Levels of toxic arsenic species in native terrestrial plants from soils polluted by former mining activities

Arsenic is considered a toxic element for plants. However, the discovery of arsenic resistant and hyperaccumulating plant species has increased the interest in understanding the distribution of arsenic species in these environmental matrices. Although As can be present in plants under different chemical forms, such phytochelatins, the As remained in plants as free inorganic or methylated ions has a greater interest due to its higher mobility and toxicity, and therefore, its ecological impact and risk to health. The aim of this work consisted on the determination of the fraction of As present as toxic forms (inorganic and methylated species) present in native terrestrial plants from polluted soils by former mining activities (Mónica mine, NW Madrid, Spain), with high total arsenic concentration levels (up to 3,500 µg g-1), due to their higher mobility and the risk associated to their reintegration into other environmental compartments. Roots and aboveground parts were analysed separately, to assess possible transformations from translocation processes. Extractions were carried out with deionized water by microwave-assisted extraction, at a temperature of 90 °C and three extraction steps of 7.5 min each. Total extracted arsenic concentrations were determined by ICP-AES, showing extraction percentages from 9 to 39%. Speciation studies were performed by HPLC-(UV)-HG-AFS, and they showed the main presence of As(V) (up to 350 µg g-1), followed by As(III), in both plant parts. Monomethylarsonic acid (MMA) and trimethylarsine oxide (TMAO) were also found only in some plants. On the other hand, the use of 0.5 mol L-1 acetic acid as extractant led to higher extraction percentages (33-87%), but lower column recoveries, probably due to the extraction of arsenic compounds different to toxic free ions studied, which may come from biotransformation mechanisms carried out by plants to reduce arsenic toxicity. However, As(V) concentrations increased up to 800 µg g-1 in acid medium, indicating the probable release of As(V) from organoarsenic compounds and therefore a higher potential risk for the environment. From the easily soluble, water-extractable arsenic species concentration levels, it can be drawn that between 70 and 89% of the total arsenic in plants must have been biotransformed, so it is present under the form of different arsenic compounds. Still, high As concentration levels remain as toxic forms, predominantly As(V), reaching up to 190 µg g-1 considering roots and aboveground parts separately, and 350 µg g-1 considering the sum of both plant parts (more than double in acid medium), which may constitute an environmental risk due to its possible reintegration to the environment. Therefore, the study and control of native plants growing in As polluted soils is a relevant factor for environmental saf

Toxic arsenic compounds remain in native plant species from arsenic polluted soils

Conferencia sobre fitorremediación de suelos contaminados con metales pesados y arsénico y estudios de especiación de arsénico en las plantas autóctonas acumuladoras

Estudios de especiación de arsénico y acumulación de metales en muestras de interés medioambiental : Arsenic speciation and metal accumulation studies in environmental samples

Se ha estudiado la determinación de especies de arsénico y de contenidos totales de arsénico y metales pesados, específicamente cadmio, cromo, cobre, níquel, plomo y cinc, en muestras de interés medioambiental por su elevada capacidad acumuladora de metales, concretamente algas marinas comestibles y plantas terrestres procedentes de suelos contaminados por la actividad minera. La determinación de contenidos totales se ha llevado a cabo mediante espectrometría de emisión atómica con plasma de acoplamiento inductivo (ICP‐AES), así como por espectrometría de fluorescencia atómica con generación de hidruros (HG‐AFS), para bajos contenidos de arsénico. Las muestras fueron mineralizadas en medio ácido y calentamiento en horno de microondas. Los métodos fueron validados a través de su aplicación a materiales de referencia de matriz similar a la de las muestras, certificados en contenidos totales de los elementos seleccionados. Los resultados obtenidos mostraron su elevada capacidad de bioabsorción, especialmente en relación a los elevados contenidos de arsénico encontrados en algunas especies de algas pardas (Phaeophytas). En las plantas, se calcularon los factores de translocación, acumulación y biodisponibilidad de los elementos estudiados, permitiendo identificar a la especie Corrigiola telephiifolia como posible acumuladora de plomo e hiperacumuladora de arsénico. La determinación de especies de arsénico hidrosolubles en las muestras objeto de estudio, se llevó a cabo por cromatografía líquida de alta eficacia (HPLC) acoplado a ICP‐AES, HG‐ICP‐AES y HG‐AFS, incluyendo una etapa previa de foto‐oxidación. Los métodos desarrollados, mediante intercambio aniónico y catiónico, permitieron la diferenciación de hasta once especies de arsénico. Para el análisis de las muestras, fue necesaria la optimización de métodos de extracción, seleccionándose la extracción asistida por microondas (MAE) con agua desionizada. Asimismo, se realizaron estudios de estabilidad de arsénico total y de las especies hidrosolubles presentes en las algas, tanto sobre la muestra sólida como en sus extractos acuosos, evaluando las condiciones de almacenamiento adecuadas. En el caso de las plantas, la aplicación del diseño factorial de experimentos permitió optimizar el método de extracción y diferenciar entre las especies de arsénico presentes en forma de iones sencillos de mayor movilidad y el arsénico más fuertemente enlazado a componentes estructurales. Los resultados obtenidos permitieron identificar la presencia de arseniato (As(V)) y arsenito (As(III)) en las plantas, así como de ácido monometilarsónico (MMA) y óxido de trimetilarsina (TMAO) en algunas especies. En la mayoría de las algas se encontraron especies tóxicas, tanto mayoritarias (arseniato) como minoritarias (ácido dimetilarsínico (DMA)), así como hasta cuatro arsenoazúcares. Los resultados obtenidos y su estudio a través de la legislación vigente, mostraron la necesidad de desarrollar una reglamentación específica para el control de este tipo de alimentos. La determinación de especies de arsénico liposolubles en las muestras de algas se llevó a cabo mediante HPLC, en modo fase inversa, acoplado a espectrometría de masas con plasma de acoplamiento inductivo (ICP‐MS) y con ionización por electrospray (ESI‐MS), permitiendo la elucidación estructural de estos compuestos a través de la determinación de sus masas moleculares. Para ello, fue necesaria la puesta a punto de métodos extracción y purificación de los extractos. La metodología desarrollada permitió identificar hasta catorce especies de arsénico liposolubles en las algas, tres de ellas correspondientes a hidrocarburos que contienen arsénico, y once a arsenofosfolípidos, además de dos especies desconocidas. Las masas moleculares de las especies identificadas fueron confirmadas mediante cromatografía de gases acoplada a espectrometría de masas (GC‐MS) y espectrometría de masas de alta resolución (HR‐MS). ABSTRACT The determination of arsenic species and total arsenic and heavy metal contents (cadmium, chromium, cooper, nickel, lead and zinc) in environmental samples, with high metal accumulator capacity, has been studied. The samples studied were edible marine algae and terrestrial plants from soils polluted by mining activities. The determination of total element contents was performed by inductively coupled plasma atomic emission spectrometry (ICP‐AES), as well as by hydride generation atomic fluorescence spectrometry (HG‐AFS) for low arsenic contents. The samples studied were digested in an acidic medium by heating in a microwave oven. The digestion methods were validated against reference materials, with matrix similar to sample matrix and certified in total contents of the elements studied. The results showed the high biosorption capacity of the samples studied, especially regarding the high arsenic contents in some species of brown algae (Phaeophyta division). In terrestrial plants, the translocation, accumulation and bioavailability factors of the elements studied were calculated. Thus, the plant species Corrigiola telephiifolia was identified as possible lead accumulator and arsenic hyperaccumulator. The determination of water‐soluble arsenic species in the samples studied was carried out by high performance liquid chromatography (HPLC) coupled to ICP‐AES, HG‐ICP‐AES and HG‐AFS, including a prior photo‐oxidation step. The chromatographic methods developed, by anion and cation exchange, allowed us to differentiate up to eleven arsenic species. The sample analysis required the optimization of extraction methods, choosing the microwave assisted extraction (MAE) with deionized water. On the other hand, the stability of total arsenic and water‐soluble arsenic species in algae, both in the solid samples and in the water extracts, was studied, assessing the suitable storage conditions. In the case of plant samples, the application of a multivariate experimental design allowed us to optimize the extraction method and differentiate between the arsenic species present as simple ions of higher mobility and the arsenic more strongly bound to structural components. The presence of arsenite (As(III)) and arsenate (As(V)) was identified in plant samples, as well as monomethylarsonic acid (MMA) and trimethylarsine oxide (TMAO) in some cases. Regarding algae, toxic arsenic species were found in most of them, both As(V) and dimethylarsinic acid (DMA), as well as up to four arsenosugars. These results were discussed according to the current legislation, showing the need to develop specific regulations to control this kind of food products. The determination of lipid‐soluble arsenic species in alga samples was performed by reversed‐phase HPLC coupled to inductively coupled plasma and electrospray mass spectrometry (ICP‐MS and ESI‐MS), in order to establish the structure of these compounds by determining the corresponding molecular masses. For this purpose, it was necessary to develop an extraction method, as well as a clean‐up method of the extracts. The method developed permitted the identification of fourteen lipid‐soluble arsenic compounds in algae, corresponding to three arsenic‐hydrocarbons and eleven arsenosugarphospholipids, as well as two unknown compounds. Accurate mass measurements of the identified compounds were performed by gas chromatography coupled to mass spectrometry (GC‐MS) and high resolution mass spectrometry (HR‐MS)

Single extraction tests for risk assessment of arsenic and heavy metals in polluted soils from mining activities

Se presentan los resultados obtenidos en la aplicación de 3 métodos de extracción simple, para evaluar el grado de movilidad y biodisponibilidad de As, Cd, Cu, Pb y Zn, en suelos afectados por la actividad minera. Los métodos desarollados están basados en el empleo de agentes extractantes de diferente naturaleza y UPS, con determinación por ICP-AES. El estudio estadístico de los resultados obtenidos, permitió establecer el método más adecuado en función del elemento analizado, debido a su diferente asociación a los componentes del suelo

Arsenic speciation in plants by HPLC-(UV)-HG-AFS: Optimisation of the extraction method and application to native plant species from soils polluted by mining activities

Se ha optimizado el método de extracción de especies de arsénico hidrosolubles en muestras de plantas terrestres procedentes de suelos contaminados por actividad minera. La especies extraídas se han determinado mediante HPLC-(UV)-HG-AFS

Application of ultrasound probe sonication for arsenic and heavy metal extraction in soils

Se ha optimizado un método de extracción de arsénico y metales pesados en suelos contaminados procedentes de actividad minera, mediante la aplicación de la sonda de ultrasonidos focalizada

Assessment of total arsenic and arsenic species stability in alga samples and their aqueous extracts

In order to achieve reliable information on speciation analysis, it is necessary to assess previously the species stability in the sample to analyse. Furthermore, in those cases where the sample treatment for species extraction is time-consuming, an assessment of the species integrity in the extracts is of paramount importance. Thus, the present paper reports total arsenic and arsenic species stability in alga samples (Sargassum fulvellum and Hizikia fusiformis), as well as in their aqueous extracts, which were stored in amber glass and polystyrene containers at different temperatures. Total arsenic determination was carried out by inductively coupled plasma atomic emission spectroscopy (ICP-AES), after sample acid digestion in a microwave oven, while arsenic speciation was conducted by anion exchange high performance liquid chromatography on-line coupled to ICPAES, with and without sample introduction by hydride generation (HPLC-ICP-AES and HPLC-HG-ICP-AES), after aqueous microwave-assisted extraction. The results obtained for solid alga samples showed that total arsenic (for Hijiki alga) and arsenic species present (As(V) for Hijiki and NIES No. 9 Sargasso) are stable for at least 12 months when samples are stored in polystyrene containers at +20 ◦C. On the other hand, a different behaviour was observed in the stability of total arsenic and As(V) species in aqueous extracts for both samples, being the best storage conditions for Sargasso extracts a temperature of −18 ◦C and polystyrene containers, under which they are stable for at least 15 days, while Hijiki extracts must be stored in polystyrene containers at +4 ◦C in order to ensure the stability for 10 days

Virtual lab practice for quantitative analysis of metals in polluted soils

En este trabajo se muestra el desarrollo de una práctica de laboratorio virtual de Experimentación Química, para los alumnos de Ciencia Medioambiental de la ETSIC de la UPM

Determination of toxic arsenic species and arsenosugars in edible seaweed by HPLC-(UV)-HG-AFS

Arsenic is a toxic element widely distributed in the environment, and the estimation of its toxicity requires knowledge of the individual arsenic species present in biological materials. Marine algae contribute substantial amounts of arsenic to the human diet in Asian countries, and nowadays their popularity in western countries is increasing due to their high mineral content and their recognized therapeutic properties1. It is known that marine organisms can accumulate considerable arsenic concentrations, up to ug g"1 level, which may be harmful to human beings. In seaweed, the main arsenic species are usually arsenoribosides (arsenosugars), which are considered to be non-toxic to living organisms and can be present at trace levéis, so analysis techniques of high sensitivity are needed to carry out their determination

Determination of soluble toxic arsenic species in alga samples by microwave-assisted extraction and high performance liquid chromatography-hydride generation-inductively coupled plasma-atomic emission spectroscopy

A microwave-based procedure for arsenic species extraction in alga samples (Sargassum fulvellum, Chlorella vulgaris, Hizikia fusiformis and Laminaria digitata) is described. Extraction time and temperature were tested in order to evaluate the extraction efficiency of the process. Arsenic compounds were extracted in 8 ml of deionised water at 90 °C for 5 min. The process was repeated three times. Soluble arsenic compounds extracted accounted for about 78–98% of total arsenic. The results were compared with those obtained in a previous work, where the extraction process was carried out by ultrasonic focussed probe for 30 s. Speciation studies were carried out by high performance liquid chromatography–hydride generation–inductively coupled plasma-atomic emission spectrometry (HPLC-HG-ICP-AES). The chromatographic method allowed us to separate As(III), As(V), monomethylarsonic acid and dimethylarsinic acid in less than 13 min. The chromatographic analysis of the samples allowed us to identify and quantify As(V) in Hizikia sample and Sargasso material, while the four arsenic species studied were found in Chlorella sample. In the case of Laminaria sample, none of these species was identified by HPLC-HG-ICP-AES. However, in the chromatographic analysis of this alga by HPLC-ICP-AES, an unknown arsenic species was detected