CN106749542A - A kind of synthetic method of Fertirelin - Google Patents
A kind of synthetic method of Fertirelin Download PDFInfo
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- CN106749542A CN106749542A CN201611227425.1A CN201611227425A CN106749542A CN 106749542 A CN106749542 A CN 106749542A CN 201611227425 A CN201611227425 A CN 201611227425A CN 106749542 A CN106749542 A CN 106749542A
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- China
- Prior art keywords
- resin
- dmf
- fertirelin
- peptide
- pro
- Prior art date
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- DGCPIBPDYFLAAX-YTAGXALCSA-N fertirelin Chemical compound CCNC(=O)[C@@H]1CCCN1C(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CC(C)C)NC(=O)CNC(=O)[C@@H](NC(=O)[C@H](CO)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CC=1NC=NC=1)NC(=O)[C@H]1NC(=O)CC1)CC1=CC=C(O)C=C1 DGCPIBPDYFLAAX-YTAGXALCSA-N 0.000 title claims abstract description 43
- 229950001491 fertirelin Drugs 0.000 title claims abstract description 42
- 108700020627 fertirelin Proteins 0.000 title claims abstract description 42
- 238000010189 synthetic method Methods 0.000 title claims abstract description 21
- 239000011347 resin Substances 0.000 claims abstract description 72
- 229920005989 resin Polymers 0.000 claims abstract description 72
- 238000006243 chemical reaction Methods 0.000 claims abstract description 50
- 108090000765 processed proteins & peptides Proteins 0.000 claims abstract description 35
- 102000007079 Peptide Fragments Human genes 0.000 claims abstract description 14
- 108010033276 Peptide Fragments Proteins 0.000 claims abstract description 14
- -1 Chlorotrityl Chemical group 0.000 claims abstract description 8
- XFWCSGJOVUQCME-YUMQZZPRSA-N pEH Chemical compound C([C@@H](C(=O)O)NC(=O)[C@H]1NC(=O)CC1)C1=CNC=N1 XFWCSGJOVUQCME-YUMQZZPRSA-N 0.000 claims abstract description 8
- 238000000746 purification Methods 0.000 claims abstract description 4
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims description 113
- 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 99
- 239000003153 chemical reaction reagent Substances 0.000 claims description 32
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 claims description 26
- 238000005859 coupling reaction Methods 0.000 claims description 26
- 230000008878 coupling Effects 0.000 claims description 23
- 238000010168 coupling process Methods 0.000 claims description 23
- 238000005406 washing Methods 0.000 claims description 23
- 239000000243 solution Substances 0.000 claims description 20
- 230000000903 blocking effect Effects 0.000 claims description 14
- QUSNBJAOOMFDIB-UHFFFAOYSA-N Ethylamine Chemical compound CCN QUSNBJAOOMFDIB-UHFFFAOYSA-N 0.000 claims description 12
- 239000002904 solvent Substances 0.000 claims description 12
- 125000003088 (fluoren-9-ylmethoxy)carbonyl group Chemical group 0.000 claims description 11
- JFLSOKIMYBSASW-UHFFFAOYSA-N 1-chloro-2-[chloro(diphenyl)methyl]benzene Chemical compound ClC1=CC=CC=C1C(Cl)(C=1C=CC=CC=1)C1=CC=CC=C1 JFLSOKIMYBSASW-UHFFFAOYSA-N 0.000 claims description 10
- AHDSRXYHVZECER-UHFFFAOYSA-N 2,4,6-tris[(dimethylamino)methyl]phenol Chemical compound CN(C)CC1=CC(CN(C)C)=C(O)C(CN(C)C)=C1 AHDSRXYHVZECER-UHFFFAOYSA-N 0.000 claims description 10
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical group OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 claims description 10
- NPZTUJOABDZTLV-UHFFFAOYSA-N hydroxybenzotriazole Substances O=C1C=CC=C2NNN=C12 NPZTUJOABDZTLV-UHFFFAOYSA-N 0.000 claims description 10
- 238000003756 stirring Methods 0.000 claims description 10
- 239000003795 chemical substances by application Substances 0.000 claims description 8
- 238000005520 cutting process Methods 0.000 claims description 8
- 150000007530 organic bases Chemical class 0.000 claims description 8
- 150000001413 amino acids Chemical class 0.000 claims description 7
- 239000012043 crude product Substances 0.000 claims description 7
- 238000006467 substitution reaction Methods 0.000 claims description 7
- ZPGDWQNBZYOZTI-SFHVURJKSA-N (2s)-1-(9h-fluoren-9-ylmethoxycarbonyl)pyrrolidine-2-carboxylic acid Chemical group OC(=O)[C@@H]1CCCN1C(=O)OCC1C2=CC=CC=C2C2=CC=CC=C21 ZPGDWQNBZYOZTI-SFHVURJKSA-N 0.000 claims description 6
- 238000001514 detection method Methods 0.000 claims description 6
- 239000000706 filtrate Substances 0.000 claims description 6
- FEMOMIGRRWSMCU-UHFFFAOYSA-N ninhydrin Chemical compound C1=CC=C2C(=O)C(O)(O)C(=O)C2=C1 FEMOMIGRRWSMCU-UHFFFAOYSA-N 0.000 claims description 6
- 229910052698 phosphorus Inorganic materials 0.000 claims description 6
- 239000011574 phosphorus Substances 0.000 claims description 6
- 230000009514 concussion Effects 0.000 claims description 5
- XWBDWHCCBGMXKG-UHFFFAOYSA-N ethanamine;hydron;chloride Chemical compound Cl.CCN XWBDWHCCBGMXKG-UHFFFAOYSA-N 0.000 claims description 5
- PNGLEYLFMHGIQO-UHFFFAOYSA-M sodium;3-(n-ethyl-3-methoxyanilino)-2-hydroxypropane-1-sulfonate;dihydrate Chemical compound O.O.[Na+].[O-]S(=O)(=O)CC(O)CN(CC)C1=CC=CC(OC)=C1 PNGLEYLFMHGIQO-UHFFFAOYSA-M 0.000 claims description 5
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 5
- DKIDEFUBRARXTE-UHFFFAOYSA-N 3-mercaptopropanoic acid Chemical compound OC(=O)CCS DKIDEFUBRARXTE-UHFFFAOYSA-N 0.000 claims description 4
- FGUUSXIOTUKUDN-IBGZPJMESA-N C1(=CC=CC=C1)N1C2=C(NC([C@H](C1)NC=1OC(=NN=1)C1=CC=CC=C1)=O)C=CC=C2 Chemical compound C1(=CC=CC=C1)N1C2=C(NC([C@H](C1)NC=1OC(=NN=1)C1=CC=CC=C1)=O)C=CC=C2 FGUUSXIOTUKUDN-IBGZPJMESA-N 0.000 claims description 4
- 238000001914 filtration Methods 0.000 claims description 4
- 239000000203 mixture Substances 0.000 claims description 4
- 238000002360 preparation method Methods 0.000 claims description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical group CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 3
- 244000248349 Citrus limon Species 0.000 claims description 3
- 235000005979 Citrus limon Nutrition 0.000 claims description 3
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N Phenol Chemical compound OC1=CC=CC=C1 ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 claims description 3
- 239000007864 aqueous solution Substances 0.000 claims description 3
- 239000004327 boric acid Substances 0.000 claims description 3
- 239000012141 concentrate Substances 0.000 claims description 3
- 230000006837 decompression Effects 0.000 claims description 3
- 238000011068 loading method Methods 0.000 claims description 3
- 239000011259 mixed solution Substances 0.000 claims description 3
- 238000002390 rotary evaporation Methods 0.000 claims description 3
- 150000003839 salts Chemical class 0.000 claims description 3
- 229920006395 saturated elastomer Polymers 0.000 claims description 3
- 239000007787 solid Substances 0.000 claims description 3
- 238000000967 suction filtration Methods 0.000 claims description 3
- 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 claims description 2
- ODHCTXKNWHHXJC-VKHMYHEASA-N 5-oxo-L-proline Chemical compound OC(=O)[C@@H]1CCC(=O)N1 ODHCTXKNWHHXJC-VKHMYHEASA-N 0.000 claims description 2
- 241000254173 Coleoptera Species 0.000 claims description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 2
- 239000002253 acid Substances 0.000 claims description 2
- 210000004899 c-terminal region Anatomy 0.000 claims description 2
- 150000002148 esters Chemical class 0.000 claims description 2
- 239000012071 phase Substances 0.000 claims description 2
- 150000003053 piperidines Chemical class 0.000 claims description 2
- 238000010532 solid phase synthesis reaction Methods 0.000 claims description 2
- 238000005292 vacuum distillation Methods 0.000 claims description 2
- ODHCTXKNWHHXJC-GSVOUGTGSA-N Pyroglutamic acid Natural products OC(=O)[C@H]1CCC(=O)N1 ODHCTXKNWHHXJC-GSVOUGTGSA-N 0.000 claims 1
- ODHCTXKNWHHXJC-UHFFFAOYSA-N acide pyroglutamique Natural products OC(=O)C1CCC(=O)N1 ODHCTXKNWHHXJC-UHFFFAOYSA-N 0.000 claims 1
- 235000015110 jellies Nutrition 0.000 claims 1
- 239000008274 jelly Substances 0.000 claims 1
- 239000007788 liquid Substances 0.000 claims 1
- 230000003647 oxidation Effects 0.000 claims 1
- 238000007254 oxidation reaction Methods 0.000 claims 1
- 238000000034 method Methods 0.000 abstract description 9
- 230000008569 process Effects 0.000 abstract description 6
- 238000003786 synthesis reaction Methods 0.000 abstract description 4
- 230000015572 biosynthetic process Effects 0.000 abstract description 3
- 238000004519 manufacturing process Methods 0.000 abstract description 3
- 238000011835 investigation Methods 0.000 abstract description 2
- ZPGDWQNBZYOZTI-UHFFFAOYSA-N 1-(9h-fluoren-9-ylmethoxycarbonyl)pyrrolidine-2-carboxylic acid Chemical compound OC(=O)C1CCCN1C(=O)OCC1C2=CC=CC=C2C2=CC=CC=C21 ZPGDWQNBZYOZTI-UHFFFAOYSA-N 0.000 abstract 1
- 230000008961 swelling Effects 0.000 description 8
- 239000012317 TBTU Substances 0.000 description 4
- CLZISMQKJZCZDN-UHFFFAOYSA-N [benzotriazol-1-yloxy(dimethylamino)methylidene]-dimethylazanium Chemical compound C1=CC=C2N(OC(N(C)C)=[N+](C)C)N=NC2=C1 CLZISMQKJZCZDN-UHFFFAOYSA-N 0.000 description 4
- 150000002469 indenes Chemical class 0.000 description 4
- 238000007086 side reaction Methods 0.000 description 4
- UHOVQNZJYSORNB-UHFFFAOYSA-N Benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 description 3
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 3
- 230000009286 beneficial effect Effects 0.000 description 3
- 238000004140 cleaning Methods 0.000 description 3
- 238000005336 cracking Methods 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 229920001184 polypeptide Polymers 0.000 description 3
- 102000004196 processed proteins & peptides Human genes 0.000 description 3
- 230000004044 response Effects 0.000 description 3
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 2
- 239000007983 Tris buffer Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000012634 fragment Substances 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 239000003960 organic solvent Substances 0.000 description 2
- WROMPOXWARCANT-UHFFFAOYSA-N tfa trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F.OC(=O)C(F)(F)F WROMPOXWARCANT-UHFFFAOYSA-N 0.000 description 2
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 2
- XXMYDXUIZKNHDT-QNGWXLTQSA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-3-(1-tritylimidazol-4-yl)propanoic acid Chemical compound C([C@@H](C(=O)O)NC(=O)OCC1C2=CC=CC=C2C2=CC=CC=C21)C(N=C1)=CN1C(C=1C=CC=CC=1)(C=1C=CC=CC=1)C1=CC=CC=C1 XXMYDXUIZKNHDT-QNGWXLTQSA-N 0.000 description 1
- 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 1
- ADOHASQZJSJZBT-SANMLTNESA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-3-[1-[(2-methylpropan-2-yl)oxycarbonyl]indol-3-yl]propanoic acid Chemical compound C12=CC=CC=C2N(C(=O)OC(C)(C)C)C=C1C[C@@H](C(O)=O)NC(=O)OCC1C2=CC=CC=C2C2=CC=CC=C21 ADOHASQZJSJZBT-SANMLTNESA-N 0.000 description 1
- JAUKCFULLJFBFN-VWLOTQADSA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-3-[4-[(2-methylpropan-2-yl)oxy]phenyl]propanoic acid Chemical compound C1=CC(OC(C)(C)C)=CC=C1C[C@@H](C(O)=O)NC(=O)OCC1C2=CC=CC=C2C2=CC=CC=C21 JAUKCFULLJFBFN-VWLOTQADSA-N 0.000 description 1
- CBPJQFCAFFNICX-IBGZPJMESA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-4-methylpentanoic acid Chemical compound C1=CC=C2C(COC(=O)N[C@@H](CC(C)C)C(O)=O)C3=CC=CC=C3C2=C1 CBPJQFCAFFNICX-IBGZPJMESA-N 0.000 description 1
- 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 1
- ZOXJGFHDIHLPTG-UHFFFAOYSA-N Boron Chemical compound [B] ZOXJGFHDIHLPTG-UHFFFAOYSA-N 0.000 description 1
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 1
- 239000007821 HATU Substances 0.000 description 1
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 1
- NANWDEYOYMUHHG-UHFFFAOYSA-N N1(CCCC1)[P] Chemical compound N1(CCCC1)[P] NANWDEYOYMUHHG-UHFFFAOYSA-N 0.000 description 1
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 1
- BLRPTPMANUNPDV-UHFFFAOYSA-N Silane Chemical compound [SiH4] BLRPTPMANUNPDV-UHFFFAOYSA-N 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 229910021529 ammonia Inorganic materials 0.000 description 1
- 239000002585 base Substances 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 229910052796 boron Inorganic materials 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 239000000460 chlorine Substances 0.000 description 1
- 229910052801 chlorine Inorganic materials 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 230000003292 diminished effect Effects 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 125000000031 ethylamino group Chemical group [H]C([H])([H])C([H])([H])N([H])[*] 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 238000007654 immersion Methods 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 125000001449 isopropyl group Chemical group [H]C([H])([H])C([H])(*)C([H])([H])[H] 0.000 description 1
- 230000001294 luteotrophic effect Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 239000000376 reactant Substances 0.000 description 1
- 239000003488 releasing hormone Substances 0.000 description 1
- 229910000077 silane Inorganic materials 0.000 description 1
- 230000007704 transition Effects 0.000 description 1
- ZGYICYBLPGRURT-UHFFFAOYSA-N tri(propan-2-yl)silicon Chemical compound CC(C)[Si](C(C)C)C(C)C ZGYICYBLPGRURT-UHFFFAOYSA-N 0.000 description 1
- 238000010792 warming Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/23—Luteinising hormone-releasing hormone [LHRH]; Related peptides
-
- 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
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- Life Sciences & Earth Sciences (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Endocrinology (AREA)
- Peptides Or Proteins (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
Comprised the following steps the invention provides a kind of synthetic method of Fertirelin：Prepare the Chlorotrityl Resin of resin Fmoc Pro 2；Prepare peptide resin：Pyr‑His(trt)‑Trp(boc)‑Ser(tbu)‑Tyr(tbu)‑Gly‑Leu‑Arg(pbf)‑Pro‑2CTC Resin；Prepare full guard peptide fragments：Pyr‑His(trt)‑Trp(boc)‑Ser(tbu)‑Tyr(tbu)‑Gly‑Leu‑Arg(pbf)‑Pro‑OH；Prepare full guard Fertirelin；Excision side chain obtains Fertirelin；Thick peptide purification.By the further investigation to process route, optimum synthesis route and condition, the synthetic method operation for obtaining is simple, and reaction condition is gentle for the present invention, process warm and, low production cost, high income.
Description
Technical field
The invention belongs to polypeptide drugs preparation field, it is related to a kind of synthetic method of Fertirelin.
Background technology
Fertirelin (Fertirelina) is a kind of luteotropic hormone releasing factor, and polypeptide sequence is Glp-His-
Trp-Ser-Tyr-Gly-Leu-Arg-Pro-NHEt, its structural formula is as follows：
There is severe reaction conditions, high cost, the low defect of yield in the synthetic method of existing Fertirelin.
The content of the invention
A kind of at least one deficiency in order to overcome prior art of the invention, there is provided synthetic method of new Fertirelin,
To realize that reaction condition is gentle, process is simple, low cost, the purpose of high income.
To achieve these goals, the present invention takes following technical proposals to realize：
A kind of synthetic method of Fertirelin, specifically includes following steps：
Step a, prepares resin：Fmoc-Pro-2-Chlorotrityl Resin, with dichloromethane as solvent, add
Fmoc-Pro-OH dissolves and 2- chlorine trityl chloride resin (2-Chlorotrityl Chloride Resin, abbreviation 2-CTC
Resin), organic base DIPEA (DIPEA) is added, reaction obtains Fmoc-Pro-2-Chlorotrityl
The substitution degree 0.3-1.1mmol/g of Resin, wherein 2-Chlorotrityl Chloride Resin.
Step b, prepares peptide resin：Pyr-His(trt)-Trp(boc)-Ser(tbu)-Tyr(tbu)-Gly-Leu-Arg
(pbf)-Pro-2CTC Resin；It is starting with Fmoc-Pro-2-Chlorotrityl Resin, is coupled successively using solid phase method
Amino acid with blocking group, synthesizes peptide resin.
Step c, prepares full guard peptide fragments：Pyr-His(trt)-Trp(boc)-Ser(tbu)-Tyr(tbu)-Gly-
Leu-Arg(pbf)-Pro-OH；The peptide resin that step b is obtained is added and cuts peptide reagent, obtains full guard peptide fragments.
Step d, prepares full guard Fertirelin：The C-terminal of full guard peptide fragments is carried out ethylamine to obtain full guard husband
For Rayleigh.
Step e, excision side chain obtains Fertirelin：Ice cutting reagent low temperature is added to keep in full guard Fertirelin
After 10-20min, 25 DEG C of reaction 2-3h are heated to, obtain Fertirelin crude product, wherein cutting reagent is trifluoroacetic acid (TFA)：Three
Isopropyl base silane (Tris)：Mercaptopropionic acid (MPA)：Phenol：H2O=90:4:3:2:1.
Cracking reaction initial stage in the present invention, moment is highly exothermic, and temperature rises rapidly, and high temperature easily makes polypeptide be denatured simultaneously
Many side reactions are produced, increases the danger of experiment.Present invention selection adds ice cutting reagent and low temperature keeps, steady beneficial to safety
Fixed carrying out is tested, and reduces side reaction.
Cutting reagent considers the structure of Fertirelin in the present invention, promotes the cracking of different blocking groups, captures quilt
The group of cracking, while preventing the side reaction under complex environment.
Step f, thick peptide purification.
Further, in the step a, room temperature reaction 2-4 hours, repeatedly washed after draining, be subsequently adding without water beetle
The active group of Fmoc-Pro-OH is not connected with alcohol closing resin.
Further, in the step a, repeatedly washing, is respectively adopted DMF and washes twice, is then washed twice with DCM, finally uses
Absolute methanol is washed twice, is subsequently added into absolute methanol stirring or concussion reaction 1-3 hours is not connected with Fmoc-Pro- on closing resin
The active group of OH, drains, and obtains Fmoc-Pro-2-Chlorotrityl Resin.
The washing of step a in the present invention, first using the DMF that dissolubility is best, the reaction residue attachment of dissolving previous step
Reactant on resin, but DMF is not volatile, is difficult to pump.
DCM is also good dissolubility, and DMF dissolves each other, and dissolubility is relatively good, volatile also easily to pump.In this step,
The order of DCM and DMF can be exchanged, and influence smaller, it is preferred that DMF is preceding.
Absolute methanol needs to be placed on finally, and its intersolubility is good, and dissolubility can make resin volume big with shrinkage resin
It is big to reduce, also conform to the reaction that next step absolute methanol closes the unnecessary active group of resin.
Wherein absolute methanol can wash and can shrinkage resin, resin volume is diminished saving reagent dosage, but if most
First washed with absolute methanol and be unfavorable for that resin is thoroughly washed, the material of previous step reaction residual is easy when resin volume diminishes
It is wrapped, is unfavorable for thoroughly cleaning.
Small volume, dry resin are adapted to preserve.And DMF and DCM can allow resin swelling, volume to become big.If without
Absolute methanol is washed, and has DMF or DCM residuals when next step is reacted, it is necessary to be closed with more absolute methanol reagents, shunk.
Secondly, the reaction that next step absolute methanol closes the unnecessary active group of resin has been complied with：Closed with absolute methanol
The active group of the not connected proline of chlorine resin, reagent, can be realized preferable transition by existing cleaning function again here.
Preferably, the substitution degree of 2-Chlorotrityl Chloride Resin is 0.4-0.7mmol/ in the step a
g。
The difficulty of the substitution degree amino acid couplings in it can increase organic synthesis too high, increases impurity, extends manufacture cycle, drop
Low yield.The too low synthesis of coupling of substitution degree easily, but prepares the resin and reagent for needing to use substantial amounts of costliness, also extends overall
Experimental period, increased cost, reduce efficiency.Therefore above-mentioned factor, 2- in currently preferred step a are considered
The substitution degree of Chlorotrityl Chloride Resin is 0.4-0.7mmol/g.
Further, the preparation method of peptide resin is in the step b：
(1) resin Fmoc-Pro-2-Chlorotrityl Resin are taken, 20-40min in dichloromethane is immersed in, are drained,
Washed with DMF three times, drained；
(2) reagent of raising one's hat is added, reaction twice, removes Fmoc blocking groups；Washing at least 6 times, washing sequence：Dimethyl
Formamide (DMF), absolute methanol, DMF, dichloromethane (DCM)+DMF, DMF, DMF are drained；With DMF as solvent, Fmoc- is added
The combination of Arg (pbf)-OH and N, N- DIC (DIC) and I-hydroxybenzotriazole (HOBT) or N, N- bis-
The combination of wopropyl ethyl amine (DIPEA) and condensing agent, wherein condensing agent are selected from 2- (7- aoxidizes BTA)-N, N, N', N'-
Tetramethylurea hexafluorophosphoric acid ester (HATU), O- BTAs-N, N, N', N'- tetramethylurea tetrafluoro boric acid (TBTU), hexafluoro phosphorus
Sour BTA -1- bases-epoxide tripyrrole alkyl phosphorus (PYBOP), O- BTAs-N, N, N', N'- tetramethylurea tetrafluoro boron
Mixture, the hexafluorophosphoric acid BTA -1- bases-epoxide tripyrrole alkyl phosphorus of sour (TBTU) and I-hydroxybenzotriazole (HOBT)
(PYBOP) one kind and in the mixture of I-hydroxybenzotriazole (HOBT)；Coupling 1-3 hours, ninhydrin detection reaction process；
Coupling takes out reaction solution after terminating, and is washed with DMF 3 times；(2) above-mentioned steps are repeated until completing last amino acid Jiao's paddy ammonia
Sour PGlu；DMF is washed three times, and DCM is washed three times, and absolute methanol is washed three times, and absolute methanol each 5-10min is dried to obtain after washing
Peptide resin.
In this step, washing at least 6 times, washing sequence why are designed：Dimethylformamide (DMF), absolute methanol,
DMF, dichloromethane (DCM)+DMF, DMF, are that applicant constantly observes in experiment is invented, and are thought deeply, and practice is obtained：
DMF dissolubilities preferably, can dissolve and take away a large amount of reaction residues, therefore first by.
Methyl alcohol can make resin shrinkage, volume diminish, and reduce resin surface attachment, and resin volume reduces and also reduces washing
Required amount of reagent.But form by-product if absolute methanol residual can easily participate in reaction during coupling reaction afterwards
Thing.So can be only placed at used above, it is impossible to washed with below.
Resin wash when one contracting one it is swollen can be issued to more preferable clean result in less reagent dosage, also save examination
Agent.
DMF has appropriate swellability to resin, between methyl alcohol and DCM, opens gentle swelling of resin, is easy to wash
Wash.
DCM has preferable dissolubility and very strong swellability, make resin fully further it is swelling open, be easy to well washing
Resin, the abundant swelling state opened is also beneficial to coupling reaction below.
Resin swelling is more beneficial for carrying out after opening in coupling reaction, so needing DCM again fully swelling to open resin.
Washed with the best DMF of dissolubility below, be further ensured that cleaning is thorough, it is to avoid residue enters influence next step
Reaction produces side reaction.
Further, each step amino acid is 1.3-4 times of resin mole, N, N- DIC in step b
It it is 2.6-24 times of resin mole, condensing agent is 1.3-4 times of resin mole.
Further, reagent of being raised one's hat in step b is piperidines (the DBLK)/DMF of volume fraction 20%.
Further, first set reaction 10min, the second secondary response 5min after reagent of raising one's hat are added in step b.
It is found by the applicant that primary first-order equation can only remove most FMOC blocking groups.Need to add new examination of raising one's hat again
Agent, secondary response reaches relatively complete removal degree.And once add excessive reagent of raising one's hat also be difficult to reach it is relatively complete
Removal degree, removal effect is optimal after secondary response.
Further, in step c, peptide resin that step b is obtained add cut the trifluoroacetic acid of peptide reagent volume fraction 2%/
DCM mixed solutions；Peptide resin is added at -20 to 0 DEG C, concussion/stirring keeps low temperature 10-20min；25 DEG C are then heated to, instead
Answer 2-3 hours；Suction filtration, collects filtrate, plus organic base adjusts pH to neutrality, and vacuum distillation obtains full guard peptide fragments.
Further, in step d, full guard peptide fragments are dissolved in DMF, plus hexafluorophosphoric acid BTA -1- bases-epoxide three
Pyrrolidinyl phosphorus and ethylamine hydrochloride；It is 8-9, stirring reaction 2-6 hours to add organic base regulation pH；Added after ethylamine end
The saturated lemon aqueous solution separates out white solid, drains, and dries, and obtains full guard Fertirelin, the wherein addition of ethylamine hydrochloride
Measure is 1.2-2 times of full guard peptide fragments mole.
Further, it is in step f that Fertirelin crude product is soluble in water, filtered with 0.45 μm of micro-porous filtration plate；Filtrate passes through
C18 prepares the preparation of post loading, and wash-out collects target peak peptide solution；High-purity peptide solution turns salt by reversed-phase column, into turning acetate
Form；Solution decompression concentrated by rotary evaporation, it is powdered Fertirelin that concentrate is dried by deep freeze refrigerator.
The all methyl alcohol wherein used in the present invention are absolute methanol.
The present invention has the following technical effect that：
The present invention is by the further investigation to process route, optimum synthesis route and condition, the synthetic method operation for obtaining
Simply, reaction condition is gentle, process warm and, low production cost, high income.
It is that above and other objects of the present invention, feature and advantage can be become apparent, preferred embodiment cited below particularly,
It is described in detail below.
Specific embodiment
Embodiment 1
The synthetic method of Fertirelin：
1. resin processed：Fmoc-Pro-2-Chlorotrityl Resin.
50ml organic solvent DCM are added in reactor, 4.5g Fmoc-Pro-OH dissolvings are added, 10g 2- are added
Chlorotrityl Chloride Resin (2-CTC Resin), immersion, stirring or concussion make it fully swelling；Add 10ml
Organic base DIPEA (DIPEA) reacts 3 hours at room temperature, takes out.
Washing：DMF*2+DCM*2+ methyl alcohol * 2 is respectively adopted to be washed, adds the stirring of 60ml methyl alcohol or concussion reaction 2 small
When closing resin on be not connected with Fmoc-Pro-OH active group.Drain, obtain Fmoc-Pro-2-Chlorotrityl
Resin。
Ultraviolet method detection resin substitution degree 0.6mmol/g.
2. peptide resin is prepared：Pyr-His(trt)-Trp(boc)-Ser(tbu)-Tyr(tbu)-Gly-Leu-Arg
(pbf)-Pro-2CTC Resin。
It is starting with the resin Fmoc-Pro-2-Chlorotrityl Resin for connecting.
(1) 10g resin Fmoc-Pro-2-Chlorotrityl Resin are taken, is immersed in 100ml organic solvents DCM
30min, being aided with stirring makes it fully swelling.Drain, washed with DMF three times, take out.
(2) 100ml is added to raise one's hat reagent：20%DBLK+DMF, reacts twice, first time 10min, second 5min respectively
To remove Fmoc blocking groups.It is washed out 6 times, specific washing sequence：DMF+ methyl alcohol+DMF+ (DCM+DMF)+DMF+DMF, takes out
It is dry.With DMF as solvent, 9.7g Fmoc-Arg (pbf)-OH, 10ml DIPEA and TBTU (4.8g)+HOBT (2g), normal temperature are added
Lower coupling 2 hours, ninhydrin detection reaction process, coupling is taken out reaction solution, is washed with DMF 3 times after terminating.
(3) 100ml is added to raise one's hat reagent：20%DBLK+DMF, reacts twice, first time 10min, second 5min respectively
To remove Fmoc blocking groups.It is washed out 6 times, specific washing sequence：DMF+ methyl alcohol+DMF+ (DCM+DMF)+DMF+DMF, takes out
It is dry.With DMF as solvent, coupling under 5.3g Fmoc-Leu-OH, 10ml DIPEA and TBTU (4.8g)+HOBT (2g) normal temperature is added
2 hours, ninhydrin detection reaction process, coupling was taken out reaction solution, is washed with DMF 3 times after terminating.
(4) 100ml is added to raise one's hat reagent：20%DBLK+DMF, reacts twice, first time 10min, second 5min respectively
To remove Fmoc blocking groups.It is washed out 6 times, specific washing sequence：DMF+ methyl alcohol+DMF+ (DCM+DMF)+DMF+DMF, takes out
It is dry.With DMF as solvent, 4.5g Fmoc-Gly-OH, 5ml DIC and 2g HOBT are added, coupling 2 hours under 29 degree, ninhydrin inspection
Reaction process is surveyed, coupling is taken out reaction solution, washed with DMF 3 times after terminating.
(5) 100ml is added to raise one's hat reagent：20%DBLK+DMF, reacts twice, first time 10min, second 5min respectively
To remove Fmoc blocking groups.It is washed out 6 times, specific washing sequence：DMF+ methyl alcohol+DMF+ (DCM+DMF)+DMF+DMF, takes out
It is dry.With DMF as solvent, 6.9g Fmoc-Tyr (tbu)-OH, 5ml DIC and 2g HOBT are added, coupling 2 hours, indenes under 29 degree
Triketone detects that reaction process, coupling take out reaction solution after terminating, and is washed with DMF 3 times.
(6) 100ml is added to raise one's hat reagent：20%DBLK+DMF, reacts twice, first time 10min, second 5min respectively
To remove Fmoc blocking groups.It is washed out 6 times, specific washing sequence：DMF+ methyl alcohol+DMF+ (DCM+DMF)+DMF+DMF, takes out
It is dry.With DMF as solvent, 5.7g Fmoc-Ser (tbu)-OH, 5ml DIC and 2g HOBT are added, coupling 2 hours, indenes under 29 degree
Triketone detects that reaction process, coupling take out reaction solution after terminating, and is washed with DMF 3 times.
(7) 120ml is added to raise one's hat reagent：20%DBLK+DMF, reacts twice, first time 10min, second 5min respectively
To remove Fmoc blocking groups.It is washed out 6 times, specific washing sequence：DMF+ methyl alcohol+DMF+ (DCM+DMF)+DMF+DMF, takes out
It is dry.With DMF as solvent, 7.9g Fmoc-Trp (boc)-OH, 5ml DIC and 2g HOBT are added, coupling 2 hours, indenes under 29 degree
Triketone detects that reaction process, coupling take out reaction solution after terminating, and is washed with DMF 3 times.
(8) 120ml is added to raise one's hat reagent：20%DBLK+DMF, reacts twice, first time 10min, second 5min respectively
To remove Fmoc blocking groups.It is washed out 6 times, specific washing sequence：DMF+ methyl alcohol+DMF+ (DCM+DMF)+DMF+DMF, takes out
It is dry.With DMF as solvent, 9.3g Fmoc-His (trt)-OH, 5ml DIC and 2g HOBT are added, coupling 2 hours, indenes under 29 degree
Triketone detects that reaction process, coupling take out reaction solution after terminating, and is washed with DMF 3 times.
(9) 120ml is added to raise one's hat reagent：20%DBLK+DMF, reacts twice, first time 10min, second 5min respectively
To remove Fmoc blocking groups.It is washed out 6 times, specific washing sequence：DMF+ methyl alcohol+DMF+ (DCM+DMF)+DMF+DMF, takes out
It is dry.With DMF as solvent, 2g H-Pyr-OH, 5ml DIC and 2g HOBT are added, coupling 2 hours under 29 degree, ninhydrin detection is anti-
Process, coupling is answered to take out reaction solution after terminating.Washed with DMF 3 times, DCM is washed 3 times, methyl alcohol is washed 3 times, the wherein each 8min of methyl alcohol,
Complete washing and resin shrinkage.It is dried to obtain peptide resin.
3. full guard peptide fragments are prepared：Pyr-His(trt)-Trp(boc)-Ser(tbu)-Tyr(tbu)-Gly-Leu-
Arg(pbf)-Pro-OH。
Trifluoroacetic acid (TFA) DCM mixed solutions of configuration 2%.Peptide resin is added at -10 DEG C, is shaken, keep low temperature
15min.25 degree are warming up to, are reacted 2.5 hours.Suction filtration, collects filtrate, plus a small amount of organic base：N, N- diisopropylethylamine are reconciled
To neutrality, vacuum rotary steam obtains full guard fragment PGluP-9 to pH.
4. full guard Fertirelin is prepared.
It is dissolved in full guard fragment PGluP-9 is obtained in 3 in appropriate DMF, adds PYBOP and ethylamine hydrochloride.Addition has
Machine alkali:N, N- diisopropylethylamine adjust pH=8.5.5 hours .HPLC tracking reaction process of stirring reaction.After ethylamine end
Add the saturated lemon aqueous solution to separate out white solid, drain, dry, obtain full guard Fertirelin.
5. excision side chain obtains Fertirelin crude product.
Ice cutting reagent (trifluoroacetic acid TFA is added in glass container：Tri isopropyl silane Tris：Mercaptopropionic acid mpa：Benzene
Phenol：H2O=90：4:3:2:1) low temperature keeps 15min.It is heated to 25 degree or so reaction 2.5h.After reaction terminates.Add ice anhydrous
Ether is precipitated, and is centrifuged/is collected by filtration precipitation, then is washed 5 times with normal temperature absolute ether, is vacuum dried, and obtains Fertirelin crude product.
HPLC detects purity.Weigh 6.94g.
6. thick peptide purification
6.94g Fertirelin crude products is soluble in water, filtered with 0.45 μm of micro-porous filtration plate.Filtrate prepares post by C18
Prepared by loading, wash-out, collects target peak peptide solution.High-purity peptide solution turns salt into acetate form.Solution decompression concentrated by rotary evaporation,
It is powdered Fertirelin 2.19g, purity 99.6%, yield 31.6% that concentrate is dried by deep freeze refrigerator.
Although the present invention is disclosed as above with preferred embodiment, it is not for limiting the present invention, any this area
Technical staff without departing from the spirit and scope of the present invention, may be by the methods and techniques content of the disclosure above to this hair
Bright technical scheme makes possible variation and modification, therefore, every content without departing from technical solution of the present invention, according to the present invention
Technical spirit to any simple modification, equivalent variation and modification made for any of the above embodiments, belong to technical solution of the present invention
Protection.
Claims (10)
1. a kind of synthetic method of Fertirelin, it is characterised in that the synthetic method specifically includes following steps：
Step a, prepares resin：Fmoc-Pro-2-Chlorotrityl Resin, with dichloromethane as solvent, add Fmoc-
Pro-OH dissolves and 2-Chlorotrityl Chloride Resin, adds organic base DIPEA (DIPEA),
Reaction obtains Fmoc-Pro-2-Chlorotrityl Resin, the wherein replacement of 2-Chlorotrityl Chloride Resin
Degree 0.3-1.1mmol/g；
Step b, prepares peptide resin：Pyr-His(trt)-Trp(boc)-Ser(tbu)-Tyr(tbu)-Gly-Leu-Arg
(pbf)-Pro-2CTC Resin；It is starting with Fmoc-Pro-2-Chlorotrityl Resin, using solid-phase synthesis one by one
Amino acid of the coupling with blocking group, synthesizes peptide resin；
Step c, prepares full guard peptide fragments：Pyr-His(trt)-Trp(boc)-Ser(tbu)-Tyr(tbu)-Gly-Leu-
Arg(pbf)-Pro-OH；The peptide resin that step b is obtained is added and cuts peptide reagent, obtains full guard peptide fragments；
Step d, prepares full guard Fertirelin：The C-terminal of full guard peptide fragments is carried out ethylamine to obtain full guard husband for auspicious
Woods；
Step e, excision side chain obtains Fertirelin：Ice cutting reagent low temperature is added to keep 10- in full guard Fertirelin
After 20min, 25 DEG C of reaction 2-3h are heated to, obtain Fertirelin crude product, wherein cutting reagent is trifluoroacetic acid：Triisopropyl silicon
Alkane：Mercaptopropionic acid：Phenol：H2O=90:4:3:2:1；
Step f, thick peptide purification.
2. the synthetic method of Fertirelin according to claim 1, it is characterised in that in the step a, room temperature reaction 2-
4 hours, repeatedly washed after draining, be subsequently adding on absolute methanol closing resin and be not connected with the active group of Fmoc-Pro-OH
Group.
3. the synthetic method of Fertirelin according to claim 2, it is characterised in that in the step a, repeatedly washing,
DMF is respectively adopted to wash twice, is then washed twice with DCM, finally washed twice with absolute methanol, be subsequently added into absolute methanol stirring or
The active group for Fmoc-Pro-OH being not connected with closing resin in concussion reaction 1-3 hours；Drain, obtain Fmoc-Pro-2-
Chlorotrityl Resin。
4. the synthetic method of Fertirelin according to claim 1, it is characterised in that 2- in the step a
The substitution degree of Chlorotrityl Chloride Resin is 0.4-0.7mmol/g.
5. the synthetic method of Fertirelin according to claim 1, it is characterised in that the system of peptide resin in the step b
Preparation Method is：
(1) resin Fmoc-Pro-2-Chlorotrityl Resin are taken, 20-40min in dichloromethane is immersed in, is drained, used
DMF is washed three times, is drained；
(2) reagent of raising one's hat is added, reaction twice, removes Fmoc blocking groups；Washing at least 6 times, washing sequence：DMF, without water beetle
Alcohol, DMF, DCM+DMF, DMF, DMF are drained；With DMF as solvent, Fmoc-Arg (pbf)-OH, and I-hydroxybenzotriazole are added
And the combination of N, N- DIC or DIPEA and condensing agent combination, wherein condensing agent is selected from 2- (7-
Oxidation BTA)-N, N, N', N'- tetramethylurea hexafluorophosphoric acid ester, O- BTAs-N, N, N', N'- tetramethylurea
Tetrafluoro boric acid, hexafluorophosphoric acid BTA -1- bases-epoxide tripyrrole alkyl phosphorus, O- BTAs-N, N, N', N'- tetramethyl
Urea tetrafluoro boric acid and I-hydroxybenzotriazole mixture, hexafluorophosphoric acid BTA -1- bases-epoxide tripyrrole alkyl phosphorus and 1- hydroxyls
One kind in base BTA mixture；Coupling 1-3 hours, ninhydrin detection reaction process, coupling takes out reaction solution after terminating,
Washed with DMF 3 times；(2) above-mentioned steps are repeated until completing last amino acid pyroglutamic acid PGlu；DMF washes three times, DCM
Wash three times, absolute methanol is washed three times, absolute methanol each 5-10min is dried to obtain peptide resin after washing.
6. the synthetic method of Fertirelin according to claim 5, it is characterised in that each step amino acid is in step b
1.3-4 times of resin mole, DIPEA is 2.6-24 times of resin mole, and condensing agent is resin mole
1.3-4 times.
7. the synthetic method of Fertirelin according to claim 5, it is characterised in that reagent of being raised one's hat in step b is volume
Hexahydro piperidines/the dimethylformamide of fraction 20%；First set reaction 10min after reagent of raising one's hat is added in step b, second anti-
Answer 5min.
8. the synthetic method of Fertirelin according to claim 1, it is characterised in that in step c, step b is obtained
Peptide resin adds the trifluoroacetic acid/DCM mixed solutions for cutting peptide reagent volume fraction 2%；Peptide resin is added at -20 to 0 DEG C, is shaken
Swing/stir, keep low temperature 10-20min；25 DEG C are then heated to, is reacted 2-3 hours；Suction filtration, collects filtrate, plus organic base is adjusted
PH to neutrality is saved, vacuum distillation obtains full guard peptide fragments.
9. the synthetic method of Fertirelin according to claim 1, it is characterised in that in step d, full guard peptide fragments
It is dissolved in DMF, plus hexafluorophosphoric acid BTA -1- bases-epoxide tripyrrole alkyl phosphorus and ethylamine hydrochloride；Add organic base regulation
PH is 8-9, stirring reaction 2-6 hours；Add the saturated lemon aqueous solution to separate out white solid after ethylamine end, drain, do
It is dry, full guard Fertirelin is obtained, the wherein addition of ethylamine hydrochloride is 1.2-2 times of full guard peptide fragments mole.
10. the synthetic method of Fertirelin according to claim 1, it is characterised in that by Fertirelin crude product in step f
It is soluble in water, filtered with 0.45 μm of micro-porous filtration plate；Filtrate prepares post loading and prepares by C18, and wash-out collects target peak peptide molten
Liquid；High-purity peptide solution turns salt by reversed-phase column, into acetate form；Solution decompression concentrated by rotary evaporation, concentrate passes through low temperature cold
It is powdered Fertirelin that jelly machine is dried.
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