On-line solid-phase extraction to enhance sensitivity in peptide biomarker analysis by microseparation techniques coupled to mass spectrometry: capillary liquid chromatography versus capillary electrophoresis

Abstract

In this study, on-line solid-phase extraction capillary liquid chromatography-mass spectrometry (SPE-CapLC-MS) and on-line solid-phase extraction capillary electrophoresis-mass spectrometry (SPE-CE-MS) were compared for the analysis of the opioid peptide biomarkers dynorphin A (1-7) (DynA), endomorphin 1 (End 1), and methionine-enkephalin (Met). First, a capillary liquid chromatography-mass spectrometry (CapLC-MS) method was established, which allowed limits of detection (LODs) of 0.5 μg/mL for Dyn A and Met, and 0.1 μg/mL for End 1. Then, a column switching setup operated by a 2-position/6-port micro-valve with a C18 enrichment column was assembled for SPE-CapLC-MS. Under optimized conditions, the LODs for the three peptides were lowered up to 1000-fold compared to CapLC-MS, until detecting 0.5 ng/mL concentrations. Repeatability (<0.2 % and <11 % RSD for retention times and peak areas, respectively), linearity (0.5-100 ng/mL), and durability (20 runs) of the enrichment column were appropriate, and the method was applied to analyze human plasma samples. Finally, the established SPE-CapLC-MS method was compared with a valve-free C18-SPE-CE-MS method previously described by our group for the analysis of these opioid peptides, using the same mass spectrometer. Both methods presented an evident difference regarding the need of a valve for the operation and allowed high preconcentration factors and quite similar LODs (until 0.5 and 0.1 ng/mL by SPE-CaLC-MS and SPE-CE-MS, respectively). Some other distinctions related to the instrumental set-up, procedure and method performance were also disclosed and discussed in detail.