Trifluoroacetic acid → Diphenyl n-(6-biotinamidohexanoyl)amino(4-amidinophenyl)methanephosphonate

HRID 1470060

Reaktionsgleichung

Reaktionsgleichung von HRID 1470060
[NH2:1][CH:2]([C:19]1[CH:24]=[CH:23][C:22]([C:25](=[NH:27])[NH2:26])=[CH:21][CH:20]=1)[P:3]([O:12][C:13]1[CH:18]=[CH:17][CH:16]=[CH:15][CH:14]=1)(=[O:11])[O:4][C:5]1[CH:10]=[CH:9][CH:8]=[CH:7][CH:6]=1.[C:28]([NH:43][CH2:44][CH2:45][CH2:46][CH2:47][CH2:48][C:49](O)=[O:50])(=[O:42])[CH2:29][CH2:30][CH2:31][CH2:32][C@H:33]1[C@@H:41]2[C@@H:36]([NH:37][C:38]([NH:40]2)=[O:39])[CH2:35][S:34]1.C1CN([P+](ON2N=NC3C=CC=CC2=3)(N2CCCC2)N2CCCC2)CC1.F[P-](F)(F)(F)(F)F.C(O)(C(F)(F)F)=O.C(ON1C(=O)CCC1=O)(=O)CCCCCCC(ON1C(=O)CCC1=O)=O.OC(CCCC[C@H]1[C@@H]2[C@@H](NC(N2)=O)CS1)=O>O.C(#N)C.CN(C=O)C.ClCCl>[C:28]([NH:43][CH2:44][CH2:45][CH2:46][CH2:47][CH2:48][C:49]([NH:1][CH:2]([C:19]1[CH:24]=[CH:23][C:22]([C:25](=[NH:26])[NH2:27])=[CH:21][CH:20]=1)[P:3]([O:4][C:5]1[CH:10]=[CH:9][CH:8]=[CH:7][CH:6]=1)(=[O:11])[O:12][C:13]1[CH:18]=[CH:17][CH:16]=[CH:15][CH:14]=1)=[O:50])(=[O:42])[CH2:29][CH2:30][CH2:31][CH2:32][C@H:33]1[C@@H:41]2[C@@H:36]([NH:37][C:38]([NH:40]2)=[O:39])[CH2:35][S:34]1

Komponenten

11

Edukt 6

HCID 6422Trifluoroacetic acid3×HCID 100658N-Hydroxysuccinimide suberic acid esterHCID 171548BiotinHCID 4469056-[5-(2-Oxo-Hexahydro-Thieno[3,4-D]Imidazol-4-Yl)-Pentanoylamino]-Hexanoic AcidHCID 2724699PyBOPHCID 10407461Diphenyl amino(4-amidinophenyl)methanephosphonate

Produkt 1

HCID 87648867Diphenyl n-(6-biotinamidohexanoyl)amino(4-amidinophenyl)methanephosphonate

Reagenzien, Katalysatoren, Lösungsmittel 4

HCID 962WaterLösungsmittelHCID 6228DimethylformamideLösungsmittelHCID 6342AcetonitrileLösungsmittelHCID 6344DichloromethaneLösungsmittel

Versuchsvorschrift

CALs. Diphenyl N-(6-biotinamidohexanoyl)amino(4-amidinophenyl)methanephosphonate (1) was prepared from diphenyl amino(4-amidinophenyl)methanephosphonate (19,20) and 6-biotinamidohexanoic acid (Anaspec; San Jose, Calif.) by the aid of PyBOP (Novabiochem; San Diego, Calif.). The HPLC-purified material [retension time 20.76 min, purity 95% (220 nm); YMC ODS-AM column (4.6×250 mm), 0.05% TFA in water (A):0.05% TFA in acetonitrile (B) 90:10 to 20:80 in 45 min (1.0 ml/min)] was characterized by ESI-MS [Observed m/z 721.3 (MH+; calculated MH+ for C36H45N6O6PS, 721.3)] and stored as 10 mM solution in DMF at −70° C. The active ester 2 was prepared by acylating the same precursor amine with disuccinimidyl suberate (Pierce; Rockford, Ill.) and characterized in the same way [Observed m/z, 635.3 (MH+; calculated MH+ for C32H35N4O8P, 635.2)]. VIP-CAL (3) was synthesized as follows. The VIP sequence with N-terminal biotin was constructed on Rink amide MBHA resin (0.72 mmol/g; Novabiochem) by the standard 9-fluorenylmethoxycarbonyl protocol (21) except that 4-methyltrityl (22) was used for side-chain protection of Lys20. The peptide-resin was treated with 1% TFA in dichloromethane (5 min×10) to remove the 4-methyltrityl group and the deprotected amino group of Lys20 was acylated with 2 in 1-methyl-2-pyrrolidinone containing 0.1 mM N,N-diisopropylethylamine. The peptide resin was treated with TFA-ethanedithiol-thioanisole-phenol (90:1:1:8) at room temperature for 2 h. After removing the resin by filtration, diethyl ether was added to the solution to afford a precipitate, which was collected by centrifugation and washed with diethyl ether. The HPLC-purified material [retention time 50.25 min, purity 96% (220 nm); Vydac 214TP C4 column (4.6×250 mm); A:B 90:10 to 60:40 in 60 min (1.0 ml/min)] was characterized by ESI-MS [Observed m/z, 4071.4 (MH+; calculated MH+ for C185H282N49O49PS2, 4072.0) and stored as 10 mM solution in DMSO at −70° C.

Aufarbeitung

  1. customThe active ester 2 was prepared
  2. customto remove the 4-methyltrityl group
  3. additionThe peptide resin was treated with TFA-ethanedithiol-thioanisole-phenol (90:1:1:8) at room temperature for 2 h
  4. customAfter removing the resin
  5. filtrationby filtration, diethyl ether
  6. additionwas added to the solution
  7. customto afford a precipitate, which
  8. customwas collected by centrifugation
  9. washwashed with diethyl ether
  10. customstored as 10 mM solution in DMSO at −70° C.

Quellen und Nachweise

ORD-Eintrag
ord-fddc5bfff4ab4fa3a404fb3c2e54ad7d
Quelldatensatz
uspto-grants-2015_04
Patentnummer
US09006388B2

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