Kinetics of interaction of Cr(VI) and Cr(III) with serum constituents and detection of Cr species in human serum at physiological concentration levels

Abstract

A novel analytical method, based on monolithic convective interaction media (CIM) chromatography coupled to UV and inductively coupled plasma mass spectrometry (ICP-MS) detectors, was developed to investigate the kinetics of interactions of Cr(VI) and Cr(III) with serum constituents, and perform Cr speciation at physiological concentration levels. Cr(VI) was separated from Cr-transferrin (Cr-Tf) and Cr-albumin (Cr-HSA) on a CIM diethylamino (DEAE) column using linear gradient elution from 100% buffer A (50 mM Tris-HCl + 10 mM NaHCO₃, pH 7.4) to 60% buffer B (buffer A + 2 M NH₄Cl) in 10 min at a flow rate of 1 mL min^-1. Good column recovery of separated Cr species (close to 100%) and satisfactory method repeatability (RSDs below 8%) was obtained. Kinetics of interaction of Cr(VI) and Cr(III) with serum constituents were followed after human serum was doubly spiked with enriched ⁵⁰Cr(VI) and ⁵³Cr(III) solutions, and speciation analysis applied from 5 min up to 48 h after spiking. The results showed that in serum ⁵³Cr(III) rapidly interacted with Tf, while ⁵⁰Cr(VI) reduction was slow. 48 h after spiking, more than 90% of added ⁵³Cr(III) was bound to Tf and the remaining 10% associated with HSA. About 20% of spiked ⁵⁰Cr(VI) was still present in serum, while the resulting ⁵⁰Cr(III) was bound predominantly to Tf. High sensitivity of the developed speciation procedure enabled detection of Cr-Tf complex as the main Cr species in human serum at physiological concentration levels. To the best of our knowledge, Cr-Tf concentrations in serum of unexposed individuals have not been reported yet.

Keywords: Cr speciation; Human serum; Physiological concentration levels; Stable isotopic tracers.