Recent advances in on-line coupling of capillary electrophoresis to atomic absorption and fluorescence spectrometry for speciation analysis and studies of metal-biomolecule interactions

Speciation information is vital for the understanding of the toxicity, mobility and bioavailability of elements in environmental or biological samples. Hyphenating high resolving power of separation techniques and element-selective detectors provides powerful tools for studying speciation of trace elements in environmental and biological systems. During the last five years several novel hybrid techniques based on capillary electrophoresis (CE) and atomic spectrometry have been developed for speciation analysis and metal-biomolecule interaction study in our laboratory. These techniques include CE on-line coupled with atomic fluorescence spectrometry (AFS), chip-CE on-line coupled with AFS, CE on-line coupled with flame heated quartz furnace atomic absorption spectrometry (FHF-AAS), and CE on-line coupled with electrothermal atomic absorption spectrometry (ETAAS). The necessity for the development of these techniques, their interface design, and applications in speciation analysis and metal-biomolecule interaction study are reviewed. The advantages and limitations of the developed hybrid techniques are critically discussed, and further development is also prospected.     

Trace element speciation by ICP-MS in large biomolecules and its potential for proteomics

Latest studies on the chemical association of trace elements to large biomolecules and their importance on the bioinorganic and clinical fields are examined. The complexity of the speciation of metal-biomolecules associations in various biological fluids is stressed. Analytical strategies to tackle speciation analysis and the-state-of-the-art of the instrumentation employed for this purpose are critically reviewed. Hyphenated techniques based on coupling chromatographic separation techniques with ICP-MS detection are now established as the most realistic and potent analytical tools available for real-life speciation analysis. Therefore, the status and potential of metal and semimetals elemental speciation in large biocompounds using ICP-MS detection is mainly focused here by reviewing reported metallo-complexes separations using size-exclusion (SEC), ion-exchange (IE), reverse phase chromatography (RP) and capillary electrophoresis (CE). Species of interest include coordination complexes of metals with larger proteins (e.g. in serum, breat milk, etc.) and metallothioneins (e.g. in cytosols from animals and plants) as well as selenoproteins (e.g. in nutritional supplements), DNA-cisplatin adducts and metal/semimetal binding to carbohydrates. An effort is made to assess the potential of present trace elements speciation knowledge and techniques for "heteroatom-tagged" (via ICP-MS) proteomics.     

Arsenic speciation analysis

Nearly two dozen arsenic species are present in the environmental and biological systems. Differences in their toxicity, biochemical and environmental behaviors require the determination of these individual arsenic species. Considerable analytical progresses have been made toward arsenic speciation analysis over the last decade. Hyphenated techniques involving a highly efficient separation and a highly sensitive detection have become the techniques of choice. Methods based on high-performance liquid chromatography separation with inductively coupled plasma mass spectrometry, hydride generation atomic spectrometry, and electrospray mass spectrometry detection have been shown most useful for arsenic speciation in environmental and biological matrices. These hyphenated techniques have resulted in the determination of new arsenic species, contributing to a better understanding of arsenic metabolism and biogeochemical cycling. Methods for extracting arsenic species from solid samples and for stabilizing arsenic species in solutions are required for obtaining reliable arsenic speciation information.     

Non-chromatographic atomic spectrometric methods in speciation analysis: A review

In recent years, knowledge of the different chemical forms of the elements has gained increasing importance. There has been significant progress in methods that hyphenate chromatographic separations with atomic spectrometry. These hyphenated methods can provide the most complete information on the species distribution and even structure. However, they can be lengthy, relatively costly and difficult to bring to the routine. On the other hand, it is important to remember that chromatographic techniques represent only a minor part of the separation procedures available and, in certain cases, the application of basic chemistry to sample treatments can give quantitative information about specific chemical forms. In this sense, non-chromatographic procedures can provide methods that offer sufficient information on the elemental speciation for a series of situations. Moreover, these non-chromatographic strategies can be less time consuming, more cost effective and available, and present competitive limits of detection. Thus, non-chromatographic speciation analysis continues to be a promising research area and has been applied to the development of several methodologies that facilitate this type of analytical approach. In view of their importance, the present work overviews and discusses different non-chromatographic methods as alternatives for the speciation analysis of clinical, environmental and food samples using atomic spectrometry for detection.     

Modern trends in the speciation of selenium by hyphenated techniques

The complexity of selenium speciation in the environment and in living organisms results in broad analytical challenges. The importance of the selective determination of the particular species of this element, to understand its metabolism and biological significance in clinical chemistry, biology, toxicology, and nutrition, calls for state-of-the-art analytical techniques. In this paper hyphenated techniques are evaluated with particular emphasis on interfaced separation with element-selective detection and identification of the selenium compounds detected.     

Recent on-line processing procedures for biological samples for determination of trace elements by atomic spectrometric methods

Few of the elements present in nature play a metabolic role in living organisms. According to their abundance, these elements are classified as macro-, micro- or trace elements, representing 93%, 5% and around 1% respectively, of the total body weight. The remaining percentage could be attributed to those elements with unknown biological functions, to others which are present only because of the exposure to polluted environment or to those intentionally introduced into the body for a special treatment. This review summarizes and discusses the most recent publications related to the on-line processing of biological samples for trace element determination using atomic spectrometry-based detectors. Preconcentration/separation procedures based on solid phase or cloud point extractions, electrochemical deposition, microdialysis, as well as chemical vapor generation are the common practice for improving the sensitivity and selectivity of the available atomic spectrometric techniques. The advantages of using isotope dilution mass spectrometry in speciation studies are also emphasized. Digestion or leaching in oxidizing acidic mixtures aided by heat or by ultrasound or microwave radiation, performed off- or on-line, is necessary to previous steps when processing solid biological samples. The most relevant analytical figures of merit such as detection limits, enrichment factors and sample throughput as well as some aspects related to the on-line system configurations and accuracy assessments are critically presented.     

Sample treatment in chromatography-based speciation of organometallic pollutants.

Speciation analysis is nowadays performed routinely in many laboratories to control the quality of the environment, food and health. Chemical speciation analyses generally include the study of different oxidation state of elements or individual organometallic compounds. The determination of the different chemical forms of elements is still an analytical challenge, since they are often unstable and concentrations in different matrices of interest are in the microg l(-1) or even in the ng l(-1) range (e.g., estuarine waters) or ng g(-1) in sediments and biological tissues. For this reason, sensitive and selective analytical atomic techniques are being used as available detectors for speciation, generally coupled with chromatography for the time-resolved introduction of analytes into the atomic spectrometer. The complexity of these instrumental couplings has a straightforward consequence on the duration of the analysis, but sample preparation to separate and transfer the chemical species present in the sample into a solution to be accepted readily by a chromatographic column is the more critical step of total analysis, and demands considerable operator skills and time cost. Traditionally, liquid-liquid extraction has been employed for sample treatment with serious disadvantages, such as consumption, disposal and long-term exposure to organic solvent. In addition, they are usually cumbersome and time-consuming. Therefore, the introduction of new reagents such as sodium tetraethylborate for the simultaneous derivatization of several elements has been proposed. Other possibilities are based in the implementation of techniques for efficient and accelerated isolation of species from the sample matrix. This is the case for microwave-assisted extraction, solid-phase extraction and microextraction, supercritical fluid extraction or pressurized liquid extraction, which offer new possibilities in species treatment, and the advantages of a drastic reduction of the extraction time and the embodiment into on-line flow analysis systems. This new generation of treatment techniques constitutes a good choice as fast extraction methods for feasible species-selective analysis of organometallic compounds under the picogram level, that can be used for national regulatory agencies, governmental and industrial quality control laboratories, and consequently, for manufacturers of analytical instrumentation.     

Electrospray mass spectrometry: a tool for elemental speciation

Recent progress in the development of electrospray mass spectrometry (ESMS) as a tool for elemental speciation is reviewed. Reports wherein ESMS is used to qualitatively determine the presence of metal ions (inorganic, organometallic and complexed) and non-metallic inorganic species have grown exponentially over the last decade. In addition to elemental speciation, impact in other areas such as gas-phase chemistry, inorganic–organometallic chemistry and biological mass spectrometry has been prolific. The review is structured to cover each of the areas listed above, and also includes a brief introduction, discussion of the electrospray process, discussion of instrumentation and other relevant application areas. An overview of the types of species/complexes studied is given in each section along with a brief discussion of the application objectives and analytical aspects. Analytical considerations for the development of ESMS as a tool for elemental speciation are also raised, including, application, quantitation, sensitivity, limitations and future directions. The impact of speciation strategies involving stand-alone ESMS, ESMS coupled with on-line separation techniques and the inclusion of ESMS in dual (multiple) technique strategies are presented. High backgrounds due to chemical noise and signal suppression (matrix effects) appear to be two important factors limiting sensitive detection of most analytes. The use of sample pre-treatment, pre-concentration or separation techniques is necessary to alleviate these problems. Although ESMS currently suffers from a number of limitations, continued instrumentation and methods development will improve its capability and diversify the impact of ESMS as a tool for elemental speciation.     

Trends in speciation analysis of vanadium in environmental samples and biological fluids--a review

A comprehensive review is presented addressing recent trends in the speciation and determination of vanadium in environmental and biological sample matrices, including important analytical aspects such as sample clean up, pre-concentration and method development. Methodology based on both separation and spectroscopic techniques for the determination of vanadium speciation is discussed. A brief outline of analytical principles, together with an overview of the recent developments and applications of vanadium speciation determination is included. The newer methods for detecting metal ions including hyphenated spectroscopic techniques and sample preparation schemes are also discussed.     

Multidimensional approaches in biochemical speciation analysis

An understanding of the mechanisms controlling the essentiality and toxicity of trace elements in biological systems at the molecular level depends critically on the possibility of the identification, characterization, and quantification of chemical forms of these elements involved in life processes.Hyphenated techniques based on the combination of (electro)chromatography with ICP MS have become a routine tool for the analysis for metallospecies present in biological tissues. Finer analytical information on the true (down to individual species) speciation of trace elements in living organisms can be obtained by adding additional dimensions to the separation and detection steps, consisting of a sequential use of different HPLC separation mechanisms and capillary electrophoresis at the separation level, and of the use of electrospray MS, including collision induced dissociation MS, on the detection level. The value of the instrumental analytical data is decisively enhanced by the complementary use of molecular biology approaches involving gene identification, cloning and in vitro reproduction of the metal-controlled processes. A brief summary of the recent progress in biochemical speciation analysis is presented in the context of the latest research carried out in the authors' laboratory.     

Speciation of oxidation states of elements by capillary electrophoresis

Progress made in the last five years in the application of capillary electrophoresis methods to chemical speciation of elements is reported on the basis of over 100 literature references. The main trends observed include development of new on‐ and off‐capillary derivatization methods, application of new detection methods, and especially coupling of CE separation systems to powerful atomic spectroscopy and mass spectrometry instruments with various ionization techniques, providing either a sensitive element‐specific detection method or a third dimension for high performance separation. Besides numerous CZE and MEKC capillary electrophoresis methods only very few examples of CE speciation with capillary electrochromatography can be found. Concerning the chemical forms of elements determined, the new procedures developed are mostly focused on redox speciation of various oxidation states of elements, metal‐bound high molecular compounds, and organometallic species.     

Metal speciation in solid matrices

The literature on metal ion speciation in solid matrices is reviewed, taking into account its applications in the analysis of soil, sediment, biological materials, foodstuff and other solid samples. The pretreatment methods of various solid materials required for carrying out speciation studies have been highlighted. The basis of the methods of separation of different species from matrices, such as sequential extraction, selective extraction, etc. is discussed. The instrumental techniques used for the characterization of different chemical species in solid matrices have been mentioned. The literature survey reveals the analytical details of the developed methodologies, and these have been examined in terms of the limit of detection, precision and accuracy.     

Analysis of trace elements and their physico-chemical forms in natural waters

Trace elements in natural waters can be present in different physico-chemical forms, varying in size, charge and density properties. Knowledge of speciation is essential for understanding the transport, distribution, and biological uptake of trace elements in the environment. The development of techniques to provide reliable information on physico-chemical forms has, therefore, become a challenge within Analytical Chemistry.When selecting analytical methods for the determination of total concentrations or fractions of trace elements in natural waters, no exclusion of species should occur, or at least it must be accounted for. Furthermore, the determination limits must be sufficiently low to allow the actual concentrations to be determined with reasonable precision and accuracy. For very low concentrations, preconcentration techniques are applicable, provided the chemical yield of the spike represents that of the original species present. For methods meeting these criteria, the suitability for routine analysis should be considered.When the physico-chemical forms of trace elements are to be determined, the fractionation should take placein situ or shortly after sampling. As the concentrations involved in speciation studies may be extremely low, there is an increasing awareness of potential sources of errors influencing analytical results. Sample collection and separation/fractionation/concentration procedures prior to analysis are, therefore, essential within Analytical Chemistry, and the whole procedure must be taken into account when interpreting the results. There are, however, several requirements which should be met by techniques applicable for speciation purposes. In general, size fractionation techniques (e.g.in situ hollow fibre ultrafiltration) should be applied prior to the addition of any chemical reagents (charge fractionation techniques).     

Surveying selenium speciation from soil to cell—forms and transformations

The aim of this review is to present and evaluate the present knowledge of which selenium species are available to the general population in the form of food and common supplements and how these species are metabolized in mammals. The overview of the selenium sources takes a horizontal approach, which encompasses identification of new metabolites in yeast and food of plant and animal origin, whereas the survey of the mammalian metabolism takes a horizontal as well as a vertical approach. The vertical approach encompasses studies on dynamic conversions of selenium compounds within cells, tissues or whole organisms. New and improved sample preparation, separation and detection methods are evaluated from an analytical chemical perspective to cover the progress in horizontal speciation, whereas the analytical methods for the vertical speciation and the interpretations of the results are evaluated from a biological angle as well.     

Organised surfactant assemblies in analytical atomic spectrometry

The use of surfactant-based organised assemblies in analytical atomic spectroscopy is extensively and critically reviewed along three main lines: first, the ability of organised media to enhance detection of atomic spectroscopic methods by favourable manipulation of physical and chemical properties of the sample solution second, the extension of separation mechanisms by resorting to organised media and third a discussion of synergistic combinations of liquid chromatography separations and atomic detectors via the use of vesicular mobile phases. Changes in physical properties of sample solutions aspirated in atomic spectrometry by addition of surfactants can be advantageously used in at least four different ways: (i) to improve nebulisation efficiency; (ii) to enhance wettability of solid surfaces used for atomisation; (iii) to improve compatibility between aqueous and organic phases; and (iv) to achieve good dispersion of small particles in “slurry” techniques. Controversial results and statements published so far are critically discussed.The ability of surfactant-based organised assemblies, such as micelles and vesicles, to organise reactants at the molecular level has also been applied to enhance the characteristics of chemical generation of volalite species of metals and semi-metals (e.g., hydride or ethylide generation of As, Pb, Cd, Se, Sn, and cold vapour Hg generation) used in atomic methods. Enhancements in efficiency/transport of volatile species, increases in the reaction kinetics, stabilisation of some unstable species and changes in the selectivity of the reactions by surfactants are dealt with.Non-chromatographic cloud-point separations to design pre-concentration procedures with subsequent metal determination by atomic methods are addressed along with chromatographic separations of expanded scope by addition of surfactants to the conventional aqueous mobile phases of reversed-phase high-performance liquid chromatography.Finally, the synergistic effect of using vesicles to improve both the separation capabilities of reversed-phase HPLC and the detectability of atomic detectors by on-line vesicular hydride generation is described. In particular, the possible separation mechanisms responsible for micellar and vesicular mobile phases in reversed-phase chromatographies are analysed and compared. The possible effect of modification of stationary phases by monomers of the surfactants should also be taken into account. The application of such on-line couplings to develop new hybrid approaches to tackle modern problems of trace element speciation for As, Hg, Se, and Cd completes this revision of the present interface between analytical atomic spectroscopy and surfactant-based organised assemblies.     

Analytical methods for the determination of arsenosugars—A review of recent trends and developments

Arsenic-containing carbohydrates, generally termed arsenosugars, have been the subject of increasing analytical interest in arsenic speciation analysis. The present review gives an overview concerning achievements and trends in the field of instrumental analysis of arsenosugars. The typical experimental approaches for sample pre-treatment, extraction, separation and detection are discussed. Current possibilities and limitations of modern instrumental techniques are pointed out.     

Speciation analysis of heavy metals in natural waters: a review.

Metal speciation in natural waters is of increasing interest and importance because toxicity, bioavailability, environmental mobility, biogeochemical behavior, and potential risk in general are strongly dependent on the chemical species of metals. This paper provides an overview of the need for speciation of heavy metals in natural waters, the chemical and toxicological aspects of speciation, and the analytical procedures for separation and the different techniques for final determination that are used today. The trends and developments of speciation are also discussed. Finally, the case of chromium (Cr) was selected for a detailed presentation because the speciation of this metal has attracted a great deal of interest in view of the toxic properties of Cr(VI).     

Multielement trace-element speciation in metal-biomolecules by chromatography coupled with ICP-MS

The state-of-the-art of multi-element speciation in biological materials by hybrid techniques with inductively coupled plasma-mass spectrometric (ICP-MS) detection is critically reviewed with special attention to multi-metal speciation in metallothioneins (MT) and metallothionein-like proteins (MLP). The most common separation techniques used for the purpose include conventional size-exclusion chromatography (SEC) (with off-line detection) and anion-exchange (AE) or reversed-phase (RP) HPLC coupled on-line to ICP-MS. Advantages and limitations of the different commercial element-selective ICP-MS detectors, including the quadrupole (Q), double focusing (DF), and time-of-flight (TOF) mass analysers, are discussed. Procedures reported for speciation analysis, by use of hybrid techniques, for multi-metal speciation in mussel MLP and rabbit liver MT are illustrated with special emphasis on work performed in the authors' laboratory.