Two-Photon Microscopy and Spectroscopy Studies to Determine the Mechanism of Copper Oxide Nanoparticle Uptake by Sweetpotato Roots during Postharvest Treatment.

The interaction of engineered nanoparticles with plant tissues is still not well understood. There is a lack of information about the effects of curing (postharvest treatment) and lignin content on copper uptake by sweetpotato roots exposed to copper-based nanopesticides. In this study, Beauregard-14 (lower lignin) and Covington (higher lignin) varieties were exposed to CuO nanoparticles (nCuO), bulk CuO (bCuO), and CuCl2 at 0, 25, 75, and 125 mg/L. Cured and uncured roots were submerged into copper suspensions/solutions for 30 min. Subsequently, root segments were sliced for imaging with a 2-photon microscope, while other root portions were severed into periderm, cortex, perimedulla, and medulla. They were individually digested and analyzed for Cu content by inductively coupled plasma-optical emission spectroscopy. Microscopy images showed higher fluorescence in periderm and cortex of roots exposed to nCuO, compared with bCuO. At 25 mg/L, only bCuO showed higher Cu concentration in the periderm and cortex of Beauregard-14 (2049 mg/kg and 76 mg/kg before curing; 6769 mg/kg and 354 mg/kg after curing, respectively) and in cortex of Covington (692 mg/kg before curing and 110 mg/kg after curing) compared with controls ( p ≤ 0.05). In medulla, the most internal tissue, only Beauregard-14 exposed to 125 mg bCuO/L showed significantly ( p ≤ 0.05) more Cu before curing (17 mg/kg) and after curing (28 mg/kg), compared with control. This research has shown that the 2-photon microscope can be used to determine CuO particles in nondyed plant tissues. The lack of Cu increase in perimedulla and medulla, even in roots exposed to high CuO concentrations (125 mg/L), suggests that nCuO may represent a good alternative to protect and increase the shelf life of sweetpotato roots, without exposing consumers to excess Cu.

Spectroscopic verification of zinc absorption and distribution in the desert plant Prosopis juliflora-velutina (velvet mesquite) treated with ZnO nanoparticles.

The impact of metal nanoparticles (NPs) on biological systems, especially plants, is still not well understood. The aim of this research was to determine the effects of zinc oxide (ZnO) NPs in velvet mesquite (Prosopis juliflora-velutina). Mesquite seedlings were grown for 15 days in hydroponics with ZnO NPs (10 nm) at concentrations varying from 500 to 4000 mg L(-1). Zinc concentrations in roots, stems and leaves were determined by inductively coupled plasma optical emission spectroscopy (ICP-OES). Plant stress was examined by the specific activity of catalase (CAT) and ascorbate peroxidase (APOX); while the biotransformation of ZnO NPs and Zn distribution in tissues was determined by X-ray absorption spectroscopy (XAS) and micro X-ray fluorescence (µXRF), respectively. ICP-OES results showed that Zn concentrations in tissues (2102 ± 87, 1135 ± 56, and 628 ± 130 mg kg(-1) d wt in roots, stems, and leaves, respectively) were found at 2000 mg ZnO NPs L(-1). Stress tests showed that ZnO NPs increased CAT in roots, stems, and leaves, while APOX increased only in stems and leaves. XANES spectra demonstrated that ZnO NPs were not present in mesquite tissues, while Zn was found as Zn(II), resembling the spectra of Zn(NO(3))(2). The µXRF analysis confirmed the presence of Zn in the vascular system of roots and leaves in ZnO NP treated plants.

Effect of copper oxide nanoparticles on two varieties of sweetpotato plants.

Little information is available on the interaction of CuO nanoparticles (nCuO) with tuberous roots. In this study, Beauregard-14 (B-14, low lignin) and Covington (COV, high lignin) sweetpotato varieties were cultivated until maturity in soil amended with nCuO, bulk copper oxide (bCuO) and CuCl2 at 25-125 mg/kg. The Cu treatments had no significant influence on chlorophyll content. Gas exchange parameters were not affected in B-14. In COV, however, at 125 mg/kg treatments, bCuO reduced the intercellular CO2 (11%), while CuCl2 increased it by 7%, compared with control (p ≤ 0.035). At 25 mg/kg nCuO increased the length of COV roots (20.7 ± 2.0 cm vs. 14.6 ± 0.8 cm, p ≤ 0.05). In periderm of B-14, nCuO, at 125 mg/kg, increased Mg by 232%, while the equivalent concentration of CuCl2 reduced P by 410%, compared with control (p ≤ 0.05). The data suggest the potential application of nCuO as nanofertilizer for sweetpotato storage root production.

Modeling the adsorption of Cr(III) from aqueous solution onto Agave lechuguilla biomass: study of the advective and dispersive transport.

The biosorption of Cr(III) onto packed columns of Agave lechuguilla was analyzed using an advective-dispersive (AD) model and its analytical solution. Characteristic parameters such as axial dispersion coefficients, retardation factors, and distribution coefficients were predicted as functions of inlet ion metal concentration, time, flow rate, bed density, cross-sectional column area, and bed length. The root-mean-square-error (RMSE) values 0.122, 0.232, and 0.285 corresponding to the flow rates of 1, 2, and 3 (10(-3))dm3min(-1), respectively, indicated that the AD model provides an excellent approximation of the simulation of lumped breakthrough curves for the adsorption of Cr(III) by lechuguilla biomass. Therefore, the model can be used for design purposes to predict the effect of varying operational conditions.

Biochemical and physiological effects of copper compounds/nanoparticles on sugarcane (Saccharum officinarum).

The widespread use of copper based nanomaterials has been accompanied by an increasing interest in understanding their potential risks. It is essential to understand the effects of these nanoparticles on edible crops by performing long-term experiments at relevant exposure concentrations. Sugarcane is the source of 70% of the world's sugar supply and the widespread use of refined sugar and the consumption of raw sugarcane can provide a route for nanoparticles to enter the food supply. In order to evaluate the biochemical and physiological effects of copper nanoparticle exposure, sugarcane was grown for one year in soil amended with 20, 40, and 60 mg/kg of Kocide 3000 (a copper based fungicide), copper metal nanoparticles, micro-sized CuO, and CuCl2. The results show that stress indicators such as catalase and ascorbic peroxidase enzymatic activity in the sugarcane plant were activated by all the copper based materials at different concentrations. Sugarcane plants exposed to nearly all copper treatments showed dosage dependent increases in copper concentrations in root tissues. Translocation of copper to aerial tissues was minimal, with copper concentrations not being significantly different from controls. In addition, Chlorophyll A content was higher in plants treated with Kocide 3000 at 20 and 60 mg/kg, µCuO at 20 mg/kg, and CuCl2 at 20 and 60 mg/kg. To our knowledge, this is the first report on the effects of nano-copper compounds in sugarcane crop.

Speciation and uptake of arsenic accumulated by corn seedlings using XAS and DRC-ICP-MS.

ICP-MS was used to investigate the uptake of As(III) and As(V) from hydroponics growth media by corn seedlings. It was found that arsenic uptake by the plant roots for the arsenic(V) and arsenic(III) treatments were 95 and 112 ppm, respectively. However, in the shoots of the arsenic (V) treatments had 18 ppm whereas arsenic(III) treatments had 12 ppm. XANES studies showed that As for both treatments arsenic was present as a mixture of an As(III) sulfur complex and an As(V) oxygen complex. The XANES data was corroborated by the EXAFS studies showing the presence of both oxygen and sulfur ligands coordinated to the arsenic. Iron concentrations were found to increase by 4 fold in the As(V) contaminated growth media and 7 fold in the As(III) treatment compared to the control iron concentration of 500 ppm. Whereas, the total iron concentration in the shoots was found to decrease by approximately the same amount for both treatments from 360 ppm in the control to approximately 125 ppm in both arsenic treatments. Phosphorus concentrations were found to decrease in both the roots and shoots compared to the control plants. The total sulfur in the roots was found to increase in the arsenic(III) and arsenic(V) treatments to 560 ppm and 800 ppm, respectively, compared to the control plants 358 ppm. In addition, the total sulfur in shoots of the plants was found to remain relatively constant at approximately 1080 ppm. The potassium concentrations in the plants were found to increase in the roots and decrease in the shoots.

Use of X-ray absorption spectroscopy and biochemical techniques to characterize arsenic uptake and reduction in pea (Pisum sativum) plants.

Arsenite (As(III)) and arsenate (As(V)) uptake by peas was investigated using inductively coupled plasma/optical emission spectroscopy (ICP-OES) at pH below 4 and at pH 5.8. Additionally, total amylolitic activity and alpha-amylase (1,4-alpha-d-glucan glucanohydrolase; EC 3.2.1.1) activity was assayed in plants exposed to arsenic treatments. At pH below 4, the uptake for As(III) and As(V) in roots was 137 and 124 mg As kg(-1) dry weight (d wt), respectively. Translocation of arsenic to the aerial part was relatively low ( approximately 5mg As kg(-1) d wt). The uptake for As(III) and As(V) in roots at pH 5.8 was about 43 and 30 mg As kg(-1) d wt, respectively, and translocation of As to the aerial part was not detectable. None of the arsenic treatments affected the total amylolitic activity in roots; however, the shoots from all treatments showed an increase in the total amylolitic activity. Alpha-amylase activity in the pea leaves was not significantly affected by arsenic treatments. X-ray absorption spectroscopy (XAS) studies showed a reduction of As(V) to As(III) in the roots. From linear combination X-ray absorption near edge structure (LC-XANES) fittings, it was determined that arsenic was present as a mixture of As(III) oxide and sulfide in pea roots.

X-ray absorption near edge structure and extended X-ray absorption fine structure analysis of standards and biological samples containing mixed oxidation states of chromium(III) and chromium(VI).

For the first time a method has been developed for the extended X-ray absorption fine structure (EXAFS) data analyses of biological samples containing multiple oxidation states of chromium. In this study, the first shell coordination and interatomic distances based on the data analysis of known standards of potassium chromate (Cr(VI)) and chromium nitrate hexahydrate (Cr(III)) were investigated. The standards examined were mixtures of the following molar ratios of Cr(VI):Cr(III), 0:1, 0.25:0.75, 0.5:0.5, 0.75:0.25, and 1:0. It was determined from the calibration data that the fitting error associated with linear combination X-ray absorption near edge structure (LC-XANES) fittings was approximately +/-10% of the total fitting. The peak height of the Cr(VI) pre-edge feature after normalization of the X-ray absorption (XAS) spectra was used to prepare a calibration curve. The EXAFS fittings of the standards were also investigated and fittings to lechuguilla biomass samples laden with different ratios of Cr(III) and Cr(VI) were performed as well. An excellent agreement between the XANES data and the data presented in the EXAFS spectra was observed. The EXFAS data also presented mean coordination numbers directly related to the ratios of the different chromium oxidation states in the sample. The chromium oxygen interactions had two different bond lengths at approximately 1.68 and 1.98 A for the Cr(VI) and Cr(III) in the sample, respectively.

Examination of arsenic(III) and (V) uptake by the desert plant species mesquite (Prosopis spp.) using X-ray absorption spectroscopy.

This study describes the effects of Arsenic(III) and (V) on the growth and their uptake by the desert plant mesquite (Prosopis spp.). Seedlings were sown in agar-based medium containing a modified Hoagland's nutrient solution. After 1 week, the seedlings were transplanted to arsenic (As) treated agar media that contained 5 mgL(-1) of As either As(III) (As(2)O(3)) or As(V) (As(2)O(5)). The plants were harvested after 14 days of growth and sectioned into roots, stems, and leaves. After digestion, As concentrations in the roots, stems, and leaves were determined using inductively coupled plasma-optical emission spectroscopy (ICP-OES). Our results showed that the As concentrations from As(V) were significantly higher than the As concentrations from As(III) in all portions of the plant. Plants exposed to As(V) concentrated (mg As kg(-1) d wt) about 770+/-191, 326+/-94, and 119+/-18 in roots, stems, and leaves, respectively. X-ray absorption spectroscopy (XAS) showed that As(V) was reduced to As(III) inside the mesquite plant. In addition, greater than 90% of the As(III) found in the mesquite plants was bound to sulfur ligands in the roots, stems and leaves.

Physiological and biochemical responses of sunflower (Helianthus annuus L.) exposed to nano-CeO2 and excess boron: Modulation of boron phytotoxicity.

Little is known about the interaction of nanoparticles (NPs) with soil constituents and their effects in plants. Boron (B), an essential micronutrient that reduces crop production at both deficiency and excess, has not been investigated with respect to its interaction with cerium oxide NPs (nano-CeO2). Considering conflicting results on the nano-CeO2 toxicity and protective role as antioxidant, their possible modulation on B toxicity in sunflower (Helianthus annuus L.) was investigated. Sunflower was cultivated for 30 days in garden pots containing original or B-spiked soil amended with nano-CeO2 at 0-800 mg kg-1. At harvest, Ce and B concentrations in tissues, biomass, and activities of stress enzymes in leaves were determined. Results showed that in the original soil, Ce accumulated mainly in roots, with little translocation to stems and leaves, while reduced root Ce was observed in plants from B-spiked soil. In the original soil, higher levels of nano-CeO2 reduced plant B concentration. Although morphological effects were not visible, changes in biomass and oxidative stress response were observed. Sunflower leaves from B-spiked soil showed visible symptoms of B toxicity, such as necrosis and chlorosis in old leaves, as well as an increase of superoxide dismutase (SOD) activity. However, at high nano-CeO2 level, SOD activity decreased reaching values similar to that of the control. This study has shown that nano-CeO2 reduced both the B nutritional status of sunflower in original soil and the B phytotoxicity in B-spiked soil.

Potential of Agave lechuguilla biomass for Cr(III) removal from aqueous solutions: thermodynamic studies.

Thermodynamic studies on the bioadsorption of Cr(III) onto Agave lechuguilla biomass were conduced. The experimental results at different temperatures were modeled using the Langmuir and Freundlich isotherms to obtain the characteristic parameters of each model. Both the Freundlich and Langmuir models were found to represent the bioadsorption process. The average adsorption capacities calculated from Freundlich (4.7 mg/g) and Langmuir (14.2 mg/g) isotherms showed A. lechuguilla to be an effective biomass in the removal of Cr(III) from an aqueous solution. Thermodynamic parameters (deltaG0, deltaH0 and deltaS0) determined in the temperature range from 10 to 40 degrees C along with the parameters of the Dubinin-Radushkevick equation support the idea that the binding of Cr(III) may be caused by interactions with functional groups such as carboxyl groups located on the outer surface of the cell tissue of the bioadsorbent.

Lessons learned: Are engineered nanomaterials toxic to terrestrial plants?

The expansion of nanotechnology and its ubiquitous applications has fostered unavoidable interaction between engineered nanomaterials (ENMs) and plants. Recent research has shown ambiguous results with regard to the impact of ENMs in plants. On one hand, there are reports that show hazardous effects, while on the other hand, some reports highlight positive effects. This uncertainty whether the ENMs are primarily hazardous or whether they have a potential for propitious impact on plants, has raised questions in the scientific community. In this review, we tried to demystify this ambiguity by citing various exposure studies of different ENMs (nano-Ag, nano-Au, nano-Si, nano-CeO2, nano-TiO2, nano-CuO, nano-ZnO, and CNTs, among others) and their effects on various groups of plant families. After scrutinizing the most recent literature, it seems that the divergence in the research results may be possibly attributed to multiple factors such as ENM properties, plant species, soil dynamics, and soil microbial community. The analysis of the literature also suggests that there is a knowledge gap on the effects of ENMs towards changes in color, texture, shape, and nutritional aspects on ENM exposed plants.

Use of silica-immobilized humin for heavy metal removal from aqueous solution under flow conditions.

Humin extracted from Sphagnum peat moss was immobilized in a silica matrix and column experiments were performed in order to evaluate the removal and recovery of metal ions from aqueous solution under flow conditions. These experiments also allowed testing the recycling capacity of the column. Single-element solutions of Cu(II) and Pb(II), and a multi-metal solution containing Cd(II), Cu(II), Pb(II), Ni(II), and Cr(III) were passed through the columns at a flow rate of 2 ml/min. A 0.5 M sodium citrate solution was used as the stripping agent in the metal-ion recovery process. Humin immobilized in the silica matrix exhibited a similar, and in some cases, even a higher capacity than other biosorbents for the removal of metal ions from aqueous solutions under flow conditions. The sodium citrate was effective in removing Cu(II), Pb(II), Cd(II), and Ni(II) from the metal saturated column. The selectivity of the immobilized biomass was as follows: Cr(III)>Pb(II)>Cu(II)>Cd(II)>Ni(II). This investigation provides a new, environmentally friendly and cost-effective possibility to clean up heavy-metal contaminated wastewaters by using the new silica-immobilized humin material.

Bioaccumulation of cadmium, chromium and copper by Convolvulus arvensis L.: impact on plant growth and uptake of nutritional elements.

The remediation of heavy metal-contaminated sites using plants presents a promising alternative to current methodologies. In this study, the potential accumulation of Convolvulus arvensis L. for Cd(II), Cr(VI), and Cu(II) was determined using an agar-based medium. The shoots of C. arvensis plants exposed to 20 mgl(-1) of these heavy metals, demonstrated capability to accumulate more than 3800 mg of Cr, 1500 mg of Cd, and 560 mg of Cu per kg of dry tissue. The outcome of this study and the field data previously reported corroborate that C. arvensis is a suitable candidate for the phytoremediation of Cd(II), Cr(VI), and Cu(II) contaminated soils. Furthermore, the concentration of Cr determined in the dry leaf tissue (2100 mgkg(-1)) indicates that C. arvensis could be considered as a potential Cr-hyperaccumulator plant species.

Effect of mixed cadmium, copper, nickel and zinc at different pHs upon alfalfa growth and heavy metal uptake.

Alfalfa plants were grown in soil-pots contaminated with a mixture of Cd(II), Cu(II), Ni(II), and Zn(II), (at 50 mg/kg each) at pHs of 4.5, 5.8, and 7.1. The plants were fertilized using a nutrient solution, which was adjusted appropriately to the same pH. Plants in the control treatment were grown in the absence of the heavy metals mixture. The growth of the control plants was the same at the three pHs studied and the heavy metal stressed plants also showed similar behavior at each pHs. There were statistically significant differences (P<0.05) between the shoot length of the control treatment plants and the length of plants grown in the presence of the heavy metal mixture. Under the effects of the heavy metal mixture, nickel was the most accumulated element in the shoot tissue, with 437, 333, and 308 ppm at pH 7.1, 5.8, and 4.5, respectively. Cadmium was found to be second in accumulated concentrations with 202 ppm, 124 ppm, and 132 ppm at pH 7.1, 5.8, and 4.5, respectively, while zinc was third, followed by copper. The maximum relative uptakes (element in plant/element in soil-water-solution) were found to be 26 times for nickel, 23 times for cadmium, 12 times for zinc. and 6 times for copper. We considered these relations as indicative of the ability of alfalfa plants to take up elements from a soil matrix contaminated with a mixture of cadmium, copper, nickel, and zinc.

Utilization of ICP/OES for the determination of trace metal binding to different humic fractions.

In this study, the use of inductively coupled plasma/optical emission spectrometry (ICP/OES) to determine multi-metal binding to three biomasses, Sphagnum peat moss, humin and humic acids is reported. All the investigations were performed under part per billion (ppb) concentrations. Batch pH profile experiments were performed using multi-metal solutions of Cd(II), Cu(II), Pb(II), Ni(II), Cr(III) and Cr(VI). The results showed that at pH 2 and 3, the metal affinity of the three biomasses exposed to the multi-metal solution that included Cr(III) presented the following order: Cu(II), Pb(II)>Ni(II)>Cr(III)>Cd(II). On the other hand, when Cr(VI) was in the heavy metal mixture, Sphagnum peat moss and humin showed the following affinity: Cu(II), Pb(II)>Ni(II)>Cr(VI)>Cd(II); however, the affinity of the humic acids was: Cu(II)>Pb(II), Cr(VI)>Ni(II)>Cd(II). The results demonstrated that pH values of 4 and 5 were the most favorable for the heavy metal binding process. At pH 5, all the metals, except for Cr(VI), were bound between 90 and 100% to the three biomasses. However, the binding capacity of humic acids decreased at pH 6 in the presence of Cr(VI). The results showed that the ICP/OES permits the determination of heavy metal binding to organic matter at ppb concentration. These results will be very useful in understanding the role of humic substances in the fate and transport of heavy metals, and thus could provide information to develop new methodologies for the removal of low concentrations of toxic heavy metals from contaminated waters.

Supported and unsupported nanomaterials for water and soil remediation: are they a useful solution for worldwide pollution?

Remediation technologies for wastes generated by industrial processes include coagulation, reverse osmosis, electrochemistry, photoelectrochemistry, advanced oxidation processes, and biological methods, among others. Adsorption onto activated carbon, sewage sludge, zeolites, chitosan, silica, and agricultural wastes has shown potential for pollutants' removal from aqueous media. Recently, nanoscale systems [nanoparticles (NPs) supported on different inorganic adsorbents] have shown additional benefits for the removal/degradation of several contaminants. According to the literature, NPs enhance the adsorption capacity of adsorbent materials and facilitate degradation of pollutants through redox reactions. In this review we analyzed relevant literature from 2011 to 2013, dealing with water and soil remediation by nanomaterials (NMs), either unsupported or supported upon inorganic adsorbents. Despite the outstanding reported results for some NMs, the analysis of the literature makes clear the necessity of more studies. There is lack of information about NMs regeneration and reusability, their large-scale application, and their efficiency in actual industrial wastewaters and contaminated soils. Additionally, little is known about NMs' life cycle, release of metal ions, disposal of pollutant loaded NMs, and their impacts on different ecosystems.

Exposure studies of core-shell Fe/Fe(3)O(4) and Cu/CuO NPs to lettuce (Lactuca sativa) plants: Are they a potential physiological and nutritional hazard?

Iron and copper nanomaterials are widely used in environmental remediation and agriculture. However, their effects on physiological parameters and nutritional quality of terrestrial plants such as lettuce (Lactuca sativa) are still unknown. In this research, 18-day-old hydroponically grown lettuce seedlings were treated for 15 days with core-shell nanoscale materials (Fe/Fe(3)O(4), Cu/CuO) at 10 and 20mg/L, and FeSO(4)·7H(2)O and CuSO(4)·5H(2)O at 10mg/L. At harvest, Fe, Cu, micro and macronutrients were determined by ICP-OES. Also, we evaluated chlorophyll content, plant growth, and catalase (CAT) and ascorbate peroxidase (APX) activities. Our results showed that iron ions/NPs did not affect the physiological parameters with respect to water control. Conversely, Cu ions/NPs reduced water content, root length, and dry biomass of the lettuce plants. ICP-OES results showed that nano-Cu/CuO treatments produced significant accumulation of Cu in roots compared to the CuSO(4)·5H(2)O treatment. In roots, all Cu treatments increased CAT activity but decreased APX activity. In addition, relative to the control, nano-Cu/CuO altered the nutritional quality of lettuce, since the treated plants had significantly more Cu, Al and S but less Mn, P, Ca, and Mg.

Citric acid modifies surface properties of commercial CeO2 nanoparticles reducing their toxicity and cerium uptake in radish (Raphanus sativus) seedlings.

Little is known about the mobility, reactivity, and toxicity to plants of coated engineered nanoparticles (ENPs). Surface modification may change the interaction of ENPs with living organisms. This report describes surface changes in commercial CeO2 NPs coated with citric acid (CA) at molar ratios of 1:2, 1:3, 1:7, and 1:10 CeO2:CA, and their effects on radish (Raphanus sativus) seed germination, cerium and nutrients uptake. All CeO2 NPs and their absorption by radish plants were characterized by TEM, DLS, and ICP-OES. Radish seeds were germinated in pristine and CA coated CeO2 NPs suspensions at 50mg/L, 100mg/L, and 200mg/L. Deionized water and CA at 100mg/L were used as controls. Results showed ζ potential values of 21.6 mV and -56 mV for the pristine and CA coated CeO2 NPs, respectively. TEM images showed denser layers surrounding the CeO2 NPs at higher CA concentrations, as well as better distribution and smaller particle sizes. None of the treatments affected seed germination. However, at 200mg/L the CA coated NPs at 1:7 ratio produced significantly (p ≤ 0.05) more root biomass, increased water content and reduced by 94% the Ce uptake, compared to bare NPs. This suggests that CA coating decrease CeO2 NPs toxicity to plants.

Arsenic speciation in biological samples using XAS and mixed oxidation state calibration standards of inorganic arsenic.

The speciation of elements without pre-edge features preformed with X-ray absorption near edge structure (XANES) can lead to problems when the energy difference between two species is small. The speciation of arsenic (As) in plant samples was investigated using the mixtures As2S3/As2O5, As2S3/As2O3, or As2O3/As2O5. The data showed that the energy separation (eV) between As2O5 and As2S3 was 5.8, between As2O3 and As2O5 was 3.6, and between As2S3 and As2O3 was 2.1. From the intensity of the white-line feature and the concentration of As species, calibration curves showing a limit of detection of approximately 10% were generated. In addition, an error of +/-10% was determined for the linear combination-XANES (LC-XANES) fitting technique. The difference between the LC-XANES fittings and calculations from the calibration curves was <10%. The data also showed that the speciation of As in a sample can be determined using EXAFS (extended X-ray absorption fine structure). Finally, it was also shown that both EXAFS and XANES of the sample should be examined to determine the true speciation of an element. Even though there is a difference of 2 eV between As(III) bound to O and As(III) bound to S, in the EXAFS region the As(III)-S and As(III)-O ligands are clearly visible. However, distinction between the As(III)-O and As(V)-O ligands in the EXAFS spectra was not clearly visible in this study.