Environ. Sci. Technol. 2004, 38, 2865-2872
Influence of Natural Organic Matter Source on Copper Speciation As Demonstrated by Cu Binding to Fish Gills, by Ion Selective Electrode, and by DGT Gel Sampler C H A D D . L U I D E R , † J O H N C R U S I U S , ‡,§ RICHARD C. PLAYLE,+ AND P . J E F F C U R T I S * ,† Earth and Environmental Science Department, Okanagan University College, Kelowna, British Columbia, Canada V1V 1V7, Department of Earth and Ocean Sciences, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z4, and Department of Biology, Wilfrid Laurier University, Waterloo, Ontario, Canada N2L 3C5
Rainbow trout (Oncorhynchus mykiss, 2 g) were exposed to 0-5 µM total copper in ion-poor water for 3 h in the presence or absence of 10 mg C/L of qualitatively different natural organic matter (NOM) derived from water spanning a large gradient in hydrologic residence time. Accumulation of Cu by trout gills was compared to Cu speciation determined by ion selective electrode (ISE) and by diffusive gradients in thin films (DGT) gel sampler technology. The presence of NOM decreased Cu uptake by trout gills as well as Cu concentrations determined by ISE and DGT. Furthermore, the source of NOM influenced Cu binding by trout gills with high-color, allochthonous NOM decreasing Cu accumulation by the gills more than low-color autochthonous NOM. The pattern of Cu binding to the NOM measured by Cu ISE and by Cu accumulation by DGT samplers was similar to the fish gill results. A simple Cugill binding model required an NOM Cu-binding factor (F) that depended on NOM quality to account for observed Cu accumulation by trout gills; values of F varied by a factor of 2. Thus, NOM metal-binding quality, as well as NOM quantity, are both important when assessing the bioavailability of metals such as Cu to aquatic organisms.
Introduction Natural organic matter (NOM) acts as a complexing agent for many metals, decreasing their bioavailability to fish and other aquatic organisms (e.g., refs 1-4). Thus, inferring the impact of metals in natural water samples on aquatic life requires some means of evaluating how much of the total metal concentration is available for uptake by biota. Some researchers have used direct methods that involve measuring the uptake of metals by aquatic organisms; others have used analytical techniques that quantify certain metal species, * Corresponding author phone: (250)762-5445, ext. 7521; fax: (250)470-6005; e-mail:
[email protected]. † Okanagan University College. ‡ University of British Columbia. § Present address: U.S. Geological Survey, Woods Hole, MA 02543. + Wilfrid Laurier University. 10.1021/es030566y CCC: $27.50 Published on Web 04/17/2004
2004 American Chemical Society
relying on experimental evidence as to which metal species are taken up by biota. One direct method of inferring metal availability to freshwater fish has been to measure metal accumulation by the gills (5-10). This type of work has led to the development of metal-gill binding models, which have been extended to predict metal toxicity and are now generally known as biotic ligand models (BLM; refs 11-14). Conceptually, these models consider the biological membrane as a ligand with a particular metal binding strength (usually as conditional equilibrium binding constants; Figure 1), which can be inserted into aquatic geochemistry programs that normally calculate metal speciation in the water column. Complexing agents such as NOM, OH-, and carbonate bind cationic metal in solution, thereby decreasing metal binding to the gills and therefore decreasing metal toxicity. In addition, cations such as Ca2+, Mg2+, Na+, and H+ compete at metal binding sites at the gills, decreasing the binding of otherwise available metal (Figure 1). Biotic ligand models bridge the gap between measures of water chemistry and measures of metal toxicity to aquatic organisms by specifically considering complexation in the water column and competition at the biological membrane (10-14). Most analytical approaches for inferring metal availability to biota quantify the free metal ion, as formulated in the free ion activity model (FIAM) of metal toxicity to aquatic organisms (15). One such analytical approach uses ion selective electrodes (ISE), where the activity of a single metal (e.g., Cu2+) is determined from the ISE mV response (16). Extensive work has been done with the Cu ISE because the methodology is relatively simple, and alternatives for measuring free ionic Cu are limited (17), although care must be taken to control for ionic composition and especially sample pH (18, 19). The difference between total dissolved Cu and free labile Cu2+determined by ISE is usually taken as an approximation of complexed Cu (20). Free labile Cu2+ is correlated with Cu toxicity when all other water quality parameters remain constant (21-23), consistent with the FIAM. Another analytical method to infer bioavailable metal concentration is the diffusive gradients in thin films (DGT) approach (24), which sequesters an operationally defined DGT-labile fraction of divalent metals. Divalent metals are concentrated onto a high-affinity cation-exchange resin (Chelex) after diffusing through a permeable polyacrylimide gel layer. Metal species detected by the DGT approach are those that can both diffuse through the gel layer and be sequestered by the resin, including free Cu2+ plus inorganic and organic Cu species that can dissociate to Cu2+ within the gel layer. Colloidal and particulate metal species do not diffuse through the gel (25, 26). Highly colored, allochthonous (terrestrially derived) NOM decreases metal accumulation by fish gills and decreases the toxicity of a mixed-metal solution better than does less colored, autochthonous-like NOM (algal derived), as we have demonstrated previously (3). Allochthonous NOM is typically derived from lignin-containing plants, the degradation of which yields fulvic acids relatively rich in aromatic ring content (e.g., high aromaticity; ref 27). These aromatic rings absorb a portion of visible light (so allochthonous NOM is brown) and especially absorb ultraviolet light (28-30). The complexation capacity of humic substances for Cu (and H+) correlates with both absorbance of ultraviolet light (at 254 nm) and with aromaticity measured by 13C nuclear magnetic resonance (31). Measures of NOM quality such as these have the potential to shed light on systematic influences on metal bioavailability to aquatic biota. VOL. 38, NO. 10, 2004 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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FIGURE 1. Schematic diagram of a simple Cu-gill binding model. The asterisk represents a Cu-binding site on a trout gill. Numbers represent the log conditional equilibrium constants for the gill and for natural organic matter (NOM) entered into the MINEQL+ program; the higher the number, the stronger the binding. The CO3, OH, and Cl values were already in the program. F is the Cu-binding factor for a particular NOM. Modified from refs 6 and 11. See text and Supporting Information for more details.
The objectives of our current research were 3-fold. First, we determined, under controlled conditions, whether different NOMs alter the availability of Cu to aquatic organisms to varying degrees, as indicated by different amounts of Cu accumulating on fish gills. Second, we compared the fishgill assay results with ISE and DGT measures of potentially bioavailable Cu in water. And finally, we related the biologically relevant differences in Cu binding to easily measured optical properties of NOM.
Materials and Methods Natural Organic Matter Isolation by Reverse Osmosis. Natural organic matter (NOM) was collected near Kelowna, in the southern interior of British Columbia, in June 2001. Sampling locations were in an unnamed, first-order creek in the upper watershed (unnamed creek, 50°03′ N, 119°14′ W) and along a chain of lakes (Duck Lake, 50°00′ N, 119°24′ W and Kalamalka Lake, 50°10′ N, 119°20′ W; a map of the study area is available in ref 32) that demonstrate increasing water residence time from 0.5 to 81 years (32). Most of the water for the entire system originates at high elevation because the low-elevation study lakes have small direct drainages and have a semi-arid climate. Soils in the headwaters are luvisols, and the forests are dominated by lodgepole pine, white spruce, and western redcedar. These sites were chosen to yield samples ranging from high color, allochthonousdominated NOM (unnamed creek, Duck Lake) to low color, autochthonous-like NOM (Kalamalka Lake). A mesocosm was used to produce pure autochthonous NOM from algae. The mesocosm was constructed from clear Plexiglass (approximately 100% transmission throughout the visible light range and no detectable transmision of UV light radiation below 368 nm) and fitted with a Plexiglass lid. The mesocosm was filled with 1000 L of deionized water and amended with reagent grade salts to simulate 1:10 global average river water (33) and with nitrogen and phosphorus (107.1 mM N and 3.2 mM P; 146 µM total alkalinity). The mesocosm was inoculated with algae by filtering 2 L of water from Okanagan Lake (49°50′ N, 119°30′ W), an oligotrophic lake that dominates the Okanagan Valley. The mesocosm 2866
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was outside in a locked compound from January to April 2001, where it was aerated and heated to 4 °C to prevent freezing. The procedure for standardizing each NOM was as follows. Approximately 200 L of water was pumped through silicon tubing by a peristaltic pump and was filtered using 144 mm diameter glass fiber filters (nominal pore size 1 µm; Geotech Environmental Equipment, Denver, CO) before being concentrated using a stainless steel, portable reverse osmosis unit (Limnological Research Corporation, Kelowna) with a molecular weight cutoff of about 400 Da (FilmTec FT30 US Filters thin composite RO membrane, Minneapolis, MN). Once concentrated to approximately 5 L, metals were displaced from NOM binding sites and removed from solution by exposing the NOM concentrates to a H+-cation-exchange resin (Amberlite IR-118H) to a final pH
