Advancing Trace-Element Speciation in the Environment, Food, and Biota

The speciation of trace elements has become much more common in recent years, and in some cases is now required by certain regulatory bodies - such as for the analysis of hexavalent chromium in drinking water. At the same time, speciation methods are becoming sophisticated as researchers try to address more complex questions about how different metal species are transported and interact with the environment and specifically with plants, animals, and humans, often forming complexes with other compounds that are more complicated to analyze. Prof. Jörg Feldmann of the University of Aberdeen, in Scotland, has been advancing speciation methods for more than 20 years, and is currently using these methods to address a range of questions, such as which cultivars of rice can best resist taking up arsenic into the rice grain, and how metalloproteins and metallolipids are transported in fish. Feldman won the 2015 European Award for Plasma Spectrochemistry for this work, and he recently talked to us about it.

You have been working on the speciation of trace elements for more than 20 years, and this field has gained much more attention in recent years. Given the progress that you and others have made in advancing speciation, what are some of the biggest questions that researchers are trying to answer today in this field, and what are some of the challenges involved in developing analytical methods to try to answer them?

The speciation analysis field has matured over the last 20 years. At first, it was primarily used in its traditional format, in which chromatography was only coupled to element-specific detectors for identification of metal speciation. In later years, the methods became more sophisticated, and similar concepts started to be used for biomolecules such as metalloproteins or metal-containing lipids. Going forward, the challenges are twofold. The first is to develop novel analytical methodologies that are capable of identification and quantification of traces of complex organometallic compounds such as arsenolipids without having any arsenolipid standard available. The second challenge is to develop analytical methodologies that are amenable to identifying and quantifying relatively labile organometallic compounds with high biological activities such as metal peptides or metalloproteins.   

One major focus of your work has been the identification of processes in the environment and in biota with regards to trace elements, and specifically arsenic.  In one recent study in this area (1), you developed a method that combines reversed-phase high performance liquid chromatography (HPLC) with inductively coupled plasma–mass spectrometry (ICP-MS) to improve the ability to identify arsenic compounds in a lipid matrix such as in fish oil. What are the advantages of this method over previous methods for analyzing these compounds?

In fact, arsenic in foodstuff has been the focus of our work for the last decade. Inorganic arsenic is a class I carcinogen and may soon be regulated by implementing a WHO/FAO suggestion to have a maximum level of inorganic rice in polished rice. Analyzing rice for inorganic arsenic typically is done by coupling HPLC and ICP-MS where the ICP-MS guarantees arsenic-specific detection. However, if the arsenic species become more complex, such as in the classes of arsenolipids that can be found in marine food or nutraceuticals such as fish oils and that have been tested to be as cytotoxic as inorganic arsenic, new methodologies are needed. We couple HPLC on-line not only to ICP-MS but also directly in parallel to a mass spectrometer for molecular analysis. Since the compounds are easy to ionize, electrospray ionization mass spectrometry (ESI-MS) is used. Hence, reversed-phase HPLC separates the species; ICP-MS gives the arsenic-specific detection; using species-independent calibration enables quantification without having access to an arsenolipid standard; and ESI-MS delivers molecular information through accurate mass and fragmentation pattern.

Using this method, you have been able to identify previously unreported arsenolipid compounds. What can we learn from identifying these compounds?

Arsenolipids occur mainly in marine organisms. We have found them in fish oils, which are commonly used as food supplements, and also in commercial marine products such as herring, cod, oysters, seaweed, and mackerel. These compounds seem to occur in the milligram-per-kilogram range in all fish or seafood, but differences can be seen amongst the different classes of arsenolipids. In fish and seafood, arsenic-containing hydrocarbons, fatty acids and fatty alcohols can be identified, but in seaweeds arsenic-containing phospholipids have been identified in addition to the arsenic-containing hydrocarbons. Arsenic-containing hydrocarbons have recently been identified as cytotoxic and show biological activity in a similar concentration range as inorganic arsenic by influencing the developmental stages of a Drosophila fly in an in vivo experiment.

In another study (2), you developed a method using HPLC–ICP-MS to look at how different cultivars of rice plants take up arsenic from the soil through their roots and transfer them to their shoots and grains. How does your method address this question?

In this study, we wanted to evaluate how the different rice cultivars take up arsenic and transport them into the grain. Using dual mass spectrometer detection (HPLC–ICP-MS/ESI-MS), it was possible to identify how arsenic is complexed by so-called phytochelatins in the roots and subsequently trapped in the vacuoles of the root cells. The arsenic phytochelatin complexes are very labile and can only be detected by a chromatographic system that directly detects the arsenic on-line by ICP-MS and the entire molecule by ESI-MS. This complexation prevents the arsenic from being translocated farther up the plants and finally accumulating in the grain.  The cultivars with higher amounts of certain phytochelatins are less likely to have high amounts of carcinogenic inorganic arsenic in the rice grain.

Another method you developed to study trace elements in the environment uses preconcentration liquid chromatography followed by cold vapor atomic fluorescence spectroscopy to detect methylmercury - the form of mercury that presents the most concern in the food chain - in water at the sub-part-per-trillion level (3). How is this method different from previous methods?

Methylmercury is a neurotoxin, and there has been evidence that this neurotoxin can occur in staple foods such as rice when rice plants have been exposed to elevated levels of mercury. Hence, the quick and cheap biomonitoring of rice is necessary. Since even low levels of methylmercury are biologically active and detrimental when taken up by humans, there has been a need for a method to analyze methylmercury in foodstuff and water that is quick and cheap but also sensitive and robust. We developed an on-line method that can even be automated with affordable instrumentation: We used solid-phase extraction (SPE) to trap the methylmercury and then elute this again by a mobile phase together with inorganic mercury, which is then separated by a reversed-phase column. Subsequently the organomercury is converted by oxidation to inorganic mercury. This is followed by normal cold vapor atomic fluorescence detection (SPE–HPLC–CV-AFS). With this technique it is possible to detect 20 ppb from 1 ppm (20 ppq) of methylmercury in water and parts-per-billion or parts-per-million concentrations of methylmercury in seafood or rice.

You have coupled laser ablation to multicollector ICP-MS for analyzing trace elements in tissues in a method you called dynamic bioimaging (4). How does “dynamic bioimaging” work, and what can it tell us that other methods cannot?  

We were the first to use laser ablation coupled directly to ICP-MS for bioimaging of trace elements in biological tissues more than a decade ago. Elemental concentrations down to the parts-per-million range can be determined with a spatial resolution of 10 μm. Hence, pathological anomalies like abscesses can be determined but not subcellular structures. This field has rapidly expanded. However, stable isotope metal tracers have not been used in combination with bioimaging. In her PhD study, my student Dr. Dagmar Urgast investigated the use of tracers of the essential element zinc to study how zinc is taken up into different parts of an organ and how it is accumulated and excreted. The stable isotope tracers were given to the animal at different time points before the animal was sacrificed. The isotope signature remains, however, and thin sections of organs and subsequent interrogation with a laser along with simultaneous measurement of the zinc isotopes reveals different isotope maps - for example, the differences in time-dependent zinc accumulation can be seen directly in one brain; hence we exclude here any biological variability. Since we have not only the concentration of the element or isotope and its spatial distribution but also with the timed tracer experiment, the time factor is incorporated into the image. We call this dynamic bioimaging. This will help to identify how an organism guarantees homeostasis in the most important regions of the organs or the kinetics of how a toxic element is taken up and either detoxified or accumulated. 

References

(1)      K.O. Amayo, A. Raab, E.M. Krupp, H. Gunnlaugsdottir, and J. Feldmann, Anal. Chem.85(19), 9321–9327 (2013), DOI dx.doi.org/10.1021/ac4020935

(2)      B.L. Batista, M. Nigar, A. Mestrot, B. Alves Rocha, F. Barbosa Jr., A.H. Price, A. Raab, and J. Feldmann, J. Exp. Bot. 65(6), 1467–1479 (2014), doi:10.1093/jxb/eru018

(3)      C.-C. Brombach, B. Chen, W.T. Corns, J. Feldmann, and E.M. Krupp, Spectrochim. Acta Part B, 105, 103–108 (2015), http://dx.doi.org/10.1016/j.sab.2014.09.014

 (4)     D.S. Urgast and J. Feldmann, J. Anal At. Spectrom. 28, 1367–1371 (2013)

Jörg Feldmann joined the University of Aberdeen (Scotland) in 1997 as a lecturer, and became a full professor of Environmental Analytical Chemistry in 2004. He became the head of the chemistry department in 2012.

He has published approximately 200 peer-reviewed papers, about one third of which have been published in analytical chemistry journals whilst the others have been published in applied sciences journals, such as Science, PNAS, Plant Physiology, Environmental Sciences, Toxicology and Medical Sciences. He has received more than 8000 citations and currently has an h-index of 51. He recently received the biennial European Prize for Plasma Spectrochemistry 2015.

Feldmann received his PhD from the University of Essen (Germany), where he developed the first coupling of gas chromatography with inductively coupled plasma-mass spectrometry (GC–ICP-MS) to measure cryotrapped volatile metal compounds from landfill sites. He was awarded the prize of the University of Essen for the best PhD thesis that year.