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Rapid synthesis of short peptide segments via the FMOC-amino acid chloride solution technique (FAACST method)
Abstract
At the time the present work was started, a new technique, the so-called FMOC/4-AMP method for rapid continuous solution synthesis of peptides, involving coupling by crystalline FMOC-N-$\alpha$-protected amino acid chlorides and deblocking via 4-(aminomethyl)piperidine, had been described. However methods for the synthesis of FMOC-amino acids and the corresponding acid chlorides, especially those bearing t-butyl-based side chain protecting groups were not wholly satisfactory and in addition there was some uncertainty regarding the actual structures of the acid chlorides. Analytical methods for assaying the purity of the acid chlorides were not quantitative nor were convenient and accurate racemization tests available. Finally in the case of certain sequences, separation of the growing peptide from the 4-AMP/DBF adduct was complicated by formation of emulsions or separation of voluminous precipitates during buffer extraction. We have now examined all of these questions and further improved the standard protocols for the use of FMOC-amino acid chlorides in rapid syntheses of short peptide segments. Most simple and side-chain protected FMOC-amino acids were synthesized via FMOC-N$\sb3$ or FMOC-OSu as acylating agents. Stable acid chlorides could not be obtained from trifunctional amino acids bearing t-butyl-derived protecting groups. Remarkably, the corresponding FMOC amino acid fluorides proved to be quite stable and were obtained readily in crystalline form. Most of the common amino acids were converted to such fluoride derivatives and these new compounds were shown to be excellent acylating agents for use in the FAACST synthesis. As a racemization test, a new method was developed based on the separability of the D-L, and L-L diastereomers of FMOC-Phe-Leu-OMe by HPLC on a silica gel column using 1% isopropanol in hexane. For structural studies on the acid halides the combined use of IR and $\sp{13}$C NMR spectroscopy confirmed the expected structures. Finally, substitution of tris(2-aminoethyl)amine (TAEA) for 4-AMP allowed us to overcome the problems of emulsion and precipitation previously observed during the FAACST synthesis. The practical utility of this new technique and the use of FMOC-amino acid fluorides as new peptide coupling agents were demonstrated by the synthesis of several peptides such as those listed in Table 10.
Subject Area
Organic chemistry|Biochemistry
Recommended Citation
Sadat-Aalaee, Dean, "Rapid synthesis of short peptide segments via the FMOC-amino acid chloride solution technique (FAACST method)" (1990). Doctoral Dissertations Available from Proquest. AAI9101626.
https://scholarworks.umass.edu/dissertations/AAI9101626