6) Synthesis of a fully protected decapeptide fragment (Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-OH):
to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 5.8g of Fmoc-Val-OH were added and 2.6g of DIC were added in portions. Controlling the temperature to be 25-30 ℃. The reaction was incubated for 5 hours and filtered. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. And then slowly cooling to 10-15 ℃. Filtering and drying to obtain 19.3g of product. The yield thereof was found to be 98.5%. Repeating the steps 2-3 to carry out coupling of 2, 3, 4, 5, 6, 7, 8 and 9 amino acids and coupling of deprotection to 10 amino acid Boc-His (Trt) -OH. Obtaining Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-O-TAGa; to a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above-mentioned fully protected decapeptide containing a carrier were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with nitrogen and then purging with hydrogen. Carrying out hydrogenation reaction at 30-35 ℃ and 0.02-0.04 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering and drying to obtain a fully protected decapeptide product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-OH 9.2g with the yield of 98.6 percent.
7) Synthesis of heneicosin, Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa:
148.0mL of methylene chloride and 18.5g of the fully protected pentadecapeptide with carrier were added to a 250mL three-necked reaction flask under nitrogen protection, and the solid was stirred to dissolve it. Adding 44.4mL of DMF, cooling the system to 10-15 ℃, and adding 14.8g of piperidine. The reaction was carried out at 10-15 ℃ for 30 minutes, and monitored by TLC (EA: PE: 1: 4). 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. The column was washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine.
5.5g of the fully protected hexapeptide and 0.68g of HOBt were added to the three-necked reaction flask, and the solid was stirred to dissolve and then 1.26g of DCC was added thereto. Controlling the temperature to be 25-30 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. Filtering and drying to obtain a product Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa 21.0 g. The yield thereof was found to be 95.9%.
8) Synthesis of thirty-one peptide, Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg Pbf) -Gly-Arg (Pbf) -Gly-OH: 166.0mL of methylene chloride and 20.8g of the fully protected heneicosin containing carrier described above were added to a 250mL three-necked reaction flask under nitrogen, and the solid was stirred to dissolve it. Adding 44.4mL of DMF, cooling the system to 10-15 ℃, and adding 0.85g of DBU. The reaction was carried out at 10-15 ℃ for 30 minutes, and monitored by TLC (EA: PE: 1: 4). 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. The column was washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine.
To the three-necked reaction flask, 7.6g of the fully protected decapeptide product and 0.61g of HOBt were added, and the solid was stirred to dissolve and clear, followed by addition of 1.59g of DCC. Controlling the temperature to be 25-30 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. After filtration and drying, the product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg Pbf) -Gly-OTAGa 24.6g is obtained. The yield thereof was found to be 93.7%. To a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above carrier-containing fully protected triundecapeptide were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with hydrogen. Carrying out hydrogenation reaction at 30-35 ℃ and 0.02-0.04 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering and drying to obtain a fully protected thirty-one peptide product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-OH 12.4g with the yield of 96.9%.
10) Cracking: Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-OH 12.4g, into a 250.0mL three-port reaction flask, followed by 146.4g of TFA: EDT (electro-thermal transfer coating): and (3) cracking the lysate with Tis being 90:5:5 (volume ratio) at 25-30 ℃ for 2 hours, dropwise adding the lysate into 500.0g of isopropyl ether, precipitating and filtering. The filter cake was then washed with 250.0g of acetonitrile: water 2: 1 (volume ratio), washing, filtering, and purifying the crude peptide in the mother liquor through a column. 5.64g of Desomo-L-peptide with the yield of 66.0%.
Example 3:
adding 1264.0mL of MeOH into a 2L three-mouth reaction bottle under the protection of nitrogen, adding 216.0g of trihydroxybenzoic acid, stirring to dissolve, and cooling to 0-5 ℃. 180.5mL of acetyl chloride was added dropwise, and the reaction was monitored after the addition. Reacting for 6 hours at the temperature of 20-30 ℃; and (4) removing the solvent after the heat preservation reaction is finished. Then 2.0L of EA was added to dissolve it, and 500.0mL of water was added to extract it once. 15% NaHCO for EA phase3Extracting with water for 2 times, and mixingAnd the aqueous phase was extracted 3 times with 300mL EA; and (3) combining the EA phases, washing the combined EA phases once by using 300mL of saturated sodium chloride aqueous solution, adding 100.0g of anhydrous sodium sulfate for drying, filtering, removing a solvent to obtain 230.0g of a crude product, heating to dissolve the crude product by using 240.0mL of ethyl acetate, dripping 840.0mL of petroleum ether, cooling to 10-15 ℃, filtering, and drying to obtain 213.2g of a product of the formula 2 with the yield of 91.2%.
Adding DMF1056.0 mL and 49.0g of compound of formula (2) into a 3.0L three-neck flask under the protection of nitrogen, stirring to dissolve, adding K2CO3293.8g, white and turbid, and added with 310.5g of 1-bromooctadecane to react for 16h at 100 ℃. Adding 1300mL of toluene, heating to 70 ℃ for dissolving, filtering while hot, heating the filtrate to 70 ℃, slowly cooling under mechanical stirring, and separating out solids. Filtering and drying to obtain 203.5g of a product in the formula 3 with the yield of 81.3 percent.
Under the protection of nitrogen, 2500.0mL of toluene and 500.0g of the compound of formula 3 are added into a 3L three-mouth reaction bottle, the temperature is raised to 50 ℃ for clearing, the temperature is reduced to 20 ℃, 306.8g of 70% red aluminum toluene solution is added in batches, and the reaction is carried out for 1h under the condition of heat preservation. After heat preservation is finished, 1000.0mL of water is dripped to quench, and then concentrated hydrochloric acid is used for adjusting the pH value to 4-5; the layers were separated and the organic phase was washed with 400.0mL of saturated brine. Drying 100.0g of anhydrous sodium sulfate, filtering, removing the solvent, adding 1500.0mL of acetonitrile when about 1000.0mL of the anhydrous sodium sulfate remains, separating out the solid, slowly cooling to 5-10 ℃, stirring for 20min, filtering, and drying to obtain 421.0g of the product of the formula 4, wherein the yield is 86.8%.
Under the protection of nitrogen, 35.0mL of DCM and 27.0g of DMSO are added into a 1L three-port reaction bottle, and the temperature is controlled at 0-5 ℃. 17.8g of pyridine sulfur trioxide are added in portions. After the addition was complete, the mixture was stirred for 1 hour. 32.4g of the compound of formula 4 and 21.3g of TEA were dissolved in 320.0mL of DCM, which was added to the above reaction flask. Controlling the temperature to be 5-10 ℃. Stirred for 1 hour. And then heating to room temperature of 30-35 ℃, and reacting for 15 hours under the condition of heat preservation. The reaction was quenched by the addition of 450.0mL of 12% aqueous citric acid. Layering; the aqueous phase was extracted once with 120.0mL of DCM. The combined organic phases were washed 2 times with 120.0mL 6% aqueous sodium bicarbonate; the column was washed once with 120.0mL of saturated saline. 30.0g of anhydrous sodium sulfate was dried and filtered. Desolventizing, and stopping desolventizing when about 60.0mL of residual solvent is left; raising the temperature to 40-45 ℃, and completely dissolving and cleaning the solid. 180.0mL of ethyl acetate are added dropwise. And then slowly cooling to 5-10 ℃. Stirred for 30 minutes. Filtering and drying to obtain 24.5g of a product shown in the formula 5 with the yield of 75.4%.
Under nitrogen protection, a 1L three-necked reaction flask is charged with 200.0mL of 2-MeTHF and 20.0g of the compound of formula 7, and the temperature is raised to 35-40 ℃. Dissolving the solid clearly, and then cooling to 25-30 ℃. 28.0mL of phenylmagnesium chloride is dripped, and the temperature is controlled to be 20-25 ℃. Keeping the temperature and reacting for 1 hour; adding 20.0mL of 10% citric acid aqueous solution to quench the reaction; 2-MeTHF was removed under reduced pressure, 240.0mL of DCM and 80.0mL of 10% aqueous citric acid were added, the layers were separated, and the organic phase was washed with 120.0mL of water and 120.0mL of saturated brine. 30.0g of anhydrous sodium sulfate was dried and filtered. And stopping desolventizing when the desolventizing amount is about 70.0mL, heating to 40-45 ℃, and dropwise adding 200.0mL of methanol after the desolventizing is completed. Cooling to 20-25 ℃, and stirring for 30 minutes; filtering and drying to obtain 20.0g of a product in the formula III with the yield of 92.0 percent.
Synthesis of somaglutide:
1) Fmoc-Gly-O-TAGa (coupling of first amino acid):
to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP, and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 5.4g of Fmoc-Gly-OH were added and 2.76g of DIC were added in portions. The temperature is controlled to be 15-20 ℃. The reaction was incubated for 7 hours and monitored by TLC (EA: PE ═ 1: 4); and (5) filtering. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. And then slowly cooling to 10-15 ℃. And (3) filtering and drying to obtain 19.2g of Fmoc-Gly-O-TAGa product. The yield thereof was found to be 99.8%.
2) H-Gly-O-TAGa (DeFmoc):
to a 250mL three-necked reaction flask, under nitrogen, 148.0mL of dichloromethane and 19.0g of Fmoc-Gly-TAGa were added and the solid solution was stirred. Adding 44.4mL of DMF, cooling the system to 20-25 ℃, and adding 14.8g of piperidine. The reaction was carried out at 20-25 ℃ for 30 minutes, and monitored by TLC (EA: PE: 1: 4). 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. Then washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine.
3) Fmoc-Arg (Pbf) -Gly-O-TAGa (coupling of second amino acid): under the protection of nitrogen, 11.5g of Fmoc-Arg (Pbf) -OH and 2.45g of HOBt were added to the three-necked reaction flask, and the solid was stirred to be clear, followed by addition of 4.4g of DCC. The temperature is controlled to be 20-25 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. Filtering and drying to obtain 24.3g of Fmoc-Arg (Pbf) -Gly-O-TAGa product. The yield thereof was found to be 96.2%.
4) Repeating the steps 2-3 to carry out coupling and deprotection of 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 and 14 amino acids to 15 amino acid Fmoc-Gln (Trt) -OH to obtain Fmoc-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa).
5) Synthesis of fully protected hexapeptide fragment (Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-OH): to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 5.4g Fmoc-Gly-OH was added and 2.76g DIC were added in portions. The temperature is controlled to be 20-25 ℃. The reaction was incubated for 7 hours and filtered. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. And then slowly cooling to 10-15 ℃. Filtering and drying to obtain 18.9g of product. The yield thereof was found to be 98.3%. Repeating the steps of 2-3 to carry out coupling and deprotection of 2, 3, 4 and 5 amino acids to the coupling of 6 amino acid Fmoc-Ser (tBu) -OH to obtain Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-O-TAGa; to a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above supported hexapeptide were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with nitrogen and then purging with hydrogen. Carrying out hydrogenation reaction at 35-40 ℃ and under the pressure of 0.01-0.02 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering and drying to obtain 7.7g of Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-OH of the fully-protected hexapeptide product with the yield of 99.5 percent.
6) Synthesis of a fully protected decapeptide fragment (Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-OH): to a 250mL three-necked reaction flask, under nitrogen, was added 150.0mL of dichloromethane, 0.43g of DMAP, and 15.0g of the compound of formula III, and the solid was stirred to dissolve it. 5.6g of Fmoc-Val-OH were added and 2.7g of DIC were added in portions. The temperature is controlled to be 20-25 ℃. The reaction was incubated for 5 hours and filtered. And (3) stopping desolventizing when about 80.0g of the remaining desolventizing agent is removed, raising the temperature to 40-45 ℃, and dropwise adding 180.0mL of methanol after the solid is dissolved clearly. And then slowly cooling to 10-15 ℃. Filtering and drying to obtain 19.5g of product. The yield thereof was found to be 99.4%. Repeating the steps 2-3 to carry out coupling of 2, 3, 4, 5, 6, 7, 8 and 9 amino acids and coupling of deprotection to 10 amino acid Boc-His (Trt) -OH. Obtaining Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-O-TAGa; to a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above-mentioned fully protected decapeptide containing a carrier were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with nitrogen and then purging with hydrogen. Carrying out hydrogenation reaction at 35-40 ℃ and 0.01-0.02 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering and drying to obtain a fully protected decapeptide product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-OH 9.1g with the yield of 97.5 percent.
7) Synthesis of heneicosin, Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa: 148.0mL of methylene chloride and 18.5g of the fully protected pentadecapeptide with carrier were added to a 250mL three-necked reaction flask under nitrogen protection, and the solid was stirred to dissolve it. Adding 44.4mL of DMF, cooling the system to 10-15 ℃, and adding 14.8g of piperidine. The reaction was carried out at 20-25 ℃ for 30 minutes, and monitored by TLC (EA: PE: 1: 4). 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. The column was washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine.
5.6g of the fully protected hexapeptide and 0.68g of HOBt were added to the three-necked reaction flask, and the solid was stirred to dissolve and then 1.26g of DCC was added thereto. The temperature is controlled to be 20-25 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. Filtering and drying to obtain a product Fmoc-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-O-TAGa 21.2 g. The yield thereof was found to be 96.8%.
8) Synthesis of thirty-one peptide, Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg Pbf) -Gly-Arg (Pbf) -Gly-OH: 166.0mL of methylene chloride and 20.8g of the fully protected heneicosin containing carrier described above were added to a 250mL three-necked reaction flask under nitrogen, and the solid was stirred to dissolve it. Adding 44.4mL of DMF, cooling the system to 20-25 ℃, and adding 0.9g of DBU. The reaction was carried out at 15-20 ℃ for 30 minutes, and monitored by TLC (EA: PE: 1: 4). 60.0mL of 5% citric acid aqueous solution was added, and the mixture was washed and separated. The column was washed with 60.0mL of water and 60.0mL of 5% aqueous sodium bicarbonate solution. 60.0mL of the mixture was washed with water and 60.0mL of the mixture was washed with brine. To the three-necked reaction flask, 7.7g of the fully protected decapeptide and 0.61g of HOBt were added, and the solid was stirred to dissolve and then 1.59g of DCC was added. Controlling the temperature to be 25-30 ℃. The reaction was incubated for 3 hours. Desolventizing to a viscous state, stopping desolventizing, and adding 180.0mL of methanol. Stirred for 30 minutes. Filtering and drying to obtain a product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg-Pbf) -Gly-O-TAGa 24.8 g. The yield thereof was found to be 94.4%.
To a 500mL hydrogenation reactor, 150.0mL of tetrahydrofuran and 15.0g of the above carrier-containing fully protected triundecapeptide were added, the solid solution was stirred, purged with nitrogen, and then 1.5g of 10% wet palladium on carbon (containing 50% water) was added, followed by purging with hydrogen. Carrying out hydrogenation reaction at 35-40 ℃ and 0.01-0.02 MPa. The reaction was incubated for 5 hours and monitored by TLC (EA: PE: 1: 4); filtering, and concentrating the mother liquor under reduced pressure to obtain solid. 200.0mL of methanol was added, stirred for 30 minutes, and filtered. The mother liquor was concentrated under reduced pressure to a solid, and 60.0mL of n-heptane was added thereto and stirred for 30 minutes. Filtering and drying to obtain a fully protected thirty-one peptide product Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-OH 12.5g with the yield of 97.6%.
9) Cracking: Boc-His (Trt) -Aib-Glu (OtBu) -Gly-Thr (OtBu) -Phe-Thr (OtBu) -Ser (OtBu) -Gln (OtBu) -Val-Ser (OtBu) -Tyr (OtBu) -Leu-Glu (OtBu) -Gly-Gln (Trt) -Ala-Ala-Lys [ Oct (OtBu) -Glu (AEEA-AEEA) -OtBu ] -Glu (OtBu) -Phe-Ile-Ala-Trp (Boc) -Leu-Val-Arg (Pbf) -Gly-OH 12.4g, into a 250.0mL three-port reaction flask, followed by 146.4g of TFA: EDT (electro-thermal transfer coating): and (3) cracking the lysate with Tis being 90:5:5 (volume ratio) at 30-35 ℃ for 2 hours, dropwise adding the lysate into 500.0g of isopropyl ether, precipitating and filtering. The filter cake was then washed with 250.0g of acetonitrile: water 2: 1 (volume ratio), washing, filtering, and purifying the crude peptide in the mother liquor through a column. 5.7g of Desomareuptate, and the yield is 66.7%.
In the present invention, the amino acid sequences of the polypeptides involved are shown in Table 1.
Table 1: amino acid sequence listing
While the invention has been described with respect to a preferred embodiment, it will be understood by those skilled in the art that the foregoing and other changes, omissions and deviations in the form and detail thereof may be made without departing from the scope of this invention. Those skilled in the art can make various changes, modifications and equivalent arrangements, which are equivalent to the embodiments of the present invention, without departing from the spirit and scope of the present invention, and which may be made by utilizing the techniques disclosed above; meanwhile, any changes, modifications and variations of the above-described embodiments, which are equivalent to those of the technical spirit of the present invention, are within the scope of the technical solution of the present invention.
Claims (10)
1. A diaryl benzyl alcohol compound used as a polypeptide liquid phase synthesis carrier is characterized in that the structural formula of the diaryl benzyl alcohol compound is shown as a formula I:
2. The diaryl benzyl alcohol compound as the carrier of the liquid phase synthesis of polypeptide of claim 1, wherein the structural formula of the diaryl benzyl alcohol compound is shown as formula III:
3. the preparation method of diaryl benzyl alcohol compounds as claimed in claim 1, characterized in that, the compound with the structure as shown in formula 1 is used as raw material, and the compound with the structure as shown in formula I is prepared by esterification, substitution, reduction, oxidation and addition, hydrolysis and precipitation;
4. the method for producing diarylbenzyl alcohol compounds according to claim 4, wherein the method comprises the steps of:
a) dissolving the substance shown in the formula 1 in methanol, and adding an esterification reaction reagent to perform esterification reaction to generate a compound 2;
b) adding the compound 2, an acid-binding agent and a substitution reaction reagent into a solvent, and reacting to generate a compound 3 in a formula;
c) dissolving the compound 3, adding a reducing agent a, and carrying out reduction reaction to generate a compound 4;
d) adding an oxidant a and an oxidant b into dichloromethane to serve as a solution A for later use; dissolving the compound 4 in triethylamine and dichloromethane, dropwise adding the solution into the solution A system, and carrying out oxidation reaction to generate a compound 5;
e) dissolving the compound 5, adding a solution containing a Grignard reagent, and reacting to generate the compound represented by the formula I.
5. The method of claim 4, wherein the esterification reagent in step a) comprises acetyl chloride, thionyl chloride or concentrated sulfuric acid, and the molar ratio of the substance represented by formula 1 to the esterification reagent is 1: 1.5 to 3.0.
6. The method for preparing diaryl benzyl alcohol compounds according to claim 4, wherein in the step b), the acid-binding agent comprises one or a mixture of more than two of potassium carbonate, sodium carbonate, potassium hydroxide, sodium hydroxide, diisopropylethylamine and triethylamine, the substitution reaction reagent is halogenated alkane RX, R is C12-C22 fatty chain, and X is halogen; the molar ratio of the compound 2, the substitution reaction reagent and the acid-binding agent is 1.0: 3.2-4.0: 8.0 to 10.0.
7. The method for preparing diarylbenzyl alcohol compounds according to claim 4, wherein in step c), the reducing agent a comprises red aluminum or lithium aluminum hydride, and the molar ratio of compound 3 to reducing agent a is 1: 2.5 to 4.0.
8. The method for preparing diarylbenzyl alcohol compounds according to claim 4, wherein in step d), the oxidant a comprises pyridine sulfur trioxide, oxalyl chloride or NaClO, the oxidant b comprises DMSO or TEMPO, and the molar ratio of the compound 4 to the oxidant a to the oxidant b is 1: 3.0-5.0: 10.0 to 12.0;
in step e), the Grignard reaction reagent comprises aryl magnesium chloride, aryl magnesium bromide or aryl lithium, and one or more of tetrahydrofuran, methyltetrahydrofuran and toluene as a solvent; the molar ratio of compound 5 to the format reaction reagent is 1: 2.0 to 3.0.
9. The use of a diaryl benzyl alcohol compound according to claim 1, wherein the compound according to claim 1 is used as a hydrophobic carrier for the synthesis of a fully protected polypeptide or a deprotected polypeptide in a liquid phase.
10. The use of diaryl benzyl alcohol compounds according to claim 9, wherein the compounds of formula I are hydrophobic carriers, amino acids are sequentially coupled by condensation reactions to produce protected polypeptide chains containing a start-end carrier, and deprotection is carried out to produce polypeptide chains not containing a start-end carrier.
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