The state-of-the-art of species-selective analysis for trace
metals and metalloids in biological materials by chromatographic and
electrophoretic separation techniques with element selective detection is
critically reviewed. The species of interest include organoarsenic,
organoselenium and metal complexes with bioligands such as phytochelatins,
metallothioneins, proteins and polysaccharides. The separation mechanisms
discussed include size-exclusion, anion- and cation-exchange and
reversed-phase HPLC and flatbed and capillary zone electrophoresis.
Advantages and limitations of various element selective (e.g.,
AAS, ICP-AES and ICP-MS) and molecule specific (electrospray MS/MS)
detection techniques used on- and off-line are discussed. The applications
of coupled techniques to the analysis of biological materials are
comprehensively reviewed in tabular form. Attention is paid to the sample
preparation and sources of error in bioinorganic speciation analysis.

Bio-inorganic speciation analysis by hyphenated techniques.

Liquid chromatography–inductively coupled plasma mass spectrometry

Indirect determination of cyanide ion and hydrogen cyanide by adsorptive stripping voltammetry at a mercury electrode

Selenium (Se) is an ultramicro essential nutrient and both inorganic (selenite and selenate) and organic (selenocysteine and selenomethionine) forms of Se can be used as nutritional sources. Metabolic pathways for Se in the body were studied for selenite and selenate, with the use of enriched 82Se, by speciation with separation by gel filtration HPLC and detection by element-specific mass spectrometry with ionization with inductively coupled argon plasma (HPLC-ICP MS). The concentrations of 82Se in organs and body fluids and the distributions of their constituents depending on the dose and time after the intravenous administration of 82Se-selenite and -selenate to rats were determined. Selenite was taken up by red blood cells within several minutes, reduced to selenide by glutathione, and then transported to the plasma, bound selectively to albumin and transferred to the liver. Contrary to selenite, intact selenate was either taken up directly by the liver or excreted into the urine. The 82Se of selenite origin and that of selenate origin were detected in the forms of the two Se peak materials in the liver, A and B. The former one was methylated to the latter in vivo and in vitro. The latter one was identical with the major urinary metabolite and it was identified as Se-methyl-N-acetyl-selenohexosamine (selenosugar). The chemical species-specific metabolic pathway for Se was explained by the metabolic regulation through selenide as the assumed common intermediate for the inorganic and organic Se sources and as the checkpoint metabolite between utilization for the selenoprotein synthesis and methylation for the excretion of Se.

Metabolic pathway for selenium in the body: speciation by HPLC-ICP MS with enriched Se.

Organised surfactant assemblies in analytical atomic spectrometry

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 SeIV, 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 SeIV 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 SeIV and SeVI peaks is accomplished by a second injection of the urine sample to determine only SeIV 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.

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

A vesicle mediated high performance liquid chromatographic (HPLC)–microwave digestion (MW)–hydride generation (HG) system coupled on-line with atomic absorption spectrometry (AAS) and inductively coupled plasma mass spectrometry (ICP-MS) has been assessed for selenium species separation and detection. Selenocystine, selenomethionine, selenoethionine, selenourea, SeIV and SeVI are separated by vesicle mediated chromatography prior to on-line selenocompounds microwave digestion with a KBrO3–HBr mixture to generate SeIV continuously, which is finally transformed into SeH2, in a continuous manner with a merging flow of sodium tetrahydroborate(III). Analytical characteristics of this coupling are compared with those obtained coupling HPLC–ICP-MS via conventional nebulisation. Detection limits (DLs) obtained for selenocystine, selenomethionine, selenoethionine, selenite and selenate in spiked human urine, when ICP-MS was used as detector, ranged between 1.0 and 5.3 µg l–1 (51–267 pg), while precision ranged between±3.4 and±8.4%. This continuous system, vesicle mediated HPLC–MW–HG–atomic detection, allows the separation and detection of selenocystine, selenourea, selenomethionine, selenoethionine, selenite and selenate in human urine. The analytical versatility of such coupling (with ICP-MS as the atomic detector) allows basal selenium speciation in urine. Three different normally occurring selenium species in human urine, simply diluted (1+1), have been found. The relative sophistication of the vesicle mediated–HPLC–MW–HG–ICP-MS interface, versus HPLC–ICP-MS via conventional nebulisation, can be justified because of its considerably higher sensitivity for most of the selenocompounds assayed, lower matrix interferences and the possibility of simultaneous interference free 77Se and 78Se monitoring.

Organic and inorganic selenium speciation in urine by on-line vesicle mediated high-performance liquid chromotography–focused microwave digestion–hydride generation–inductively coupled plasma mass spectrometry

Urinary selenium speciation by high-performance liquid chromatography-inductively coupled plasma mass spectrometry: advantages of detection with a double-focusing mass analyser with a hydride generation interface.

J. M. González Lafuente

Papers

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

Organic and inorganic selenium speciation in urine by on-line vesicle mediated high-performance liquid chromotography–focused microwave digestion–hydride generation–inductively coupled plasma mass spectrometry

Urinary selenium speciation by high-performance liquid chromatography-inductively coupled plasma mass spectrometry: advantages of detection with a double-focusing mass analyser with a hydride generation interface.

Differential-pulse voltammetric determination of low μg l−1 cyanide levels using EDTA, Cu(II) and a hanging mercury drop electrode