Full Paper High-speed separation of arsenic compounds using narrow-bore high-performance liquid chromatography on-line with inductively coupled plasma mass spectrometry Sunanta Wangkarn and Spiros A. Pergantis* School of Biological and Chemical Sciences, Birkbeck College, University of London, Gordon House, 29 Gordon Square, London, UK WC1H 0PP. E-mail: s.pergantis@chem.bbk.ac.uk Received 6th March 2000, Accepted 14th April 2000 Published on the Web 18th May 2000 The growing importance of arsenic speciation analysis has led to the development of a wide range of high- performance liquid chromatographic based hyphenated techniques. During the present study a method was developed for the high-speed separation of several biologically and environmentally important arsenic compounds. The method is based on the use of an octadecyldimethylsilyl reversed-phase narrow-bore HPLC column. Separation of anionic arsenic species [arsenite (Aite), dimethylarsinic acid (DMAA), monomethylarsonic acid (MMAA), and arsenate (Aate)] can be achieved using a mobile phase containing 5 mM tetrabutylammonium hydroxide as the ion-pairing reagent, at pH 6.0, in less than 2 min, when employing a ¯ow rate of 0.7 ml min 21 . Adding 4-hydroxyphenylarsonic acid as the internal standard prolongs the total separation time by 30 s. On-line coupling with inductively coupled plasma mass spectrometry affords high sensitivity, as well as low limits of detection (low ppb or pg of arsenic). The in¯uence of mobile phase pH and ion-pairing reagent concentration on the separation ef®ciency was studied. A loss of resolution occurs with increasing ion-pairing reagent concentration; the optimum pH is between 6.0 and 6.2. The ion-pair reversed- phase narrow-bore HPLC-ICP-MS method was subsequently applied to the speciation of arsenic in wine and kelp samples. Aite at trace levels was found to be the only arsenic species present in several wines. Average spike recoveries for Aite, Aate, MMAA and DMAA were 95¡3, 94¡5, 98¡1 and 92¡1%, respectively, for all wines examined. The method was also used for the speciation of four arsenosugars and DMAA in a kelp powder extract. Introduction Elemental speciation is rapidly gaining recognition as a valid approach for making risks assessments for arsenic in environ- mental and biological materials. The approach is well justi®ed because both the toxicity and environmental fate of arsenic depend strongly on oxidation state and organic substituents. 1 As a result, knowledge of arsenic species' concentrations can help to make more accurate assessments of environmental impact and health risks. 1±4 In contrast, knowledge of the total arsenic content of a sample provides limited information regarding a material's suitability for a given purpose. 5,6 Consequently, a wide range of analytical techniques and methods have been developed for the identi®cation and quanti®cation of arsenic species present in environmental and biological samples. 7±11 Most analytical methods currently used for arsenic specia- tion analysis are based on hyphenated techniques that employ high-performance liquid chromatography (HPLC) coupled on-line with very sensitive element speci®c detectors, e.g., inductively coupled plasma mass spectrometry (ICP-MS), 11±16 hydride generation atomic ¯uorescence spectrometry (HG- AFS), 7 HG atomic absorption spectrometry (HG-AAS), 17,18 HG-ICP atomic emission spectrometry (AES), 19 HG-ICP- MS 20 or electrospray tandem mass spectrometry (ES- MS). 8,10,21 Even though there have been over 150 publications in the last decade describing HPLC based methods for arsenic speciation the approach is not yet routine. Numerous researchers have tried to address this by developing methods with short analysis times, improved chromatographic resolu- tion, high precision, and accuracy. In this respect a signi®- cant advancement was made recently by Le and Ma, 7 when four arsenic species, i.e., arsenite (Aite), dimethylarsinic acid (DMAA), monomethylarsonic acid (MMAA) and arsenate (Aate), were separated within 2±4 min by using HPLC guard columns coupled with HG-AFS; however, relatively poor chromatographic resolution was achieved. On-line element speci®c detection often involves hydride generation coupled with an atomic spectrometric detector, i.e., an atomic absorption or an atomic ¯uoresence spectrometer. Both detectors are widely available and relatively inexpensive to purchase and maintain. However, it has been reported that in some cases hydride generation is less ef®cient in generating arsines from DMAA than from Aite. 22 Also, arsenobetaine (AsB), arsenocholine (AsC) and arsenosugars (AsSugar) do not form volatile hydrides under normal conditions, and must therefore be adequately decomposed prior to being converted into arsines. HPLC-ICP-MS has been used extensively to overcome some of the shortcomings of hydride generation techniques and is currently considered state-of-the-art for elemental speciation, as it combines the resolving power of HPLC with the speci®city and high sensitivity of ICP-MS. Even though ICP-MS offers several advantages for trace arsenic speciation, its relatively high operating costs prevent it from being considered the ideal detector for routine speciation analysis. Making use of high-speed chromatographic methods can alleviate this. A desired situation would involve separating a mixture of arsenic species and detecting them in the time normally required for determining the total arsenic content of a sample. For example, a method that allows for the separation of a mixture of arsenic species in less than 2 min would provide increased sample throughput and therefore reduce the costs associated with arsenic speciation analysis. This study focuses on the development of ion pair (IP) reversed-phase (RP) narrow-bore HPLC coupled on-line with ICP-MS for the speciation of arsenic compounds in DOI: 10.1039/b001810o J. Anal. At. Spectrom., 2000, 15, 627±633 627 This journal is # The Royal Society of Chemistry 2000