Speciation of inorganic selenium and selenoaminoacids by on-line reversed-phase high-performance liquid chromatography–focused microwave digestion–hydride generation-atomic detection

Abstract

A high-performance liquid chromatographic–microwave digestion–hydride generation system coupled on-line with three atomic detectors (atomic absorption, inductively coupled plasma atomic emission and inductively coupled plasma mass spectrometry) has been developed and investigated for selenium species separation and determination. Total inorganic selenium, selenomethionine and selenoethionine are separated by reversed-phase chromatography prior to on-line microwave digestion of selenocompounds with a KBrO₃–HBr mixture to form continuously Se^IV, which is finally transformed into H₂Se, also in a continuous manner, with a merging flow of sodium borohydride. Detection limits obtained for each Se species in water and urine using the three atomic detectors have been worked out and compared (i.e., for Se^IV DLs were 6.8 µg l^–1 by AAS, 30 µg l^–1 by ICP-AES and 0.16 µg l^–1 by ICP-MS). The integrated flow system, HPLC–MW digestion–HG-atomic detection, proposed here allows, in a single injection, reliable speciation in urine of the selenoaminoacids tested versus total inorganic selenium. Further speciation of the overlapped inorganic Se^IV and Se^VI peaks is accomplished by a second injection of the urine sample to determine only Se^IV by avoiding microwave heating. Results on Se speciation in human urine have shown that more speciation information of the actual species, perhaps unknown, present in real samples could be gathered by resorting to the use of two, or more, atomic detectors coupled to the same separation scheme.