Development of a SEC-ICP-MS platform for multielement metallobiomolecule profiling and quantitation using a blood serum reference material

The characterization of metallobiomolecules in biological fluids is essential for understanding metal homeostasis, biomarker discovery, as well as toxicity assessment. In this work, we present a size-exclusion chromatography inductively coupled plasma mass spectrometry (SEC-ICP-MS) platform for the comprehensive profiling of metallobiomolecules in human serum. This method enables the simultaneous detection and quantitation of ten metals and metalloids (Co, Mg, Ca, Cu, Zn, Fe, Mn, Pb, Se, Hg) within a single sample. The integration of post-column flow injection allows for element calibration, using ionic standards containing EDTA, total element determination following the injection of an acid-diluted blood serum sample, and instrument sensitivity monitoring and correction. Method validation was performed using the certified total element concentration in a human reference material (Seronorm Trace Elements Level 2). Element recoveries exceeded 80% for most analytes, both following total element determination and column elution, confirming the robustness and accuracy of the approach. An on-column EDTA injection strategy effectively mitigated metal/metalloid interactions with the stationary phase, enhancing column recoveries of Co and Zn while preventing cross-contamination between samples. This platform expands the number of elements that can be simultaneously monitored and quantified, and maximizes the information obtained for the analysis of each sample, providing an improved assessment of metal/metalloid distributions in human blood serum. The analysis of Seronorm Trace Elements Level 2 material provided novel insights into the element distribution across different relative molecular mass metallobiomolecules. Detected metallobiomolecule bands aligned with biomolecules known to be present in human serum and previous studies of this reference material, yet discrepancies in element distribution suggest possible element addition during reference material preparation. Its ability to provide detailed elemental distributions across biomolecular bands enhances its potential for applications in biomarker discovery, disease monitoring, and environmental exposure assessments.