Investigation of mercury-containing proteins by enriched stable isotopic tracer and size-exclusion chromatography hyphenated to inductively coupled plasma-isotope dilution mass spectrometry

In order to investigate trace mercury-containing proteins in maternal rat and their offspring, a method of enriched stable isotopic tracer (¹⁹⁶Hg and ¹⁹⁸Hg) combined with size-exclusion chromatography (SEC) coupled to inductively coupled plasma-isotope dilution mass spectrometry (ICP-IDMS) was developed. Prior to the analysis, ¹⁹⁶Hg- and ¹⁹⁸Hg-enriched methylmercury was administrated to the pregnant rats. Then the mercury-containing proteins in serum and brain cytosol of the dam and pup rats were separated by size-exclusion columns and the mercury was detected by ICP-MS. The ICP-MS spectrogram of the tracing samples showed significantly elevated ¹⁹⁶Hg and ¹⁹⁸Hg isotopic signals compared with the natural ones, indicating that the detection sensitivity could be increased by the tracer method. The contents of mercury in chromatographic fractions of the dam and pup rat brain cytosol were quantitatively estimated by post-column reverse ID-ICP-MS. The quantitative speciation differences of mercury in brain cytosol between the dam and pup rats were observed, indicating that such studies could be useful for toxicological estimation. Additionally, the isotopic ratio measurement of ¹⁹⁸Hg/²⁰²Hg in the tracing samples could be used to identify the artifact mercury species caused in the analytical procedure. The study demonstrates that the tracer method combined with high-performance liquid chromatography (HPLC)-ICP-IDMS could provide reliably qualitative and quantitative information on mercury-containing proteins in organisms.