Trace LC/MS quantitative analysis of polypeptide biomarkers: impact of 1-D and 2-D chromatography on matrix effects
Eduard Rogatsky1, Vlad Tomuta, Harsha Jayatillake
1Core Analytical Laboratory at General Clinical Research Center, Albert Einstein College of Medicine, Yeshiva University, Bronx, NY 10461, USA. rogatsky@aecom.yu.edu
Abstract:
We investigated the impact of one dimension (single reverse phase (RP) column) and two dimension (two different RP columns) chromatographic methods on SIM (MS) and multiple reaction monitoring (MRM; MS/MS) performance from human plasma. We find that MRM analysis is clearly preferable for 1-D applications; however, implementation of SIM detection in conjunction with 2-D separation technique resulted in an over 60-fold increase in analyte peak area and improved S/N compared to MRM for our analyte, human C-peptide. Implementation of a 2-D RP-RP technique with SIM detection is capable of eliminating matrix effects and greatly increases signal response and data quality. For two large peptides in complex biological samples, we found that a 2-D approach performed better than high quality sample preparation together with 1-D chromatography and MRM, even on a high-end mass spectrometer.
