Bioavailability, Bioaccesibility of Heavy Metal Elements and Speciation of as in Contaminated Areas of Chile

Studies on the bioavailability of As, Cr, Cu, Pb, Mn and Cd of impacted soils; the As bioaccesibility in the edible parts of carrots, beets and quinoa growing in these polluted soils thought “in vitro” gastrointestinal process; the As speciation both in the edible parts of vegetables and in their gastrointestinal extracts have been performed. Elemental analysis and As speciation have been performed by ICP-MS and LC-ICP-MS, respectively. The high As contents in the interchangeable and oxidized fractions of soil may be responsible for the high As species content in these high vegetables consumption. The Arsenic recovery after the in vitro gastrointestinal digestion was from 98, 90 and 40% for carrots, beets and quinoa, respectively; with no significant transformation of original As species. These studies provide a clearer understanding of the impact that As and other contaminant elements may present in the population of this high polluted Chilean region

Arsenic species-binding proteins in human cardiovascular and muscle tissues

The intracellular As-protein binding in cytosol and methanol–water extract of the auricle and saphene tissues of As impacted people was evaluated by bidimensional size exclusion FPLC-UV-ICP-MS. The fractionation of cytosol using Superdex, Phenomenex and MonoQ HR 5/5 columns, shows that As was distributed in a wide range of contiguous fractions of each column, being 8, 25, 50 % the percentages of As in the collected fractions, respectively. Arsenic a sulphur coelute when FPLC–UV–ICP–MS was applied, which could implicate that As is bound to bio-compounds of different molecular mass through vicinal sulphur groups. The monitoring of S, Cu and P. In the methanol: water extracts a similar study than performed with the cytosol using preparative gel chromatography on Sephadex G-75 and Shephadex G-100 columns. A very low As and protein contain were found in the different fractions of both SEC fractionating series. A similar As–protein association to that found in the cytosol after fractionating with MonoQ HR 5/5 was observed for auricle and saphene. Inorganic and methylated As speciation in the 20 - 26 cytosol fractions obtained within the Phenomenex column was performed by HPLC–ICP–MS using the Hamilton PRP-X100 column. Only As(III) and As(V) were present and the results obtained shows that the As(III)/As(V) ratio is constant in most cases. Direct evidence of the existence of As–binding peptides in auricle and saphene vein from arsenic impacted human beings has have been obtained which was previously reported by means of novo peptide synthesis

Biospeciation of tungsten in the serum of diabetic and healthy rats treated with the antidiabetic agent sodium tungstate

It is known that oral administration of sodium tungstate preserves the pancreatic beta cell function in diabetic rats. Healthy and streptozotocin-induced diabetic rats were treated with sodium tungstate for one, three or six weeks, after which the species of W in serum, were analysed. An increase in serum W with treatment time was observed. After six weeks, the serum W concentration in diabetic rats (70 mg L−1) was about 4.6 times higher than in healthy specimens. This different behaviour was also observed for Cu accumulation, while the Zn pattern follows the contrary. The patterns observed in the retention of Cu and Zn may be attributable to a normalization of glycaemia. The speciation analysis of W was performed using 2D separations, including an immunoaffinity packing and a SEC (Size Exclusion Chromatography) column coupled to an ICP-MS (Inductively Coupled Plasma Mass Spectrometry) for elemental detection. Ultrafiltration data together with SEC-ICP-MS results proved that around 80% of serum W was bound to proteins, the diabetic rats registering a higher W content than their healthy counterparts. Most of the proteinbound W was due to a complex with albumin. An unknown protein with a molecular weight higher tan 100 kDa was also found to bind a small amount of W (about 2%). MALDI-TOF (Matrix-Assisted Laser Desorption Ionization Time-of-Flight) analysis of the desalted and concentrated chromatographic fractions confirmed albumin as the main protein bound to tungstate in rat serum, while no binding to transferrin (Tf) was detected. The interaction between glutathione and W was also evaluated using standard solutions; however, the formation of complexes was not observed. The stability of the complexes between W and proteins when subjected to more stringent procedures, like those used in proteomic methodologies (denaturing with urea or SDS, boiling, sonication, acid media, reduction with -mercaptoethanol (BME) or DTT (dithiotreitol) and alkylation with iodoacetamide (IAA), was also evaluated. Our results indicate that the stability of the complexes between W and proteins is not too high enough to remain unaltered during protein separation by SDS–PAGE in denaturing and reducing conditions. However, the procedures for in-solution tryptic digestion and for ESI-MS analysis in MeOH/H2O/with 0.1% formic acid could be used for protein identification without large loss of binding between W and proteins

Identificación y cuantificación del arsénico unido a las proteínas de tejidos cardiovasculares de pacientes sometidos a cirugía de revascularización coronaria

Introducción: Los efectos de la intoxicación con Arsénico (As) como enfermedades cardiovasculares (CV), pigmentaciones y oclusiones arteriales coronarias están asociados con la ingestión de As inorgánico a través del agua de bebida y a exposiciones ambientales. La unión del As (III) a proteínas y la metilación del As podría ser una primera etapa en el mecanismo de detoxificación. Objetivo: Evaluar la unión de As a proteínas en aurícula derecha y vena safena (VS) en sujetos expuestos de la Región de Antofagasta. Métodos: Se estudió la asociación As-proteína en el citosol de AD y VS de 6 pacientes con enfermedad coronaria grave de la Región de Antofagasta. Para el fraccionamiento del citosol se utilizaron columnas de exclusión molecular de tres diferentes rangos de masas. El perfil del As se detectó por Espectrometría de Masas Inductivamente Acoplado (ICP-MS) y por Espectroscopía Ultra Violeta - Visible de las fracciones moleculares (enlaces As- tiolatos de proteínas). Resultados: En todos los casos el As estuvo ampliamente distribuido en todo el intervalo de fracciones para AUD y VS. Los porcentajes de As colectado en las fracciones de las diferentes columnas usadas fueron 10, 25 y 50%. En la especiación de As en el citosol, por Cromatografía Líquida de Alta Resolución acoplada a la Espectrometría de Masas (IC-HPLC-ICP-MS), solamente se encontró As(III) y As(V) con una distribución Gaussiana para ambas especies, siendo la relación As(III)/As(V) constante para AUD y VS. Conclusión: En los tejidos CV existe asociación As – proteína lo cual podría implicar que el As está unido a biocompuestos de diferente peso molecular a través de grupos sulfhidrilos vecinales. Es probable que el As en AUD y VS se una a fracciones proteicas de masa molecular superior a 80 kDA y a subunidades de la estructura cuaternaria de la proteína nativa

Environmental biochemistry or arsenic species in contamined areas

Arsenic in the Antofagasta Region of Chile is worldwide recognized as highly As contaminated area. Due to its abundance, mainly as As (V) and As (III), certain microorganisms such as several types of bacteria have evolved and developed the necessary genetic components which confer resistance mechanisms. These mechanisms include reactions of reduction, oxidation and methylation. The aim of this work was to study the most relevant arsenic resistant bacteria that exist in highly arsenic contaminated sediments in El Tatio geyser field. This place is a suitable habitat to study the adaptation of endemic species subjected to extreme environmental conditions. All bacterial strains isolated were grown with increasing concentration of arsenate, exhibiting high levels of arsenate resistance ranging from to 5 to 30 mM. Inductively coupled plasma mass spectrometry (ICP-MS) was employed to determine the concentration of As within intact cells of each bacterial strain. Results showed a great accumulation of this element. The separation of bacterial cells into cytoplasmic and membrane fractions were carried out by differential centrifugation in order to know the distribution of arsenic in the different cellular compartments. Most of arsenic was mainly located in the cytoplasmic fraction. Indeed, the optimization of chromatographic methods coupled to ICP-MS allowed us to separate and quantify the different arsenic species as a result of bacterial transformations. The results demonstrated that in half of the isolated strains, between 20-70% of arsenate was reduced to arsenite. Only in one case it was observed the presence of methylated species of arsenic such as DMA and MMA

Thiol-free reducing agents in electrophoretic separations and FASP proteolytic digestions for the analysis of metal-binding proteins

The analysis of the complexes between metal-based chemotherapeutic drugs and proteins in biological samples, such as cisplatin or oxaliplatin, can be a challenge due to metal strong reactivity towards S-donor molecules such as dithiothreitol (DTT) or b-mercaptoethanol (BME), usually employed as reducing agents in electrophoretic separations and proteolytic digestions for LC–MS/MS analysis. This protocol describes the use of the thiol-free reducing trialkylphosphines, such as tributylphosphine (TBP) and tris(2-carboxyethyl)phosphine (TCEP) as suitable reagents for the preservation of the metal-protein complexes during OFFGEL-IEF and SDS-PAGE separations, respectively. Moreover, the filter-aided sample preparation (FASP) method is presented as an advantageous option to perform tryptic in-solution digestions of metal–protein complexes in combination with OFFGEL-IEF separations

Total and main as species present in cardiovascular tirrues of people living in as contaminated areas

The concentration levels of As in the Chilean II Region of Antofagasta produces non cancer health outcomes such as cardiovascular diseases and in last term heart attack. On this study, the determination of total As content and main inorganic and organoarsenic species found in three heart tissues (auricle, mammary artery and fat) and the saphene vein of people living in the Chilean II Region, suffering coronary thrombosis has been carried out. Comparison with similar tissues of patients from other non-contaminated areas has been undertaken. The extraction of As species occurred in methanol: water (1:1) and As species determination have been used the tandem HPLC-ICP-MS using the Hamilton PRP X100 anion column. For total As determination has been performed by HG-AAS and ICP-MS. The auricle and in less extend the saphene support the higher As concentration (mean values of 7.7 and 2.5 µg g-1, respectively), being As(III) the predominant species. Methylarsonate (MA) and dimethylarsinate (DMA) is not a favoured mechanism. The presence of high total As and high As(III) species content in the auricle and saphene of more contaminated people, the damage found in the saphene tissue and the global characteristics of the people under study in which the As stigmas are present in all of them, suggests that As could be involved in the cardiovascular diseases. e-mail: ipizarro @uantof.cl

Estrategias bioanalíticas en estudios metalómicos de fármacos de platino

En este artículo se muestra el potencial que presentan las modernas estrategias bioanalíticas en estudios metalómicos de fármacos de platino. La combinación de técnicas de separación multidimensionales cromatográficas y/o electroforéticas con la espectrometría de masas atómica ICP-MS y molecular ESI-MS/MS se presenta como una valiosa alternativa en este tipo de estudios

Integrated transcriptomics and metabolomics analysis reveals the biomolecular mechanisms associated to the antitumoral potential of a novel silver‑based core@shell nanosystem

A combination of omics techniques (transcriptomics and metabolomics) has been used to elucidate the mechanisms responsible for the antitumor action of a nanosystem based on a Ag core coated with mesoporous silica on which transferrin has been anchored as a targeting ligand against tumor cells (Ag@MSNs-Tf). Transcriptomics analysis has been carried out by gene microarrays and RT-qPCR, while high-resolution mass spectrometry has been used for metabolomics. This multiomics strategy has enabled the discovery of the effect of this nanosystem on different key molecular pathways including the glycolysis, the pentose phosphate pathway, the oxidative phosphorylation and the synthesis of fatty acids, among others