Effect of Humic Substance Photoalteration on Lead Bioavailability to

Feb 25, 2011 - light of organic-rich riverine HS,13 alpine lake water samples,10,11 or purified peat ... ence of humic substance (HS) photoalteration ...
1 downloads 0 Views 860KB Size
Download PDF
ARTICLE pubs.acs.org/est

Effect of Humic Substance Photoalteration on Lead Bioavailability to Freshwater Microalgae Julian Spierings,‡ Isabelle A. M. Worms,‡ Pascal Mieville,§ and Vera I. Slaveykova*,†,‡ †

Aquatic Biogeochemistry and Ecotoxicology, Institute F.-A. Forel, Faculty of Sciences, University of Geneva, 10, route de Suisse, 1290 Versoix, Geneva, Switzerland ‡ Environmental Biophysical Chemistry, IIE-ENAC, Swiss Federal Institute of Technology (EPFL), Station 2, CH-1015 Lausanne, Switzerland § Laboratory of Biomolecular Magnetic Resonance, ISIC-SB, Swiss Federal Institute of Technology (EPFL), Station 6, CH-1015 Lausanne, Switzerland

bS Supporting Information ABSTRACT: The present study provides results on the influence of humic substance (HS) photoalteration on lead availability to the freshwater microalga Chlorella kesslerii. The evolution of the free lead-ion concentrations measured by the ion exchange technique [Pb]IET and intracellular lead contents was explored in the presence of Suwannee River humic (SRHA) and fulvic (SRFA) acids, as well as Aldrich humic acid (AHA) exposed at increasing radiance doses under a solar simulator. Modifications of HS characteristics highly relevant to Pb complexation and accumulation of HS to algal surfaces, including Fourier transform infrared spectroscopy, were followed. It was demonstrated that simulated sunlight exposure of HS increased [Pb]IET in the medium for SRFA and SRHA, but had no effect for AHA. No clear relationship was observed between the changes in free lead-ion concentrations and intracellular content in alga for all studied HS, suggesting that HS photodegradation products also exhibit Pb complexation properties, and that direct interactions between HS and alga are affected. Indeed, photoalteration of humic substances reduced the adsorption of HS to the algal surface; the effect was more pronounced for SRFA and AHA and less significant for SRHA. The bioavailability results were consistent with the characterization of the phototransformation of humic substances: Pb speciation changes followed the modification of the relative abundance of the carboxylic groups and their molecular environment, while the reduced HS adsorption to the alga correlated with losses of the double bond abundance and aromaticity.

’ INTRODUCTION Bioavailability is a key concept linking the changes in toxic metal concentration and speciation to their detrimental effects on biota.1 Among various freshwater constituents, humic substances (HS) play an important role in buffering toxic metal concentrations in aquatic ecosystems,2,3 thus reducing their potential detrimental effects on biota. Therefore, any environmental changes (e.g., climate and sunlight variability) that could affect HS concentration, structure, or reactivity can be expected to alter (e.g., to decrease) the HS metal-binding properties, and thus reduce their toxicity to aquatic (micro)-organisms. It is well documented that sunlight, and in particular UV-B, affects DOM quality and quantity4,5 via different processes, such as mineralization, photodegradation,6-8 or loss of suspended particulate organic carbon and the production of dissolved organic carbon (e.g., by photodissolution).9 Nonetheless, there is a paucity of studies addressing the effects of HS photoalteration on the bioavailability of toxic metals to aquatic biota. r 2011 American Chemical Society

The limited literature shows that photolysis of DOM most often results in the release of the bound metal, thus increasing the levels of free metal ion concentrations.10-15 Exposure to UV-B light of organic-rich riverine HS,13 alpine lake water samples,10,11 or purified peat humic acid15 resulted in increased free copperion concentrations. Increase of free cadmium-ion concentrations via photolytic release from cadmium-contaminated fulvic acid was found, while no increase was observed in cadmium-amended natural lake water HS.14 An enhancement of free lead-ion concentrations in estuarine samples after UV-B irradiation was reported.12 By contrast, light irradiation increased Cu complexation by wetland HS and thus decreased free copper-ion concentrations.6 Because metal bioavailability and biological Received: December 21, 2010 Accepted: February 10, 2011 Revised: February 7, 2011 Published: February 25, 2011 3452

dx.doi.org/10.1021/es104288y | Environ. Sci. Technol. 2011, 45, 3452–3458

Environmental Science & Technology effects are most often correlated with the concentration of noncomplexed metal, it can be anticipated that HS photoalteration could increase metal bioavailability via an increase of free metal-ion concentrations. Nonetheless, even more scarce and controversial is the literature about the biological availability (and toxicity) of metals in the presence of photoaltered DOM. Cu, Zn, Co, and Pb, but not Ni and Cd, toxicity to the freshwater green alga Pseudokirchneriella subcapitata increased up to 78% after 10 days of UV-B irradiation of high- and low-organic carbon-water samples.16 Pb, Cu, Ni, and Cd concentrations required to reduce algal growth by 50% were significantly decreased with both solar radiation and UV-B treatment doses; however, the changes in toxicity with UV-dosing were inconsistent among the metals tested.17 Cu toxicity to the larva Pimephales promelas increased in the presence of photooxidated DOM, following model predictions based on free copper-ion concentrations.6 Because DOM is a key parameter in water quality and cumulates the action of different environmental factors,4 the understanding of DOM’s role in metal bioavailability under changing conditions will be of prime importance in reducing the uncertainty in assessing the toxic metal effect on biota, and for the development of a predictive water quality framework for changing environment. Under well-controlled experimental conditions, the present study explores the relationship between lead availability to freshwater microalgae and Pb speciation in the presence of the HS exposed to solar radiation of increasing intensity. The specific questions to address were: (i) To what extent does the sunlightinduced transformation of HS affect Pb bioavailability? (ii) Do bioavailability alterations follow the changes in free lead-ion concentration? (iii) Is HS adsorption to alga altered by solar radiation, and what are the consequences for Pb bioavailability? Pb was chosen because of its toxicity and importance as a pollutant, and also because of the more complex interactions in the ternary system: Pb, microalgae, and HS.18,19

’ MATERIALS AND METHODS Standard fulvic and humic acids isolated from Suwannee River (SRFA and SRHA, International Humic Substances Society, St. Paul, MN) and Aldrich humic acid (AHA, Sigma-Aldrich Inc., St Louis, MO) were used as model HS. 1.0 g L-1 stock solutions of HS were prepared in Milli-Q water from freeze-dried powders; pH was adjusted to 9.0 with diluted NaOH. Solutions were stored at 4 °C in the dark for at least 24 h to ensure equilibration. pH was then readjusted to 6.0 with ultrapure HNO3 (Baker, instra) before further dilution. HS were chosen because of their different aromaticity, fluorescence fingerprints (Table S1), number, and affinity of the major metals binding sites.20 Exposure of HS to Solar Radiation of Increasing Intensity. Samples containing 10 mg C L-1 SRFA, SRHA, and AHA with pH adjusted to 6.0, in triplicate, were irradiated by the full solar spectrum for 12 h at increasing intensities under solar simulator (Sun 2000, Abet Technologies) equipped with a 1000 W Xe source lamp, an atmospheric absorption filter to mimic solar radiation distribution on Earth, and a light intensity controller. During the simulated sunlight exposure experiments, the radiation intensity was varied from 300 to 1000 W m-2. This variation corresponded to a radiance doses range from 7  106 to 2.1  107 J m-2 full solar spectrum (250-700 nm) and from 3  105 to 1  106 J m-2 UV-component (250-380 nm) measured at the surface of the irradiated samples. The radiance doses were

ARTICLE

chosen to correspond to the average annual cloudless sky UVdose in Europe ranging from 8  105 to 2  106 J m-2.21 During the experiments, the sample temperature was controlled by placing the reactors in a cooled water bath. The pH of the solutions was measured prior to and after the irradiation, and no significant changes were noted (ΔpH < 0.5 units). Pb Bioavailability to Freshwater Microalga Chlorella kesslerii. Pb bioavailability was characterized by measuring the intracellular Pb content, {Pb}int, in freshwater microalga Chlorella kesslerii exposed to 10-6 M Pb as Pb(NO3)2 for 1 h in the presence of the 10 mg C L-1, irradiated HS, and nonirradiated controls. To mimic highly impacted waters, 10-6 M total Pb concentration was chosen within the range of reported EC50 (50% effect concentration, i.e., growth inhibition) or 50% of the maximal {Pb}int for C. kesslerii.22 {Pb}int was operationally discriminated from the surface bound Pb by ethylenediaminetetraacetic acid (EDTA, ultra Fluka) extraction.22 More details about the experimental procedures can be found in the Supporting Information. Determination of Free Pb Ion Concentration by Ion Exchange Technique. The ion exchange technique (IET), in its “thermodynamic mode”, was used to determine free Pb concentrations, [Pb]IET, under the conditions of bioavailability experiments. The detailed experimental setup and procedures are described elsewhere.23 Briefly, the ion exchange column used contained 50 mg of Dowex resin in Na form. An equilibration time of 2 h was used. [Pb]IET was determined from the conditional partitioning coefficient, λ0 , following eq 1:24 ½PbIET ¼

Vel  ½Pbel λ0  mres

ð1Þ

where [Pb]el (mol L-1) is the concentration of eluted metal, determined by inductively coupled plasma-mass spectrometry (ICP-MS, Perkin-Elmer, Elan DRC II); mres (g) is the amount of resin, and Vel (L) is the elution volume. The conditional partitioning coefficient, λ0 , was determined by using standard solutions containing a known free lead-ion concentration and the same cationic composition of the sample. Accumulation of Photoaltered HS to C. kesslerii. The quantity of humic substances adsorbed to the algae was determined as the difference between the HS concentrations in the medium prior to and after contact with algae, as detailed previously,25,26 at a contact time of 1 h. The concentration of HS in the medium was measured by a UV spectrophotometer (Perkin-Elmer) at 254 nm. The absorbance measured at 254 nm was converted to HS concentrations by using a calibration graph established for each HS and irradiation condition. A control of HS-losses on the filters and container walls was performed under the accumulation study conditions in the absence of algae and was taken into account when determining the amount of adsorbed HS. The amount of HS bound to algae was presented as mg C per cm2 algal surface. Characterization of HS Photoalteration. To better understand the observed effects of HS photoalteration to Pb speciation and bioavailability, possible modification of HS characteristics was investigated by using large array of instrumental tools, probing different characteristics of these complex compounds.27 Potential photoalteration of major functional groups in the HS was followed by the Fourier transform infrared spectroscopy (FT-IR). FT-IR spectra of HS exposed to simulated sunlight of increasing intensity were compared to the spectra of 3453

dx.doi.org/10.1021/es104288y |Environ. Sci. Technol. 2011, 45, 3452–3458

Environmental Science & Technology

ARTICLE

Figure 1. (A) IET measured lead concentrations (nonirradiated controls, [Pb]IET,CTR) in solution containing 10 mg C L-1 AHA, SRFA, or SRHA enriched with 1  10-6 M Pb, pH = 6.0. (B) Effect of the radiant dose on [Pb]IET. The values were normalized to [Pb]IET,CTR. Error bars correspond to the square root of the sum of the squares of the standard deviation for [Pb]ITE and [Pb]ITE,CTR, obtained from three replicates for each condition. Student-Neuman-Keuls test demonstrated significant differences between [Pb]IET measured in the presence of photoaltered SRFA and SRHA, while no difference was found for AHA. IET-measured lead concentrations decreased in the order: [Pb]IET, SRFA > [Pb]IET, AHA g [Pb]IET, SRHA, prior to and [Pb]IET, SRFA g [Pb]IET, SRHA > [Pb]IET, AHA after irradiation.

nonirradiated control. The FT-IR frequency position was used to evaluate the chemical nature (chemical function) of photomodifications, and the changes in the IR-transmittance (in %) relative to the nonirradiated controls were used to evaluate the variation in the relative abundance of those chemical functions. General HS characteristics, such as specific UV absorbance, SUVA (L m-1 mg-1), fluorescence fingerprints, molar mass distribution, and dissolved organic carbon content (DOC), were also determined. Experimental details about each instrumental technique are provided in the Supporting Information. Data Analysis and Modeling. Control experiments (usually performed in triplicate) in the presence of the nonirradiated HS were always run in parallel with the assays in the presence of the photoaltered HS. Statistical differences within [Pb]IET or {Pb}int determined in the presence of HS irradiated at increasing radiance doses and nonirradiated controls were evaluated using the Student-Neuman-Keuls test in the Sigma Stat Software (Sigma Stat, Chicago, IL). Experimentally determined intracellular contents were compared to those predicted by using the MichaelisMenten equation and IET-measured lead concentrations,18,22 as detailed in the Supporting Information.

’ RESULTS AND DISCUSSION Pb Complexation in the Presence of Photoaltered HS. Preexposure of HS to simulated sunlight of increasing intensity resulted in a significant increase in the IET-measured Pb concentrations (Student-Neuman-Keuls test, P < 0.05) as compared to nonirradiated controls for both SRFA and SRHA. Enhanced [Pb]IET/[Pb]IET,CTR ratios of 2.2 and 2.4 were found in the presence of SRFA or SRHA exposed to a radiant dose of 2.8  106 J m-2. For AHA, [Pb]IET was not affected significantly (Student-Neuman-Keuls test, P > 0.05) as compared to the nonirradiated controls (Figure 1). No clear relationship between the radiance dose and the amount of the [Pb]IET was found for all HS. The raise in [Pb]IET was highest at 2.8  106 J m-2 exposure. Further increases in the radiance dose led to a decline in [Pb]IET released from the photoaltered HS; however, no statistically significant difference (Student-Neuman-Keuls test, P > 0.05) was found for [Pb]IET in the presence of SRFA and SRHA exposed to doses of 8.6  106 or 2.1  107 J m-2. The lack of

correlation between the increase in [Pb]IET and radiance dose suggests that photodegradation of humic substances in the aquatic environment can lead to the formation of degradation products with lead binding properties. Indeed, the evolution of the FT-IR spectra of the sunlight exposed HS revealed an increase in the relative abundance of the carboxyl groups in the photoaltered HS (see Characterization of HS Photoalteration section) as compared to nonirradiated controls. This is in agreement with literature reporting a formation of pyruvate, glyoxylate, acetate, acetaldehyde, and formate groups in photooxidized DOM,28 all exhibiting metal-binding properties. Furthermore, these observations are also consistent with a large chemical heterogeneity of the metal-binding sites typical for HS, which can be altered to a different extent under irradiation.17 Overall, exposure of SRFA and SRHA to high sunlight radiance results in a decrease of their Pb binding capacity, as demonstrated by the bigger [Pb]IET; however, no clear relationship between the amount of the uncomplexed Pb and the irradiation doses could be determined. Because the changes of the intracellular Pb content are predicted to follow free lead-ion concentration variations in the exposure medium (e.g., biotic ligand model, BLM,29 a similar pattern of the {Pb}int variations in the presence of the photoaltered HS was expected: for example, increase in {Pb}int in the presence of photoaltered SRFA and SRHA and no changes in the presence of AHA. Pb Availability to Microalgae in the Presence of Photoaltered HS. Experimental results from bioassays demonstrated, however, that no correlation existed between the increased release of Pb from the photoaltered HS (Figure 1) and intracellular Pb content in microalgae C. kesslerii (Figure 2). Preexposure of SRFA to sunlight of increasing intensity resulted in a decrease in the intracellular Pb content with respect to the nonirradiated control, opposite to about a 2-fold increase in the free metal ion concentrations (Figure 2). A decrease in the {Pb}int was also observed in the presence of photoaltered AHA, although no significant changes in [Pb]IET were found. Moreover, {Pb}int decreased in the presence of SRFA and AHA irradiated at increasing radiance doses, as illustrated by the reduction of {Pb}int/{Pb}int,CTR. By contrast, about 2.5-3-fold increase of {Pb}int/{Pb}int,CTR was determined in the presence of photoaltered SRHA, which followed (at least) qualitatively the 3454

dx.doi.org/10.1021/es104288y |Environ. Sci. Technol. 2011, 45, 3452–3458

Environmental Science & Technology

ARTICLE

Figure 2. (A) Intracellular lead content in the presence of 10 mg C L-1 AHA, SRFA, or SRHA (nonirradiated controls, {Pb}int,CTR). Total lead concentration is 10-6 M, pH = 6.0. (B) Effect of the radiance dose on {Pb}int in the presence of 10 mg C L-1 AHA, SRFA, and SRHA irradiated at increasing doses. {Pb}int were normalized to {Pb}int,CTR. Error bars correspond to the square root of the sum of the squares of the standard deviation for {Pb}int and {Pb}int,CTR, obtained from three replicates for each condition. In the absence of HS, {Pb}int = (3.6 ( 0.4)  10-12 mol cm-2 for C. kesslerii exposed to 1  10-6 M Pb for 1 h, pH = 6.0.

increase in [Pb]IET. Furthermore, the intracellular Pb content was 9, 8, and 15 times superior to that predicted by the measured [Pb]IET (Figure 1) and models such as the BLM,29 for example, for AHA, SRFA, and SRHA, similarly to the nonirradiated controls (Figure S1A). Because Pb uptake by C. kesslerii is controlled by the transport across the biological membrane rather than by the diffusion in the medium (e.g., diffusive flux of free lead-ions much greater than the internalization flux18,22), the discrepancy between measured and predicted intracellular Pb contents cannot be assigned to the contribution (e.g., via dissociation) of labile Pb-organic matter complexes formed after HS photoalteration. Furthermore, the increasing radiance doses reduced the shift between the measured and predicted Pb intracellular contents for AHA and SRFA, but had no significant effect for SRHA (Figure S3B). These observations are in agreement with previous studies about the bioavailability of Pb to microalgae in the presence of dissolved18,25,26,30 and colloidal organic matter of different composition,23 demonstrating that HS could also affect (e.g., increase) Pb bioavailability by adding supplementary binding sites26,30 and affecting the algal cell wall speciation.18 These effects relate to the capacity of HS to accumulate on microorganism surfaces.31 Accordingly, the hypothesis that the photoalteration of HS influences (e.g., reduce) the capacity of HS to accumulate to algal surfaces was further tested experimentally. Accumulation of Photoaltered HS to Algal Surface. Reduced amounts of the HS accumulated to C. kesslerii after irradiation at 2.1  107 J m-2 were measured as compared to the nonirradiated controls (Figure 3). Prior to irradiation, the amount of adsorbed HS to C. kesslerii was comparable for SRFA and AHA and higher for SRHA: {HA}ads/[HS]diss ratios were 8.9 ( 1.0, 8.7 ( 1.0, and 11.2 ( 0.5 (10-3 cm). A radiance dose of 2.1  107 J m-2 resulted in a reduction of the ratio {HA}ads/[HS]diss to 2.0 ( 0.4, 1.45 ( 0.2, and 8.0 ( 0.5 (10-3 cm) for AHA, SRFA, and SRHA, respectively, clearly indicating that photoalteration resulted in a decrease of the amount of HS adsorbed to algae in a HS-specific way. The above results demonstrate that the consequences of sunlight irradiation of organic matter on Pb availability by the microalga C. kesslerii are complex and that, in addition to the complexation properties of HS, solar radiation also alters their capacity to interact (e.g., to adsorb) with algae. To our knowledge, this is a first study demonstrating the effect of the solar

Figure 3. Adsorbed HS onto algal cells prior to and after irradiation at 2.1  107 J m-2 full solar spectrum, pH 6.0. Error bars correspond to standard deviations of three replicates.

radiation on HS0 capacity to adsorb to microalgae. An increase in the [Pb]IET is expected to enhance {Pb}int, while the reduction of the amount of HS adsorbed to C. kesslerii results in a decrease of {Pb}int for a given free lead-ion concentration. To link the above observations and elucidate the causal relationship with photoinduced HS structure modifications, the influence of the irradiation of HS on different functional groups was further evaluated. In particular, the focus was on the alteration of carboxylic groups and their environment, as well as the aromatic structures considered of high relevance to Pb complexation and HS adsorption to algae, respectively. Carboxylic groups are considered to be the major metal complexing groups in HS under circumneutral pH. Their chemical behavior is highly dependent on their structural environment such as the type and position of other chemical groups: for example, ketones and hydroxyl groups that may participate with carboxylic groups in metal chelation.32 The adsorption of HS to algae is dependent on the HS composition and seems to be significant for HS with a high hydrophobic character (and high aromaticity).31 Therefore, exploring the effect of sunlight on the aromatic structures was also of interest. 3455

dx.doi.org/10.1021/es104288y |Environ. Sci. Technol. 2011, 45, 3452–3458

Environmental Science & Technology Effect of Solar Radiation on Relevant HS Characteristics. FT-IR revealed differences between the nonirradiated samples and those receiving increasing radiation doses (Figure 4). Two regions of interest with respect to photoalteration of Pb-complexing properties and the adsorption of the HS to the algae were considered: 3500-2500 and 1700-1000 cm-1. The first spectral window is abundant in H-bonded OH stretching, aromatic, and aliphatic C-H stretching.33 The second spectral window is rich in CdO stretch from COOH and asymmetrical and symmetrical (1650-1590 cm-1) stretching from deprotonated carboxylic groups (1650-1590 cm-1) and amides.33,34 A common and prominent feature of all three HS was that the relative abundance of the deprotonated carboxylic and amide groups increased upon irradiation, as illustrated by the band at 16221597 cm-1 attributed to the CdO stretch from COO- and amide groups. Given that nitrogen represents only 1.7% in SRHA and only 0.72% in SRFA,20 the intensity of transmittance changes seems inconsistent with amide group assignment. Thus, we suppose that CdO stretch originates from COO- rather than from amide groups. Furthermore, it was found that the amide groups are very prone to photooxidation.5 The formation of hydroxyl groups from carboxylic acids and alcohols was also observed from the evolution of the broad bands, which appeared close to 3400 cm-1 for SRFA and 3309 cm-1 for SRHA, usually associated with the H-bonded OH stretching. This was most evident for SRHA (25% increase in the transmittance) rather than for SRFA (only 3-7% increase). In addition, the photoalterations were not directly proportional to the radiance dose, but rather appeared at lower doses. For example, the aldehyde groups formed at lower radiance doses and decreased when radiance doses further increased (e.g., photooxidation of aldehydes in acids). The percentage of newly formed carboxyl groups (probably from low molar mass acids) increased with the increased radiance doses for SRFA, did not change significantly for SRHA, but decreased for AHA. An opposite tendency was observed in the evolution of the newly formed hydroxyl groups: enhanced radiance doses resulted in a decrease of the % of OH-groups for SRFA, no changes for SRHA, and an increase for AHA. At the highest radiance doses, a formation of free (non H-bonded) OH was observed for AHA. A formation of both aldehyde and arene functional groups was found only in photoaltered SRHA as compared to nonirradiated control. Another feature of the FT-IR spectra evolution with the irradiation intensity, relevant with respect to the adsorption of HS to algae, was the important decrease in the relative abundance of the double bonded structures CdC, spectral band at 2852 cm-1. These modifications are most visible in the FT-IR spectra of SRFA (Figure 4B) in which aromatic carbon represents about 30% of the total organic carbon.36 FT-IR spectra evolution is also consistent with the SUVA and fluorescent fingerprint changes upon irradiation (Table S1 and Figure S2). UV-absorbance declines in the wavelength range from 250 to 450 nm after irradiation, with HS-absorbance losses higher at 325 nm than at 254 nm. Upon irradiation at 2.8  106 J m-2, a 10-15% increase in A254/A325nm was found, while the SUVA-values increased significantly in an HS-specific manner. At the highest radiance doses used in the present study (2.1  107 J m-2), the SUVA-rise was ca. 40% for SRFA, 17% for SRHA, and 14% for AHA, as compared to the nonirradiated controls. Decreased absorbance in the UV region suggest a loss of sp2-hybridized carbon bonds (e.g., double bonds in aromatics),

ARTICLE

Figure 4. Fourrier transform infrared (FT-IR) spectra of AHA (A), SRFA (B), and SRHA (C). For all spectra, the relative percent transmittance is plotted against wavenumber. Band frequency assignment based on peaks previously reported for the humic substances:33,35 3400-3200 cm-1, OH stretch from -COOH and -COH; 31002800 cm-1, CH stretch from CH, -CH2, and CH3 and aromatics; 2700-2400 cm-1, OH stretch from strongly H-bonded -COOH; 1622-1597 cm-1, CdO stretch from COO- and amide groups. Note that the position of the CdO stretch from the carboxylic groups, usually observed at 1700 cm-1, is pH dependent and shifts toward high energies when pH increases.34

which is consistent with the significant decrease in the percentage of the double bonds in FT-IR for SRFA. However, FT-IR spectra do not show significant variations in sp2-double bonds for SRHA, although the important decrease in SUVA (and aromaticity). SUVA determined at 254 nm is highly correlated to the aromaticity as determined by the 13C NMR different types of source materials;37 however, a wide range of reactivity for DOM with similar SUVA was observed, showing that SUVA does not provide information about the reactivity of DOM derived from different types of source materials. A decrease in the HS adsorption to algae upon phototransformation of HS is in qualitative agreement with a decrease in the aromaticity, as shown by SUVA evaluation (Table S1). Decrease in SUVA and UV absorbing properties in combination with no changes in the DOC concentrations (Table S1) are highly suggestive of aromatic moieties conversion to low molar mass organic compounds (Figure S3). For SRHA and SRFA, the ratio of the intensity of peaks “type A-humic” and “type C-fulvic” (F240/445/F320/445) augmented 3456

dx.doi.org/10.1021/es104288y |Environ. Sci. Technol. 2011, 45, 3452–3458

Environmental Science & Technology with the increase of radiance dose, suggesting that fulvic fluorophores are more sensitive to irradiation than humic ones (Figure S2 and Table S1). Similarly, the fraction F240/445/ F320/445 increased after a 20 min exposure to UV-visible radiation of Amazon affluent waters.38 Decreases in the aromaticity (e.g., decrease in the quinine and lignin derived polyphenoles) with increasing irradiation doses also suggests that the molecular electronegativity, related to the electron-withdrawing character of the aromatic moieties, will decrease and thus could reduce the complexation of the positively charged Pb2þ.27 Indeed, the increase in Pb released from SRFA was more pronounced than that from photoaltered AHA. It was outside of the scope of the study to explore the detailed compositional, structural, and functional characteristics of HS and propose the transformation pathways. Nonetheless, the results of the present study are consistent with the extensive literature (see ref 5 for review) demonstrating that irradiated HS acts as photosensitizers, generating short-lived, highly reactive oxygen species. The above observations also confirmed that AHA is less prone to photooxidation as compared to SRFA and SRHA; consequently, Pb speciation and uptake were insignificantly affected. The obtained results also suggest that the influence of sunlight radiation affects Pb complexation by both alteration of the relative abundance of carboxylic groups and by affecting their molecular environment and thus their Pb affinity in a HS-specific way. Therefore, in addition to the direct deleterious influence of UV-B exposure on the aquatic phytoplankton,39 an increased sunlight radiation can significantly affect contaminant biouptake by changes in the medium chemistry. DOM is a key water quality parameter encompassing the influence of rarely monitored processes related to global environmental changes. Thus, our ability to interpret and predict the impact of toxic metals on aquatic phytoplankton, representing the important primary producers, under changing conditions is directly related to our ability to quantify and foresee the effect of dissolved organic matter on metal bioavailability.

’ ASSOCIATED CONTENT

bS

Supporting Information. Excitation-emission matrices, FT-IR results, and AFlFFF fractograms of HS irradiated with sunlight of increasing doses, as well as experimental details about HS characterization, modeling, and prediction of intracellular Pb content in Chlorella kesslerii. This material is available free of charge via the Internet at http://pubs.acs.org.

’ AUTHOR INFORMATION Corresponding Author

*Phone: þ41 22 379 03 35. Fax: þ41 22 379 03 29. E-mail: vera. [email protected].

’ ACKNOWLEDGMENT We gratefully acknowledge the financial support provided by the Swiss National Science Foundation project PP002102640. Thanks are extended to T. Kohn for kindly providing access to the solar simulator, E. Alasonati for the AFlFFF-UV measurements, and J. Bolzman for HS adsorption to algae experiments.

ARTICLE

’ REFERENCES (1) Wilkinson, K. J.; Buffle, J. Critical evaluation of physicochemical parameters and processes for modelling the biological uptake of trace metals in environmental (aquatic) systems. In Physicochemical Kinetics and Transport at Biointerfaces; van Leeuwen, H. P., Koester, W., Eds.; John Wiley & Sons: Chichester, 2004; pp 447-533. (2) Slaveykova, V. I.; Wilkinson, K. J. Predicting the bioavailability of metals and metal complexes: Critical review of the biotic ligand model. Environ. Chem. 2005, 2, 9–24. (3) Campbell, P. G. C.; Errecalde, O.; Fortin, C.; Hiriart-Baer, W. R.; Vigneault, B. Metal bioavailability to phytoplankton - applicability of the biotic ligand model. Comp. Biochem. Physiol., C 2002, 133, 189–206. (4) Zepp, R. G.; Erickson, D. J., III; Paul, N. D.; Sulzberger, B. Interactive effects of solar UV and climate change on biogeochemical cycles. Photochem. Photobiol. Sci. 2007, 6, 286–300. (5) Sulzberger, B.; Durisch-Kaiser, E. Chemical characterization of dissolved organic matter (DOM): A prerequisite for understanding UVinduced changes of DOM absorption properties and bioavailability. Aquat. Sci. 2009, 71, 104–126. (6) Brooks, M. L.; Meyer, J. S.; McKnight, D. M. Photooxidation of wetland and riverine dissolved organic matter: Altered copper complexation and organic composition. Hydrobiologia 2007, 579, 95–113. (7) Moran, M. A.; Sheldon, W. M.; Zepp, R. G. Carbon loss and optical property changes during long-term photochemical and biological degradation of estuarine dissolved organic matter. Limnol. Oceanogr. 2000, 45, 1254–1264. (8) Obernosterer, I.; Benner, R. Competition between biological and photochemical processes in the mineralization of dissolved organic carbon. Limnol. Oceanogr. 2004, 49, 117–124. (9) Mayer, L. M.; Schick, L. L.; Skorko, K.; Boss, E. Photodissolution of particulate organic matter from sediments. Limnol. Oceanogr. 2006, 51, 1064–1071. (10) Sander, S.; Kim, J. P.; Anderson, B.; Hinter, K. A. Effect of UVB irradiation on Cu2þ- binding organic ligands and Cu2þ speciation in alpine lake water of New Zealand. Environ. Chem. 2005, 2, 56–62. (11) Shank, G. C.; Whitehead, R. F.; Smith, M. L.; Skrabal, S. A.; Kieber, R. J. Photodegradation of strong copper complexing ligands in organic-rich estuarine waters. Limnol. Oceanogr. 2006, 51, 884–892. (12) Lindim, C.; Mota, A. M.; Goncalves, M. L. S. Influence of UV-B radiation in lead speciation from an estuarine sample. Water Res. 2000, 34, 3325–3334. (13) Brooks, M. L.; Meyer, J. S.; Boese, C. J. Toxicity of copper to larval Pimephales promelas in the presence of photodegraded natural dissolved organic matter. Can. J. Fish. Aquat. Sci. 2007, 64, 391–401. (14) Neron, R.; Auclair, J. C.; Fortin, C. Rate of Cd2þ release from dissolved fulvic acid and natural dissolved organic carbon as a function of UVB dose. Environ. Chem. 2006, 3, 433–438. (15) Vidali, R.; Remoundaki, E.; Tsezos, M. Humic acids copper binding following their photochemical alteration by simulated solar light. Aquat. Geochem. 2010, 16, 207–218. (16) Winch, S.; Ridal, J.; Lean, D. Increased metal bioavailability following alteration of freshwater dissolved organic carbon by ultraviolet B radiation exposure. Environ. Toxicol. 2002, 17, 267–274. (17) Winch, S.; Lean, D. Comparison of changes in metal toxicity following exposure of water with high dissolved organic carbon content to solar, UV-B and UV-A radiation. Photochem. Photobiol. 2005, 81, 1469–1480. (18) Lamelas, C.; Pinheiro, J. P.; Slaveykova, V. I. Effect of humic acid on Cd(II), Cu(II), and Pb(II) uptake by freshwater algae: Kinetic and cell wall speciation considerations. Environ. Sci. Technol. 2009, 43, 730–735. (19) Lamelas, C.; Slaveykova, V. I. Pb uptake by the freshwater alga Chlorella kesslerii in the presence of dissolved organic matter of variable composition. Environ. Chem. 2008, 5, 366–372. (20) Averett, R. C.; Leenheer, J. A.; McKnight, D. M.; Thorn, K. A. Humic substances in the Suwannee River, Georgia: interactions, properties, and proposed structures. U.S. Geol. Surv. Water-Supply Pap. 1994, 2373. 3457

dx.doi.org/10.1021/es104288y |Environ. Sci. Technol. 2011, 45, 3452–3458

Environmental Science & Technology

ARTICLE

(21) Seckmeyer, G.; Darius, P.; Merle, G.; Diamantino, H.; Bjorn, J.; Ann, W.; Anna-Maria, S.; Alkis, B.; Berit, K.; Colette, B.; Jacqueline, L.; Brian, G.; Peter, K.; Tapani, K.; Jussi, K.; Beate, U.; Harry, S.; Peter den, O.; Michal, J.; Peter, W.; Julian, G.; Bernhard, M.; Alain de la, C.; Stana, S.; Fernanda, C. Variability of UV irradiance in Europe. Photochem. Photobiol. 2008, 84, 172–179. (22) Slaveykova, V. I.; Wilkinson, K. J. Physicochemical aspects of lead bioaccumulation by Chlorella vulgaris. Environ. Sci. Technol. 2002, 36, 969–975. (23) Worms, I. A. M.; Traber, J.; Kistler, D.; Sigg, L.; Slaveykova, V. I. Uptake of Cd(II) and Pb(II) by microalgae in presence of colloidal organic matter from wastewater treatment plant effluents. Environ. Pollut. 2010, 158, 369–374. (24) Cantwell, F. F.; Nielsen, J. S.; Hrudey, S. E. Free nickel ion concentration in sewage by an ion exchange column-equilibration method. Anal. Chem. 1982, 54, 1498–1503. (25) Slaveykova, V. I.; Wilkinson, K. J.; Ceresa, A.; Pretsch, E. Role of fulvic acid on lead bioaccumulation by Chlorella kesslerii. Environ. Sci. Technol. 2003, 37, 1114–1121. (26) Lamelas, C.; Wilkinson, K. J.; Slaveykova, V. I. Influence of the composition of natural organic matter on Pb bioavailability to microalgae. Environ. Sci. Technol. 2005, 39, 6109–6116. (27) Leenheer, J. A.; Croue, J. P. Characterizing aquatic dissolved organic matter. Environ. Sci. Technol. 2003, 37, 18A–26A. (28) Lepane, V.; Persson, T.; Wedborg, M. Effects of UV-B radiation on molecular weight distribution and fluorescence from humic substances in riverine and low salinity water. Estuarine, Coastal Shelf Sci. 2003, 56, 161–173. (29) Di Toro, D. M.; Allen, H. E.; Bergman, H. L.; Meyer, J. S.; Paquin, P. R.; Santore, R. C. Biotic ligand model of the acute toxicity of metals. 1. Technical basis. Environ. Toxicol. Chem. 2001, 20, 2383–2396. (30) Lamelas, C.; Slaveykova, V. I. Comparison of Cd(II), Cu(II), and Pb(II) biouptake by green algae in the presence of humic acid. Environ. Sci. Technol. 2007, 41, 4172–4178. (31) Campbell, P. G. C.; Twiss, M. R.; Wilkinson, K. J. Accumulation of natural organic matter on the surfaces of living cells: implications for the interaction of toxic solutes with aquatic biota. Can. J. Fish. Aquat. Sci. 1997, 54, 2543–2554. (32) Leenheer, J. A.; Wershaw, R. L.; Reddy, M. M. Strong-acid, carboxyl-group structures in fulvic acid from the Suwannee River, Georgia. 2. Major structures. Environ. Sci. Technol. 1995, 29, 399–405. (33) Davis, W. M.; Erickson, C. L.; Johnston, C. T.; Delfino, J. J.; Porter, J. E. Quantitative Fourier transform infrared spectroscopic investigation humic substance functional group composition. Chemosphere 1999, 38, 2913–2928. (34) Cabaniss, S. E.; Leenheer, J. A.; McVey, I. F. Aqueous infrared carboxylate absorbances: aliphatic di-acids. Spectrochim. Acta, Part A 1998, 54, 449–458. (35) Elkins, K. M.; Nelson, D. J. Fluorescence and FT-IR spectroscopic studies of Suwannee river fulvic acid complexation with aluminum, terbium and calcium. J. Inorg. Biochem. 2001, 87, 81–96. (36) McKnight, D. M.; Boyer, E. W.; Westerhoff, P. K.; Doran, P. T.; Kulbe, T.; Andersen, D. T. Spectrofluorometric characterization of dissolved organic matter for indication of precursor organic material and aromaticity. Limnol. Oceanogr. 2001, 46, 38–48. (37) Weishaar, J. L.; Aiken, G. R.; Bergamaschi, B. A.; Fram, M. S.; Fujii, R.; Mopper, K. Evaluation of specific ultraviolet absorbance as an indicator of the chemical composition and reactivity of dissolved organic carbon. Environ. Sci. Technol. 2003, 37, 4702–4708. (38) Mounier, S.; Patel, N.; Quilici, L.; Benaim, J. Y.; Benamou, C. Three-dimensional fluorescence of the dissolved organic carbon in the Amazon river. Water Res. 1999, 33, 1523–1533. (39) Haeder, D. P.; Kumar, H. D.; Smith, R. C.; Worrest, R. C. Effect of solar UV radiation on aquatic ecosystems and interactions with climate change. Photochem. Photobiol. Sci. 2007, 6, 267–285.

3458

dx.doi.org/10.1021/es104288y |Environ. Sci. Technol. 2011, 45, 3452–3458