From 2000 to the end of 2003, more than 400 scientific publications have appeared dealing with some aspect of the determination of arsenic species. This review attempts to draw out the most topical or useful of these reports, and we provide now a synopsis of some interesting areas and trends. Extraction efficiencies vary greatly between samples, thus it appears unrealistic to strive for a universal optimal extraction procedure for arsenic species. Rather, sample extraction procedures should be tailored to the particular application and desired analytes. Most arsenicals are reasonably stable and can be stored frozen for long periods. Some, however, are labile even when stored frozen, and their rate of conversion depends on the sample matrix. There is a need for a thorough study of stability of arsenicals/storage conditions which could provide information on the fundamental transformation processes at work. Such a study should be based on existing information on physico-chemical properties of redox reactions such as stability diagrams as a function of pH and redox potentials. A neglected area of arsenic speciation research is that dealing with the "insoluble" fraction which is thought to comprise "protein- bound" arsenic and/or "lipid-arsenic". Research in these areas is currently hindered by lack of suitable analytical methods, and increased attempts should be made to overcome these problems. Application of the right analytical strategy is likely to lead to novel and interesting results. Improvements in chromatographic separation have been reported using HPLC with gradient elution, but the application of these techniques to real samples may be limited because of severe matrix effects. CE provides excellent separation of many arsenicals but the small sample sizes preclude its use for real samples, and this restriction is unlikely to be overcome in the near future. HPLC-ICPMS is the most powerful and commonly used method for arsenic speciation analysis, and this usage should continue to grow as the required (but expensive) instrumentation becomes more widespread. The method provides reliable quantitative data for arsenic species at environmentally relevant levels in various and diverse matrices. Methods based on HPLC coupled to AAS, AES or AFS, and used in conjunction with HG, provide lower cost options for arsenic speciation analysis. The application of these techniques to refractory arsenicals (arsenobetaine for example) through the use of a decomposition step should be handled cautiously because the degree of degradation is strongly influenced by the matrix. We expect a decline in the coming years in research investigations into arsenic speciation analysis using optical detection systems. The application of these techniques to study various aspects of arsenic speciation, however, is likely to continue. Molecular mass spectrometric methods have provided the most interesting recent results in terms of structural elucidation of new metabolites, and they hold the promise of identifying more novel arsenicals and thereby better explaining arsenic's fate and role in biological systems. Molecular MS is also increasingly being used to provide verification of chromatographic peaks detected and quantified with ICPMS. The use of molecular MS alone, however, is likely to stay mainly in qualitative analysis. An exception may arise if there is a need to routinely analyse a specific arsenical, for toxicological purposes for example, which would encourage development of a targeted sample preparation procedure applicable to molecular MS analysis. X-ray spectroscopic methods, long used for abiotic samples, are now being applied to biological samples and they have already provided interesting data on novel arsenic species and arsenic species in situ. Their ability to handle solid samples removes most concerns regarding sample extraction and sample storage which can reduce the usefulness of the traditional coupled techniques based on chromatography. X-ray spectroscopy is currently restricted to arsenic-rich samples but further developments in fluorescence detection methods may extend its use. The most topical issues on arsenic speciation research are the reduced species MA(III) and DNA(III) in human urine, and the recently reported thio arsenic species in sheep urine. On closer inspection, the data presented so far for MA(III) and DNA(III) are not totally convincing and require confirmation with appropriate sample collection, storage and preparation procedures. On the other hand, the data showing the presence of thio arsenic species look sound and there is the likelihood that other As-S compounds will be found. Given the current uncertainty surrounding MA(III) and DMA(III), the possibility that arsenic species assigned as such may in fact be thio arsenic species should be considered.
