posted on 2012-11-12, 00:00authored byDaniel Rentsch, Christian Stähelin, Markus Obkircher, Roland Hany, Marina Simeunovic, Daniel Samson, Günther Loidl, Fritz Dick
The quality of preloaded Wang resins is very important
for the
success of solid-phase peptide syntheses (SPPS). A critical factor
is the capping of remaining hydroxyl groups after loading with the
first amino acid, since these free alcohols lead to truncated sequences
during the following SPPS steps. Because the detection of hydroxyl
groups by color tests is difficult and unreliable, the capping efficiency
is often controlled by time-consuming peptide test syntheses. Here,
we describe a two-dimensional, high resolution magic angle spinning
NMR method for the quantitative determination of remaining 4-alkoxybenzyl
alcohols in Fmoc-Xaa-Wang resins with a detection limit of 1 mol-%.
The NMR method was validated with samples of known ratios between
Fmoc-Ala-Wang and 4-alkoxybenzylalcohol resin. Application to a set
of preloaded Fmoc-Ala- and Fmoc-Thr(tBu)-Wang test resins demonstrated
that the full range of essential amino acids can be quantified without
further spectrometer calibration. Compared to established test synthesis
protocols, the NMR method represents not only advantages in terms
of time and cost savings but also eliminates all inaccuracies due
to further sample treatment like SPPS and cleavage from the resin.