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This research explores the possibilities and limitations of WD-XRF, applied as a method for quantification of 20 elements in plant material, using spiked cellulose standards for calibration. Three different analytical methods were investigated: 1) standards created from pure spiked cellulose; 2) spiked cellulose mixed with 20% of binder and 3) spiked cellulose applied as a thin layer on an inert carrier. Sensitivity, linearity, limit of detection, limit of quantification, repeatability, intralaboratory reproducibility, and accuracy were determined and compared. The accuracy of the investigated methods was tested by analysis of standard reference materials and comparison with other routinely used analytical techniques (ICP-OES and ICP-MS). The comparison included real plant samples which were collected from the environment characterized by different pollution levels. The accuracy of the semiquantitative standardless method was also considered and compared with other investigated methods. Tested methods can be very precise, with good intralaboratory reproducibility over wide linear range. Supplementary material: [https://cherry.chem.bg.ac.rs/handle/123456789/4615] This is the peer-reviewed version of the article: Orlić, J., Gržetić, I., Goessler, W., Braeuer, S., Čáslavský, J., Pořízka, J.,& Ilijević, K.. (2021). Artificial cellulose standards as calibration standards for wavelength-dispersive X-ray fluorescence analysis of elements in plant samples. in Nuclear Instruments and Methods in Physics Research Section B: Beam Interactions with Materials and Atoms Elsevier., 502, 106-117. [https://doi.org/10.1016/j.nimb.2021.06.012]

Trace elements in aquatic environments pose a risk to biological communities; this study investigates the total concentrations of arsenic (As), silver (Ag), cadmium (Cd), cobalt (Co), chromium (Cr), copper (Cu), mercury (Hg), nickel (Ni), selenium (Se), lead (Pb), and zinc (Zn) within muscle, carapace, liver, and adipose tissues of diamondback terrapins and common snapping turtles in New Jersey. The effects of tissue type, sex, size, and location upon trace element accumulation were studied. The data obtained indicates that within diamondback terrapins and common snapping turtles, trace element accumulations displayed a significant difference among tissue types and sex (p

Amanita muscaria, also known as the fly agaric mushroom, can accumulate vanadium (V), with up to several hundred mg V kg−1 dry mass. It is long known that V is present in A. muscaria as a complex called amavadin, but methods for the investigation of the distribution and biosynthesis of amavadin in mushrooms are missing. Here, we describe the development of the first sensitive method for the determination of amavadin and other V-containing compounds in environmental samples by employing high performance liquid chromatography (HPLC) and inductively coupled plasma mass spectrometry (ICPMS). A strong anion-exchange column serves as the stationary phase, and the mobile phase consists of an aqueous ammonium citrate buffer and ethylenediaminetetraacetate (EDTA). The concentration and pH of the mobile phase as well as the column temperature were evaluated to optimize the separation. With the final method, amavadin is eluted in less than 17 minutes, and its limit of detection is 0.05 μg V L−1. Moreover, the compound's two isomers are separated from each other and can be quantified independently. The method was applied to extracts of fruit-body samples of A. muscaria. The extraction efficiency was 74 ± 12%, and amavadin accounted for 75–96% of the extracted V. In addition, significant concentrations of other V species could be detected, which have never been described before. Our results demonstrate that V speciation in mushrooms is more complex than assumed until now and that more in-depth investigations on this matter are needed. The developed method enables the investigation of organic and inorganic V species in the environment, even at low concentrations.

This work describes the development of a novel method for quantitative mapping of Hg and Se in mushroom fruit body tissues with laser ablation coupled to inductively coupled plasma-mass spectrometry (LA-ICP-MS). Different parameters of the protocol for preparation of the standards used for quantification via external calibration were assessed, e.g., the dissolution temperature of gelatin standards and the addition of chitosan and l-cysteine as additives to the gelatin-based calibration droplets to better match the sample matrix. While chitosan was not suited for this purpose, the presence of l-cysteine considerably improved the figures of merit of the calibration, leading to limits of detection of 0.006 and 0.3 µg g−1 for Hg and Se, respectively, at a pixel size of 20 × 20 µm. Further, an in-house reference material, ideally suited for the validation of the method for application to mushroom samples, was successfully prepared from a paste of Boletus edulis. The newly developed method was used to investigate the distribution of Hg and Se in tissue sections of five porcini mushroom individuals of three different species (Boletus edulis, Boletus aereus, and Boletus pinophilus) and one sample of a parasol mushroom (Macrolepiota procera). For one sample, additional areas were ablated at higher spatial resolution, with a laser spot size down to 5 µm, which allows a detailed investigation of the spatial distribution of Hg and Se in mushrooms. Graphical abstract

ChemBioChem 22(4), 652 - 656 (2020). doi:10.1002/cbic.202000600, Broad substrate tolerance and excellent regioselectivity, as well as independence from sensitive cofactors have established benzoic acid decarboxylases from microbial sources as efficient biocatalysts. Robustness under process conditions makes them particularly attractive for preparative‐scale applications. The divalent metal‐dependent enzymes are capable of catalyzing the reversible non‐oxidative (de)carboxylation of a variety of electron‐rich (hetero)aromatic substrates analogously to the chemical Kolbe‐Schmitt reaction. Elemental mass spectrometry supported by crystal structure elucidation and quantum chemical calculations verified the presence of a catalytically relevant Mg$^{2+}$ complexed in the active site of 2,3‐dihydroxybenoic acid decarboxylase from Aspergillus oryzae (2,3‐DHBD_Ao). This unique example with respect to the nature of the metal is in contrast to mechanistically related decarboxylases, which generally have Zn$^{2+}$ or Mn$^{2+}$ as the catalytically active metal., Published by Wiley-VCH, Weinheim

Fruit-bodies of six Thelephora species (Fungi, Basidiomycota, Thelephoraceae) were analyzed for their trace element concentrations. In Thelephora penicillata, extremely high concentrations of Cd and As were found, followed by highly elevated concentrations of Cu and Zn. The highest accumulation ability was found for Cd with a mean concentration of 1.17 ± 0.37 g kg

Thelephora penicillata is an ectomycorrhizal mushroom that can accumulate extraordinarily high concentrations of Cd, As, Cu, and Zn in its fruit-bodies. To better understand its element accumulation ability, we compared the element concentrations in T. penicillata with 10 distinct ectomycorrhizal mushroom species growing at the same site (Karlina Pila, Czech Republic). On average, T. penicillata accumulated 330, 2130, 26, and 4 times more Cd, As, Cu, and Zn, respectively, than other mushrooms. Size-exclusion chromatography and an electrophoretic analysis of T. penicillata cell extracts indicate that intracellular Cd may be present mainly in1 kDa, presumably compartmentalized, Cd species, and partially binding with 6-kDa cysteinyl-containing peptide(s) resembling metallothioneins. The cadmium isotopic composition of mushroom fruit-bodies, soil digests, and soil extracts was investigated by thermal ionization mass spectrometry (TIMS) with double spike correction. The isotopic composition (δ

Arsenic can be toxic to living organisms, depending not only on the concentration, but also its chemical form. The aim of this study was to determine arsenic concentrations and perform arsenic speciation analysis for the first time in honeybees, to evaluate their potential as biomonitors. Highest arsenic concentrations were determined in the vicinity of coal fired thermal power plants (367 µg kg

The concentrations of major and trace elements in Argentinean commercial powdered milk samples were determined with inductively coupled plasma mass spectrometry. Also the daily intake (DI) was calculated for adults and infants. The concentrations of B, Mg, Na, K and Ca were significantly higher in skimmed milk. Cu, Mo, Fe, Mn and Zn were significantly higher in infant formula. All the DIs were below the tolerable upper intake levels. The mean As concentration (26.0 ± 8.6 ng/g) in the powdered skimmed milk samples was slightly higher than in the others. Mean Pb concentrations ranged from 4.1 ± 2.1 to 13.5 ± 8.2 ng/g. The highest mean U concentration was 7.8 ± 2.6 ng/g for whole milk. This study contributes to the knowledge of major and trace elements in powdered milk and its contribution to the diet in Argentina. Fil: Arellano, Flavia Elisa. Universidad de Buenos Aires. Facultad de Ciencias Veterinarias. Centro de Estudios Transdisciplinarios del Agua; Argentina Fil: Braeuer, Simone. University of Graz; Austria Fil: Fernandez Cirelli, Alicia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Unidad Ejecutora de Investigaciones en Producción Animal. Universidad de Buenos Aires. Facultad de Ciencias Veterinarias. Unidad Ejecutora de Investigaciones en Producción Animal; Argentina Fil: Goessler, Walter. University of Graz; Austria Fil: Pérez Carrera, Alejo Leopoldo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Unidad Ejecutora de Investigaciones en Producción Animal. Universidad de Buenos Aires. Facultad de Ciencias Veterinarias. Unidad Ejecutora de Investigaciones en Producción Animal; Argentina

Polcalcins are important respiratory panallergens, whose IgE-binding capacity depends on the presence of calcium. Since specific immunotherapy is not yet available for the treatment of polcalcin-sensitized patients, we aimed to develop a molecule for efficient and safe immunotherapy. We generated a hypoallergenic variant of the grass pollen polcalcin Phl p 7 by introducing specific point mutations into the allergen’s calcium-binding regions. We thereby followed a mutation strategy that had previously resulted in a hypoallergenic mutant of a calcium-binding food allergen, the major fish allergen parvalbumin. Dot blot assays performed with sera from Phl p 7-sensitized patients showed a drastically reduced IgE reactivity of the Phl p 7 mutant in comparison to wildtype Phl p 7, and basophil activation assays indicated a significantly reduced allergenic activity. Rabbit IgG directed against mutant rPhl p 7 blocked patients’ IgE binding to wildtype Phl p 7, indicating the mutant’s potential applicability for immunotherapy. Mass spectrometry and circular dichroism experiments showed that the mutant had lost the calcium-binding capacity, but still represented a folded protein. In silico analyses revealed that the hypoallergenicity might be due to fewer negative charges on the molecule’s surface and an increased molecular flexibility. We thus generated a hypoallergenic Phl p 7 variant that could be used for immunotherapy of polcalcin-sensitized individuals.

Background: Arsenic (As) is considered to be toxic for humans, the main routes of exposure being through drinking water and the diet. Once ingested, inorganic arsenic can be methylated sequentially to monomethyl and dimethyl arsenicals. Several factors can affect both As exposure and methylation efficiency. Objectives: To describe the urinary concentrations of the different As species and evaluate the methylation effi-ciency during pregnancy, as well as their associated factors in a birth cohort of pregnant Spanish women. Methods: Participants in this cross-sectional study were 1017 pregnant women from two areas of Spain who had taken part in the INMA (Environment and Childhood) project (2003 & ndash;2008). Total As (organic and inorganic compounds) and its main metabolites (monomethylarsonic acid, [MMA], dimethylarsinic acid, [DMA], inorganic As [iAs]) and arsenobetaine [AB]) were measured in urine samples collected during the first trimester. Socio-demographic and dietary information was collected through questionnaires. Multivariate linear regression models were used to explore the association between As species concentrations and covariates. Arsenic methylation efficiency was determined through the percentages of the metabolites and using As methylation phenotypes, obtained from principal component analysis. Results: Median urine concentrations were 33.0, 21.6, 6.5, 0.35 and 0.33 lig/g creatinine for total As, AB, DMA, MMA and iAs, respectively. Daily consumption of rice and seafood during the first trimester of pregnancy were positively associated with the concentration of As species (i.e., beta [CI95%] = 0.36 [0.09, 0.64] for rice and iAs, and 1.06 [0.68, 1.44] for seafood and AB). TAs, AB and iAs concentrations, and DMA and MMA concentrations were associated with legume and vegetable consumption, respectively. The medians of the percentage of As metabolites were 89.7 for %DMA, 5.1 for %MMA and 4.7 for %iAs. Non-smoker women and those with higher body mass index presented a higher methylation efficiency (denoted by a higher %DMA and lower %MMA). Discussion: Certain dietary, lifestyle, and environmental factors were observed to have an influence on both As species concentrations and methylation efficiency in our population. Further birth cohort studies in low exposure areas are necessary to improve knowledge about arsenic exposure, especially to inorganic forms, and its potential health impact during childhood. This study was supported by grants from Instituto de Salud Carlos III, Spain [FIS-FEDER: 07/0314; FIS 11/01007, 13/1944, 16/1288, 19/1338; Miguel Servet-FEDER: CP15/0025, CPII20/00006; FIS-FSE: 17/00260; Miguel Servet-FSE: MS15/0025, MS20/0006; PI06/0867 and FIS-PI18/01142 incl. FEDER funds] , Generalitat Valenciana, Spain BEST/2020/059, Department of Health of the Basque Government, Spain (2005111093) , Provincial Government of Gipuzkoa (DFG06/002) , and annual agreements with the municipalities in the study area (Zumarraga, Urretxu, Legazpi, Azkoitia, Azpeitia and Beasain) .

The As concentrations, along with 34 other elements, and the As speciation were investigated in wild-grown samples of the parasitic mushroom Tolypocladium ophioglossoides with inductively coupled plasma mass spectrometry (ICPMS) and high performance liquid chromatography coupled to ICPMS. The As concentrations were 0.070–3.44 mg kg−1 dry mass. More remarkable was the As speciation, where up to 56% of the extracted As were found to be an unknown As species, which was marginally retained under anion- and also cation-exchange conditions. After testing several different chromatographic settings, the compound was finally isolated and identified as 2-(sulfoxyethyl) trimethylarsonium ion (in short: arsenocholine-O-sulfate) with high resolution mass spectrometry. The compound was synthesized and further quantified in all investigated samples via ion-pair chromatography coupled to ICPMS. In addition to the high abundance of arsenocholine-O-sulfate in T. ophioglossoides, small amounts of this As species were also detected in one sample of the host mushroom, Elaphomyces asperulus. In a sample of another parasitic mushroom, Ophiocordyceps sinensis, arsenocholine-O-sulfate could not be detected, but the main species was another unknown compound that was oxidized to inorganic As(V) with hydrogen peroxide. This is the first discovery of arsenocholine-O-sulfate in nature. It is possible that it is present in many other organisms, at least in low concentrations, and just has not been detected there yet because of its unusual chromatographic behavior. The existence of arsenocholine-O-sulfate brings up questions again about the biotransformation pathways of As in the environment and the specific behavior of fungi.

The arsenic speciation was determined in macrofungi of the Ramaria genus with HPLC coupled to inductively coupled plasma mass spectrometry. Besides arsenic species that are already known for macrofungi, like arsenobetaine or arsenocholine, two compounds that were only known from marine samples so far (trimethylarsoniopropanate and dimethylarsinoylacetate) were found for the first time in a terrestrial sample. An unknown arsenical was isolated and identified as homoarsenocholine. This could be a key intermediate for further elucidation of the biotransformation mechanisms of arsenic.

The edible ink stain bolete (Cyanoboletus pulverulentus) was found to hyperaccumulate arsenic. We analyzed 39 individual collections determined as C. pulverulentus, mostly from the Czech Republic. According to our results, concentrations of arsenic in C. pulverulentus fruit-bodies may reach 1300 mg kg(−1) dry weight. In most collections, data for total and bioavailable arsenic in underlying soils were collected but no significant correlation between the soil arsenic content and arsenic concentrations in the associated fruit-bodies was found. Within the fruit-bodies, we found the majority of arsenic accumulated in the hymenium. Besides occasional traces of methylarsonic acid (MA), the arsenic speciation in all mushroom samples consisted solely of dimethylarsinic acid (DMA) and no inorganic arsenic was detected. Because of the carcinogenic potential of DMA, C. pulverulentus should not be recommended as an edible mushroom and its consumption should be restricted.

The total arsenic concentration and the arsenic speciation in urine and feces samples of the two Giant Pandas living at Vienna zoo and of their feed, bamboo, were determined with ICPMS and HPLC-ICPMS. Urine was the main excretion route and accounted for around 90% of the ingested arsenic. The urinary arsenic concentrations were very high, namely up to 179 μg/L. Dimethylarsinic acid (DMA) was the dominating arsenic compound in the urine samples and ranged from 73 to 92% of the total arsenic, which is unusually high for a terrestrial mammal. The feces samples contained around 70% inorganic arsenic and 30% DMA. The arsenic concentrations in the bamboo samples were between 16 and 920 μg/kg dry mass. The main arsenic species in the bamboo extracts was inorganic arsenic. This indicates that the Giant Panda possesses a unique way of very efficiently methylating and excreting the provided inorganic arsenic. This could be essential for the survival of the animal in its natural habitat, because parts of this area are contaminated with arsenic.

We report the first halogen speciation analysis study by high performance liquid chromatography coupled with inductively coupled plasma tandem mass spectrometry (HPLC-ICPMS/MS) in the fruiting bodies of various mushroom species. Non-targeted speciation analysis revealed the occurrence of dichloroacetic acid (DCAA) in the edible mushroom Russula nigricans. Multiple samples of this mushroom (n = 5) collected from different geographic non-industrial regions in two different countries confirmed the consistent presence of this species at a relatively narrow concentration range (23–37 mg kg−1), whereas no other chlorinated acetic acid (e.g. chloroacetic acid and trichloroacetic acid) was detected. Neither DCAA nor any other chlorinated acetic acid were detected in any of the other mushroom species investigated in the present study, including seven different mushroom species of the same genus Russula, even though all mushrooms were collected from the same non-industrial geographic regions. Together with the previously reported biological activities of DCAA, these findings collectively suggest biosynthesis of this compound as an explanation for its dominant presence in R. nigricans, and constitute the first example of the dominant natural occurrence of this compound over other chlorinated acetic acids in a living organism. This may warrant a change in our view of the occurrence of dichloroacetic acid in nature, where primarily considered as a pollutant arising from water disinfection. Version of record

Biodegradable magnesium implants are under investigation because of their promising properties as medical devices. For enhancing the mechanical properties and the degradation resistance, rare earth elements are often used as alloying elements. In this study Mg10Gd pins were implanted into Sprague–Dawley® rats. The pin volume loss and a possible accumulation of magnesium and gadolinium in the rats’ organs and blood were investigated in a long-term study over 36 weeks. The results showed that Mg10Gd is a fast disintegrating material. Already 12 weeks after implantation the alloy is fragmented to smaller particles, which can be found within the intramedullary cavity and the cortical bones. They disturbed the bone remodeling until the end of the study. The results concerning the elements’ distribution in the animals’ bodies were even more striking, since an accumulation of gadolinium could be observed in the investigated organs over the whole time span. The most affected tissue was the spleen, with up to 3240 μg Gd/kg wet mass, followed by the lung, liver and kidney (up to 1040, 685 and 207 μg Gd/kg). In the brain, muscle and heart, the gadolinium concentrations were much smaller (less than 20 μg/kg), but an accumulation could still be detected. Interestingly, blood serum samples showed no accumulation of magnesium and gadolinium. This is the first time that an accumulation of gadolinium in animal organs was observed after the application of a gadolinium-containing degradable magnesium implant. These findings demonstrate the importance of future investigations concerning the distribution of the constituents of new biodegradable materials in the body, to ensure the patients’ safety. Statement of Significance In the last years, biodegradable Mg alloys are under investigation due to their promising properties as orthopaedic devices used for bone fracture stabilization. Gadolinium as Rare Earth Element enhances the mechanical properties of Mg-Gd alloys but its toxicity in humans is still questionable. Up to now, there is no study investigating the elements’ metabolism of a REE-containing Magnesium alloy in an animal model. In this study, we examined the gadolinium distribution and accumulation in rat organs during the degradation of Mg10Gd. Our findings showed that Gd is accumulating in the animal organs, especially in spleen, liver and kidney. This study is of crucial benefit regarding a safe application of REE-containing Magnesium alloys in humans.

Since the discovery of the "fish arsenic" arsenobetaine in a mushroom, arsenic species have been studied in all kinds of mushrooms, with very diverse results, for more than 20 years. Depending on the fungal species, inorganic arsenic, methylarsonic acid, dimethylarsinic acid, arsenobetaine and a couple of other arsenic-containing compounds have been detected in all kinds of concentrations in natural samples. A rule to predict the arsenic speciation of a new mushroom is yet to be found. Some research has been devoted to the question about the origin of the different arsenic species in mushrooms. It is still not clear whether mushrooms are able to transform arsenic by themselves or if they just take it up from the environment. For the determination of the arsenic species in mushrooms, high performance liquid chromatography coupled to inductively coupled plasma mass spectrometry is used most of the times, but a few alternative approaches exist. This review will give an overview of these different techniques and critically discuss them. The arsenic species that have been found in mushrooms so far will be summarized, and the work that has been devoted to unraveling the origins of these compounds will be discussed in depth. The interesting phenomenon of arsenic hyperaccumulation by mushrooms will be reviewed as well.

The Violet Crown Cup, Sarcosphaera coronaria, is a rather inconspicuous mushroom, but with an interesting and unresolved mystery. In earlier days, the mushroom was considered edible, but several poisonings were reported in the early 20th century. The reason for the seemingly sporadic toxicity of S. coronaria is still unknown. One possible explanation is arsenic, since Crown Cups can take up high amounts of this element. We investigated the arsenic concentration and arsenic speciation in S. coronaria with inductively coupled plasma mass spectrometry (ICPMS) and HPLC coupled to ICPMS and found up to incredible 0.9% As (dry mass). Most of it was present as methylarsonic acid (MA), a less toxic form of this element. However, low concentrations of the highly toxic methylarsonous acid [MA (III)] were also detected. The amounts were too low to pose an acute risk for consumers, but the concentration of MA (III) significantly increased during simulated gastric digestion. We could not unambiguously identify arsenic as the toxic constituent of S. coronaria, but we demonstrated that the extremely toxic MA (III) can be formed under certain circumstances, which should be carefully investigated in future.

Selenium (Se) and sulfur (S) speciation analysis in edible and medicinal Se enriched P. pulmonarius extracts was performed. Mycelium, colonized substrate, and fruiting bodies at different harvesting times were analyzed using ion-pairing reversed-phase chromatography coupled to an ICPMS/MS detector. Extraction efficiencies in enzymatically digested and aqueous extracts were between 45.3 and 109% for Se, depending on the sample type. Selenomethionine (Se-Met) was found to be the major Se-compound, together with a number of unknown Se-species. Cystine (Cys2), methionine (Met), and sulfate were also detected and quantified in all samples. Most of the Se-Met (84.0%) and Met (75.8%) were found to be in free form in the fruiting body, in contrast with the mycelium where 53.4% of Se-Met and 80.5% of Met is incorporated into proteins.

Collections of Cystoderma carcharias sporocarps were sampled from clean and smelter-polluted sites and analyzed for Ag, As, Cd, Cu, Pb, Se, and Zn contents. Concentrations of all elements were significantly higher in samples from the smelter-polluted area. Except for As and Pb, all elements were effectively accumulated in the sporocarps at both clean and polluted sites. With the highest concentration of 604 mg Cd kg−1, C. carcharias can be considered as Cd hyperaccumulator. As revealed by HPLC-ICPQQQMS analysis, the As species in sporocarps from clean and polluted areas involved besides the major arsenobetaine a variety of known and unknown arsenicals; the occurrence of dimethylarsinoylacetate and trimethylarsoniopropionate is reported for the first time for gilled fungi (Agaricales). Size-exclusion chromatography of C. carcharias extracts supported by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and RP-HPLC data indicated that detoxification of intracellular Cd and Cu may largely rely on metallothioneins (MT) or MT-like peptides, not phytochelatins.

Total arsenic and water-soluble arsenic species were determined in canned fish bought in Graz (n = 6) and Prague (n = 8) with inductively-coupled plasma mass spectrometry and anion-exchange liquid chromatography. A difference in total arsenic content between samples from Prague (mean 3.4; standard deviation (s.d.) 1.2 mg kg–1) and Graz (mean 2.5; s.d. 0.7 mg kg–1) was not significant, however Prague samples differed in abundance of arsenobetaine (mean 83% of extractable arsenic; s.d. 16%) vs. Graz samples (mean 66%; s.d. 10%). No differences in total arsenic were caused by fish species (mackerel, sardine, herring, tuna) and origin (Northeast and Eastern-central Atlantic).

Excessive arsenic content in drinking water poses health risks to millions of people worldwide. Inorganic arsenic (iAs) in groundwater exceeding the 10 μg/l maximum contaminant level (MCL) set by the World Health Organization (WHO) is characteristic for intermediate-depth aquifers over large areas of the Pannonian Basin in Central Europe. In western Romania, near the border with Hungary, Arad, Bihor, and Timis counties use drinking water coming partially or entirely from iAs contaminated aquifers. In nearby Arad and Bihor counties, more than 45,000 people are exposed to iAs over 10 μg/l via public drinking water sources. However, comparable data are unavailable for Timis County. To begin to address this data gap, we determined iAs in 124 public and private Timis County drinking water sources, including wells and taps, used by pregnant women participating in a case-control study of spontaneous loss. Levels in water sources were low overall (median = 3.0; range = < 0.5–175 μg/l), although higher in wells (median = 3.1, range = < 0.5–175) than in community taps (median = 2.7, range = < 0.5–36.4). In a subsample of 20 control women we measured urine biomarkers of iAs exposure, including iAs (arsenite and arsenate), dimethylarsinic acid (DMA), and methylarsonic acid (MMA). Median values were higher among 10 women using iAs contaminated drinking water sources compared to 10 women using uncontaminated sources for urine total iAs (6.6 vs. 5.0 μg/l, P = 0.24) and DMA (5.5 vs. 4.2 μg/l, P = 0.31). The results suggested that the origin of urine total iAs (r = 0.35, P = 0.13) and DMA (r = 0.31, P = 0.18) must have been not only iAs in drinking-water but also some other source. Exposure of pregnant women to arsenic via drinking water in Timis County appears to be lower than for surrounding counties; however, it deserves a more definitive investigation as to its origin and the regional distribution of its risk potential.