Vesicle-mediated high performance liquid chromatography coupled to hydride generation inductively coupled plasma mass spectrometry for cadmium speciation in fish cytosols

Abstract

The use of vesicles as mobile phases in high performance liquid chromatography (HPLC) separations, coupled to Cd hydride generation inductively coupled plasma mass spectrometry (ICP-MS) detection, allows a synergic combination which is exploited here for cadmium speciation in commercially available standards of rabbit liver metallothioneins (MT) and eel liver and kidney. The two main isoforms of rabbit liver MT were readily resolved by using a concentration gradient (2–200 mM) of Tris-HCl buffer solution (pH 7.4) containing didodecyldimethylammonium bromide (DDAB) vesicles and a C18 reversed-phase column, previously modified by DDAB solution. Chromatographic behaviour of rabbit liver Cd MT fractions with this technique was similar to that observed by anion-exchange liquid chromatography. The highly selective and sensitive on-line cadmium hydride generation ICP-MS continuous detection provided a 5-fold sensitivity enhancement in comparison with ICP-MS detection with conventional nebulization. Unfortunately, only 3-fold improvements were observed for the corresponding metal detection limits, owing to the comparatively high blank Cd signal and higher signal-to-noise ratio characterising the volatile species generation procedure. The observed precisions were always better than ±4% for the Cd content of each chromatographic peak. Application of this hybrid technique, calibrating with Cd-rabbit liver MT standards, to real life cadmium speciation in cytosol samples of eel liver and kidney has been addressed. Most of the cadmium in such eel cytosols seems to occur bound to an isoform similar to MT 2 of rabbit liver. The validity of the proposed methodology for Cd speciation in such complex matrix real samples was checked by comparing results with those obtained with anion-exchange HPLC-HG-ICP-MS as a complementary technique. Cadmium recoveries were found to range from 95 to 102% for eel cytosol samples by the proposed vesicle-mediated HPLC-HG-ICP-MS technique.