Determination of Metallothionein Isoforms in Fish by Cadmium Saturation Combined with Anion Exchange HPLC-ICP-MS

摘要

The content of metallothionein (MT) in fish can serve as a biomarker for heavy metal contamination in an aquatic environment. In this paper, an improved sample separation and quantification method using cadmium saturation combined with anion exchange high performance liquid chromatography–inductively coupled plasma mass spectrometry (HPLC–ICP-MS) was developed to measure the two major isoforms of MT (MT-I and MT-II) in fish. Replacing sulfur with cadmium as the quantitative element by cadmium saturation reduced the instrumental requirement and analysis cost. In the chromatographic separation, NH~(3)–NH~(4)Cl buffer was applied as the mobile phase instead of Tris–HCl buffer to prevent carbon deposition on the ICP cone. The chloride concentration was found to be a key factor to the separation of MT-I and MT-II, and the optimum chromatographic condition was 27?mM of NH~(3)–NH~(4)Cl in isocratic mode, with a pH of 8.6 and a flow rate of 0.8?mL?min_(?1). The limits of detection were 0.7?μg?g_(?1)for MT-I and 1.0?μg?g_(?1)for MT-II, with recoveries ranging from 96.2 to 104.2% for MT-I and 84.7 to 90.4% for MT-II in different fish tissues. The relative standard deviations were less than 8% for both isoforms. Graphical abstract.