Review article Overview of hyphenated techniques using an ICP-MS detector with an emphasis on extraction techniques for measurement of metalloids by HPLC–ICPMS William Maher a, ⁎, Frank Krikowa a , Michael Ellwood b , Simon Foster a , Rajani Jagtap a , George Raber c a University of Canberra, Ecochemistry Laboratory, Institute for Applied Ecology, Faculty of Applied Science, ACT 2601, Australia b Research School of Earth Sciences, The Australian National University, Canberra, ACT 0200, Australia c Institute of Chemistry-Analytical Chemistry, Karl-Franzens University Graz (Uni-Graz) Stremayrgasse 16/3 Stock 8010 Graz, Austria abstract article info Article history: Received 31 August 2011 Received in revised form 20 February 2012 Accepted 21 March 2012 Available online 6 April 2012 Keywords: Metalloids Species Plants and animal tissues Hyphenated ICPMS, HPLC–ICPMS Extraction In this review we present an overview of hyphenated techniques using an ICPMS detector with an emphasis on extraction techniques for the measurement of metalloids by high pressure liquid chromatography–inductively plasma mass spectrometry (HPLC–ICPMS). Five modes of using hyphenated ICPMS systems; HPLC–ICPMS, HPLC–hydride generation-ICPMS, Cryogenic trapping ICPMS, in-situ Cryogenic trapping ICPMS and Gas Chromatography–ICPMS are described together with their application for the measurement of arsenic, selenium, mercury and antimony species. Two classes of metalloid species are described; “Easy to extract species,” stable species existing as discrete molecules or relatively weakly bound to cellular constituents, and Hard to extract species,” unstable species that dissociate on extraction and species incorporated within cellular constituents such as proteins. Measurements described include, arsenic species: arsenobetaine, arsenoribosides, arsenic bound to lipids and phytochelatins and other minor arsenic species including thioarsenic species. Selenium species: selenocysteine and selenomethionine, Se-methyl selenomethionine, Se-methyl selenocysteine, y-glutamyl-Se- methylselenocysteine, dimethylselenide and dimethyldiselenide. Mercury species: inorganic Hg and methyl Hg. Antimony species: antimonite and antimonate. Germanium species: inorganic. Extraction methods are discussed in terms of their extraction efficiencies, stability of species and artifact formation. © 2012 Elsevier B.V. All rights reserved. 1. Introduction Over the last thirty years there has been a shift from measuring only total metalloid concentrations to measuring metalloid species (see Section 2). With the advent and commercialization of the inductive coupled plasma mass spectrometer (ICPMS), hyphenated techniques coupling High Performance Liquid Chromatography (HPLC) and Gas Chromatography (GC) are routinely used to separate and quantify metalloid species such as those formed by arsenic, selenium and mercury. Speciation analysis of arsenic, selenium and mercury in food and biological samples is widely documented including a number of reviews [1–12]. However, nearly all these reviews concentrate on HPLC separations and only report sample extraction techniques in passing. In this review we discuss why metalloid species need to be measured, describe the five common hyphenated ICPMS systems used and discuss two classes of metalloid species; “Easy to extract species;” stable species existing as discrete molecules or relatively weakly bound to cellular constituents, and “Hard to extract species;” unstable species that disso- ciate on extraction and species incorporated within cellular constituents such as proteins. Extraction methods are discussed in terms on their extraction efficiencies, stability of species and artifact formation. 2. What are chemical species? The International Union of Pure and Applied Chemists [13] have defined a chemical species as the “Specific form of an element defined as to its isotopic composition or oxidation state, and/or complex or molecular structure.” The speciation of an element is the “Distribution of an element amongst defined chemical species in a system” while speciation analysis is the “Analytical chemistry: analytical activities of identifying and/or measuring the quantities of one or more individual chemical species in a sample.” Some examples of metalloid species are shown in Table 1. Species may range from different isotopes and valence states to complex organic molecules. Metalloids, such as As, Se, Hg, Sb etc., are different to metals due to their ability to form covalent bonds with carbon while metals form ionic complexes. Arsenic provides an example of the diverse variety of metalloid species that can be encountered in seafood (Fig. 1) with arsenic species existing as anions, cations and zwitterions. 3. Why measure chemical species? The importance of measuring metalloid species has been appre- ciated for a long time mainly because of the wide spread knowledge of arsenic's toxicity and its use as a poison [14,15]. However, until the early 1980s it was common practice to only measure total metalloid Microchemical Journal 105 (2012) 15–31 ⁎ Corresponding author. Tel.: + 61 2 62012531; fax: + 61 2 6201 2328. E-mail address: Bill.Maher@canberra.edu.au (W. Maher). 0026-265X/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.microc.2012.03.017 Contents lists available at SciVerse ScienceDirect Microchemical Journal journal homepage: www.elsevier.com/locate/microc