Christoph Thomas, Norbert Jakubowski, Dietmar Stuewer, Dieter Klockow and Hendrik Emons
The speciation of selenocystine, selenocystamine, selenomethionine and selenoethionine was studied by on-line coupling of high performance liquid chromatography with inductively coupled plasma mass spectrometry with hydraulic high pressure nebulization (HHPN) for sample introduction employing a double-focusing magnetic sector field mass spectrometer in order to achieve the utmost sensitivity. Reversed-phase chromatography was applied to separate the organic Se compounds. The eluent was 30 mm ammonium buffer adjusted to pH 3.0 in methanol-water (5+95 v/v). The operating conditions for chromatographic separation (pH of the mobile phase, methanol content of the mobile phase, pH of the sample and sample uptake rate) were carefully optimized. With the developed procedure, detection limits below 0.02 ng ml–1 were achieved, so that even low pg levels could be detected when applying sample volume of 200 μl.