Preloaded Resins for Synthesis of Peptide Amides

Published on 20/06/2014

Used for Efficient and Economic Bulk Synthesis of Peptide APIs:

A new technology is available with a set of new preloaded resins, which enable the synthesis of peptide amides WITHOUT ANY LINKER.
Amino acid amides, which bear a functional side chain, are immobilized on acid sensitive resins. This makes process development, small scale synthesis, bulk production and in-process control very easy.

Find more technical details in our application for demonstration purposes with BIVALIRUDIN.

Used for efficient and economic synthesis of bulk peptide APIs

The Technology:

Side chain attachment of amino acids and peptides on highly acid sensitive resins.

The Advantages:

The synthesis of peptide amides happens WITHOUT typical amid linkers (Rink, Ramage, Sieber). Therefore no side reactions and for- mation of impurities through linker typical fragmentation during TFA cleavage can occur. Protected peptide acids & amides will be obtained in high yield and purity.
Cleavage from the resins is carried out with low concentration of TFA; acid sensitive protecting groups used in standard Fmoc/tBu strategy (tBu, Boc, Pbf, Trt, etc.) stay on the peptide.
Any intermediate can be cleaved as protected peptide fragment, enabling a very easy in process control through HPLC.
This is especially useful during the optimization of any large scale synthesis!

This concept can be extended to any peptide acid & amide with side chain functions.

The synthesis of head-tail cyclic peptides becomes very easy with the use of side chain immobilized aspartic and glutamic acids together with lysine or ornithine derivatives protected with highly acid sensitive groups, like trityl (Trt), p-methyltrityl (Mtt) and p-methoxy-trityl (Mmt).

K. Barlos et al. WO 2013/098802 A2.
K. Barlos, D. Gatos et al., Int. J. Peptide and Protein Res. 1991; 38: 555-561.
K. Barlos and D. Gatos, in Fmoc solid phase peptide synthesis, edit. W.C. Chan and P. D. White, Oxford University Press, 2000.
Attachment of Histidine, Histamine and Urocanic acid to Resins of the Trityl-Type. Eleftheriou, S.; Gatos, D.; Panagopoulos, A.; Stathopoulos, S.; Barlos, K.; Tetrahed- ron Letters 1999; 40(14): 2825- 2828.
G. Sabatino et al. Tetrahedron Letters 1999; 40: 809-812.
E. Atherton et al. US 7,691,968 B2.
F. Albericio et al. WO 2008/040536 A1.

Easy Process Development of the Synthesis of Bivalirudin by the Laboratories of CBL-Patras

Bivalirudine can be synthesized in bulk quantities in high purity and yield with fragment condensation techniques:

As first step fragment (11-20) will be synthesized by standard SPPS Fmoc/tBu technology on the resin, starting with Tyr¹⁹–Leu²⁰-OtBu immobilized through the side chain of Tyr¹⁹ to the highly acid sensitive 2-chlorotrity resin.
The N-terminal Boc and side chain protected fragment (1-10) will be synthesized also via standard solid phase technology on acid sensitive resin, for example on 2-chlorotrityl resins or 4-methoxybenzhydryl resin. Sequence of Bivalirudin: D-Phe¹-Pro2-Arg³-Pro⁴-Gly⁵-Gly⁶-Gly⁷-Gly⁸-Asn⁹-Gly¹⁰-Asp¹¹-Phe¹²-Glu¹³-Glu¹⁴-Ile¹⁵-Pro¹⁶-Glu¹⁷-Glu¹⁸-Tyr¹⁹-Leu²⁰
In a last step the two fragments will be coupled with DIC/HOBt in NMP. When a ratio of 1:1 of both fragments is being applied a significant part has not been reacted to the complete sequence. This easily can be determined by taking out a sample of the reactor, applying cleavage with low concentration of TFA in DCM and injection into HPLC (pic #1). Adding additional 25% of fragment (1-10) and appropriate reaction time yields complete coupling (pic #2).