Perfluoro-tert-butanol for selective on-resin detritylation: a mild alternative to traditionally used methods.

Abstract

Solid-phase synthesis of cyclic, branched or side-chain-modified peptides typically involves introduction of a residue carrying a temporary side-chain protecting group that undergoes selective on-resin removal. In particular, Nα-Fmoc-Nε-(4-methyltriphenylmethyl) (Mtt)-protected lysine and its shorter analogues are commercially available and extensively used in this context. Nevertheless, rapid reliable methods for on-resin removal of Mtt groups in the presence of tert-butyloxycarbonyl (Boc) groups are needed. Current commonly used conditions involve low concentrations (1-3%) of trifluoroacetic acid (TFA) in dichloromethane, albeit adjustment to each specific application is required to avoid premature removal of Boc groups or cleavage from the linker. Hence, a head-to-head comparison of several deprotection conditions was performed. The selected acids represent a wide range of acidity from TFA to trifluoroethanol. Also, on-resin removal of the N-(4-methoxytriphenylmethyl) (Mmt) and O-trityl groups (on serine) was investigated under similar conditions. The mildest conditions identified for Mtt deprotection involve successive treatments with 30% hexafluoroisopropanol (HFIP) or 30% perfluoro-tert-butanol [(CF3)3COH] in dichloromethane (3 × 5 or 3 × 15min, respectively), while 30% HFIP, 30% (CF3)3COH, or 10% AcOH-20% trifluoroethanol (TFE) in CH2Cl2 (3 × 5min) as well as 5% trichloroacetic acid in CH2Cl2 (3 × 2min) enabled Mmt removal. Treatment with 1% TFA with/without 2% triisopropylsilane added (3 × 5min), but also prolonged treatment with 30% (CF3)3COH (5 × 15min), led to selective deprotection of an O-Trt group on a serine residue. In all cases, the sequences also contained N-Boc or O-tBu protecting groups, which were not affected by 30% HFIP or 30% (CF3)3COH even after a prolonged reaction time of 4h. Finally, the optimized conditions involving HFIP or (CF3)3COH proved applicable also for selective deprotection of a longer resin-bound peptide [i.e., Ac-Gly-Leu-Leu-Lys(Mtt)-Arg(Pbf)-Ile-Lys(Boc)-Ser(tBu)-Leu-Leu-RAM-PS] as well as allowed for an almost complete deprotection of a Dab(Mtt) residue.