Precolumn isotope dilution analysis in nanoHPLC-ICPMS for absolute quantification of sulfur-containing peptides
Résumé
A novel generic approach based on precolumn isotope dilution nanoHPLC-ICPMS analysis was developed for the accurate absolute quantification of sulfur-containing peptides. A 34S-labeled, species-unspecific sulfur spike (sulfate), noninteracting with analyte peptides under the optimized HPLC condition, was added directly to the chromatographic eluents. Thus a generic sulfur standard permanently present during analysis was used for peptide quantification. Interference-free detection of the 32S and 34S isotopes in ICPMS was achieved by eliminating O2 + ions in a collision cell using Xe gas at 130 μL min -1. The detection limit for sulfur was 45 μg L-1 which corresponded to 1-2 pmol of individual peptides. The method was validated by the analysis of a standard peptide solution showing high accuracy (recovery 103%) and good precision (RSD 2.1%). The combination of nanoHPLC-ICP IDMS with nanoHPLC-ESI MS/MS allowed the precise quantification and identification of sulfur-containing peptides in tryptic digests of human serum albumin and salt-induced yeast protein (SIP 18) at the picomole level. © 2007 American Chemical Society.