(HMPB), and photolinkers were used as handles.27 A recent
report on the synthesis of peptides containing N-methyl amino
acids is the N-methylation of NR-acylated linear peptides and
NR-aryl sulfonyl peptides on solid support by using CH3I/Ag2O
in DMF28 and dimethylsulfate and DBU as base,29 respectively.
SCHEME 1. Synthesis of CsO by Step-by-Step Assembly of
Amino Acids
Cyclosporin O (CsO), one of the important members of the
Cs family that occurs naturally contains MeLeu and norvaline
(Nva) at positions 1 and 2 in place of (4R)-[4(E)-2-butenyl]-
4,N-dimethyl-L-threonine (MeBmt) and (2s)-2-aminobutanoic
acid (Abu), respectively. Cs compounds bearing Nva such as
CsG are reported to be less nephrotoxic than the parent CsA.30
Hence, development of an efficient method for the synthesis of
CsO becomes imminent. The synthesis of CsO both in solution
as well as by solid-phase method was performed by a combina-
tion of reagents, 2-bromo-3-ethyl-4-methyl thiazolium tetrafluo-
roborate (BEMT), 2-bromo-1-ethyl pyridinium tetrafluoroborate
(BEP), and 5-(1H-benzotriazol-1-yloxy)-3,4-dihydro-1-methyl
2H-pyrrolium hexachloroantimonate (BDMP) in the presence
of 1-hydroxy-7-azabenzotriazole (HOAt).31-33 Another success-
ful synthesis of CsO was accomplished in solution phase via 4
+ 7 fragment coupling and the cyclization of final undecapeptide
between L-Ala and D-Ala using 1-(1-pyrrolidinyl-1H -1,2,3-
triazolo[4,5-b]pyridin-1-ylmethylene)pyrrolidinium hexafluo-
rophosphate N-oxide (HAPyU). The SPPS of CsO using
bis(trichloromethyl)carbonate (BTC)/collidine and acid labile
trityl chloride polystyrene resin was reported.34,35 The couplings
were performed by in situ generation of acid chloride in the
presence of collidine and were found to be complete in 3 h.
This was an extension of the Falb et al.36 method for the
formation of N-alkyl amides on Rink amide resin. In another
report the CsO fragment, Boc-MeLeu-MeLeu-MeVal-MeLeu-
OMe was synthesized by treating its unmethylated precursor
with excess MeI and Ag2O. The fragment was further utilized
in the synthesis of CsO.37 To the best of our knowledge, most
of the earlier reports on the synthesis of Cs were either on solid
phase or by fragment coupling in solution phase. In this report
we describe the utility of acid chloride/zinc dust for the synthesis
of CsO by step-by-step assembly of amino acids using Fmoc
chemistry.
complete elimination of base-catalyzed side reactions, namely,
the formation of oxazolone and racemization. The coupling
proceeds under neutral conditions and thus can be carried out
over an extended period of time. This advantage offered by
amino acid chlorides becomes much more conspicuous for the
coupling of N-methyl amino acids. Following the reports of
difficulties encountered in the synthesis of Cs, we choose to
exploit the utility of zinc dust for the coupling of hindered
N-methyl amino acids. The overall strategy involved in the
synthesis of CsO is illustrated in Scheme 1.
The synthesis of all Fmoc-NMe-amino acid chlorides except
Fmoc-MeVal-Cl was accomplished by the SOCl2/DCM method.
The use of regular protocol for the synthesis of Fmoc-MeVal-
Cl was not satisfactory. Alternatively, an in situ generated acid
chloride was used. A suspension of Fmoc-MeVal-OH (1 mmol)
(40) For the synthesis of peptides employing Fmoc-amino acid chlorides
mediated by potassium salt of 1-hydroxy benzotriazole (KOBt) in solution
and solid phase synthesis see the chapter on acid chlorides: Beyermann,
M.; Bienert, M.; Carpino, L. A. Acid chlorides. Houben-Weyl; Thieme:
Stuttgart, Germany, 2003; Vol. E 22c, p 475 and references cited therein.
(41) For a report on the synthesis of MeLeu-Val-MeLeu-Ala-OBzl and
D- Ala-MeLeu-MeLeu-MeVal-OtBu fragments of Cs employing Fmoc-
amino acid chlorides mediated by KOBt, see: Sivanandaiah, K. M.;
Sureshbabu, V. V.; Shankaramma, S. C. Int. J. Pept. Protein Res. 1994,
44, 24.
(42) For the utility of TBDMS-OBt in racemization free coupling of
Fmoc-amino acid chlorides in solution phase see: (a) Tantry, S. J.;
Sureshbabu, V. V. Lett. Pept. Sci. 2002, 9, 35. (b) Tantry, S. J.; Mathad, R.
I.; Sureshbabu, V. V. Ind. J. Chem. 2003, 42B, 2104.
Acid chlorides, one of the powerful modes of activation of
carboxylic acids, preparation is simple as well as less expensive.
Their utility in the synthesis of peptides possessing dialkyl amino
acids is known.38 When acid chlorides are employed as acylating
agents the coupling requires a base for the abstraction of
liberated HCl. Instead, zinc dust39-45 can be used for this
purpose under non-Schotten-Baumann conditions in homoge-
neous medium employing DCM or CHCl3 resulting in the
(27) Raman, P.; Stokes, S. S.; Angell, Y. M.; Flentke, G. R.; Rich, D.
H. J. Org. Chem.1998, 63, 5734.
(28) Biron, E.; Chatterjee, J.; Kessler, H. J. Pept. Sci. 2006, 12, 213.
(29) Moretto, A.; Crisma, M.; Kaptein, B.; Broxterman, Q. B.; Toniolo,
C. Biopolymers 2006, 84, 553.
(43) For other applications of nonbasic conditions in the structure
determination of HBTU and HATU, see: Carpino, L. A.; Imazumi, H.;
El-Faham, A.; Ferrer, F. J.; Zhang, C.; Lee, Y.; Foxman, B. M.; Henklein,
P.; Hanay, C.; Mugge, C. Wenschuh, H.; Klose, J.; M. Beyermann.; M.
Bienert. Angew. Chem., Int. Ed. 2002, 41, 442.
(30) Hiestand, P. C.; Gunn, H. C.; Gale, J. M.; Ryffel, B.; Borel, J. F.
Immunology 1985, 55, 249.
(31) Li, P.; Xu, C. J. Org. Chem. 2000, 65, 2951.
(32) Li, P.; Xu, C. Tetrahedron Lett. 1999, 40, 8301.
(33) Li, P.; Xu, C. Tetrahedron 2000, 56, 9949.
(34) Thern, B.; Rudolph, J.; Jung, G. Tetrahedron Lett. 2002, 43, 5013.
(35) Sewald, N. Angew. Chem., Int. Ed. 2002, 41, 4661.
(36) Falb, E.; Yechezkel, T.; Salitra, Y.; Gilon, C. Int. J. Pept. Protein
Res. 1999, 53, 507.
(37) Keiichi, Y.; Teruya, E.; Hiroyuki, Y.; Ryoichi, K. Pept. Sci. 2004,
41, 591.
(44) For an application of AgCN in the coupling of Fmoc-amino acid
chlorides in the synthesis of cyclic hexapeptide oxytocin agonists see:
Williams, P. D.; Bock, M. G.; Tung, R. D.; Garsky, V. M.; Perlow, D. S.;
Erb, J. M.; Lundell, G. F.; Gould, N. P.; Whitter, W. L.; Hoffman, J. B.;
Kaufman, M. J.; Clineschmidt, B. V.; Pettibone, D. J.; Freidinger, R. M.;
Veber, D. F. J. Med. Chem. 1992, 35, 3905.
(45) For a study concerning the coupling of Fmoc-amino acid chlorides
in the presence of AgCN, see: (a) Sureshbabu, V. V.; Gayathri, K. Ind. J.
Chem. 1998, 37B, 1109. (b) As analyzed by HPLC using chiral stationary
phase (csp, chiralcel OD), the coupling of (S) and (R)-Fmoc-Ser(tBu)-Cl
with aniline mediated by AgCN was found to be racemization free (less
than 0.5%): Spivey, A. C.; McKendrick, J.; Srikaran, R.; Helm, B. A. J.
Org. Chem. 2003, 68, 1843.
(38) Sureshbabu, V. V.; Gopi, H. N. Tetrahedron Lett. 1998, 39, 1049.
(39) Gopi, H. N.; Sureshbabu, V. V. Tetrahedron Lett. 1998, 39, 9769.
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