Environ. Sci. Technol. 1997, 31, 603-611
Bioavailability of Cr(III) and Cr(VI) to Marine Mussels from Solute and Particulate Pathways WEN-XIONG WANG, SARAH B. GRISCOM, AND NICHOLAS S. FISHER* Marine Sciences Research Center, State University of New York, Stony Brook, New York 11794-5000
Mussels have been extensively used as biological monitors of coastal contamination. This study measured the assimilation efficiencies (AEs) of Cr(III) and Cr(VI) in the mussel Mytilus edulis from ingested food, uptake from the dissolved phase, and the physiological efflux rates following uptake. A bioenergetic-based kinetic model was then employed to determine the relative contributions of different Cr species and their accumulation pathways to the overall concentration of Cr in mussels. The concentration factors of Cr(III) in four diverse marine phytoplankters ranged between 104 and 105, whereas for Cr(VI) they were 2 to 5 × 102. Rapid reduction of Cr(VI) to Cr(III) was observed in coastal sediments. AEs of Cr(III) in mussels from ingested sediments were consistently 18%, whereas dissolved Cr(III) is more abundant at lower salinities. The bioavailability of Cr, which is complicated by its speciation and redox behavior, has been less studied than other toxic metals. Cr(VI) crosses biological membranes more readily than Cr(III) (4, 5). Most Cr in natural sediments is associated with the silicate lattice fraction and is not biologically available (21, 22). The very low assimilability of Cr(III) has led to its use as an inert tracer of ingested particulate matter to measure carbon assimilation in marine invertebrates (23), including bivalve molluscs (24, 25). However, Cr assimilation in the clam Macoma balthica is highly dependent on the food particles that the clams ingest, with assimilation efficiencies ranging from 10% for phytoplankton to as high as 90% for bacteria (26, 27). In this study, our objectives were to estimate the bioavailability of Cr(III) and Cr(VI) to the mussel Mytilus edulis, to estimate the relative contribution of dissolved and particulate uptake to the overall Cr accumulation in mussels, and to analyze the factors most responsible for Cr bioaccumulation in the mussels. We employed a kinetic approach to model Cr bioaccumulation in this mussel, which is extensively used for monitoring coastal contamination worldwide (2, 28). This kinetic model has been successfully applied to predict metal concentrations in mussel tissues (29). Unlike equilibrium partitioning models, the kinetic model can also quantify the relative importance of different pathways of metal accumulation, which permits a greater understanding of the mechanisms of metal bioavailability and bioaccumulation. The parameters necessary to construct the kinetic model (including metal assimilation efficiency, uptake rate from the dissolved phase, and efflux rates following uptake from food or water) were experimentally measured and then used to predict Cr concentrations in mussel tissues, which were compared to field measurements of Cr concentrations in mussels.
Materials and Methods Bioaccumulation of Cr(III) and Cr(VI) in Phytoplankton. Four species of phytoplankton were obtained from the Provasoli and Guillard Phytoplankton Culture Collection (Bigelow Laboratory for Ocean Sciences, West Boothbay Harbor, ME) and maintained in unialgal clonal cultures; species included the chlorophyte Chlorella autotrophica (clone CCMP243), the dinoflagellate Prorocentrum minimum (CCMP 696), the prasinophyte Tetraselmis levis (CCMP 896), and the diatom Thalassiosira pseudonana (CCMP1335, 3H). Cells of each species were resuspended into 0.2-µm filtered surface seawater (200 mL) enriched with f/2 (30) nutrient levels for N, P, Si, and vitamins, and f/20 nutrient levels for trace metals without EDTA, Cu, and Zn (31). Initial cell densities were 8 × 104, 2.1 × 104, 6 × 104, and 8 × 104 cells mL-1 or 1.6, 8.4, 6.4, and 2.3 mg dry wt L-1 for C. autotrophica,
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TABLE 1. Calculated Concentration Factors/Partition Coefficients (DCF or Kd) of Cr(III) and Cr(VI) for Phytoplankton and Natural Sedimentsa SA vol SA/vol Cr (µm2) (µm3) (µm-1) species
particle type
Phytoplankton Chlorella autotrophica
68
1.10
Prorocentrum minimum
737 1942
0.38
Tetraselmis levis
327
595
0.55
65
76
1.17
Thalassiosira pseudonana
75
III VI III VI III VI III VI
1.3 × 105 5.0 × 102 1.2 × 104 4.2 × 102 2.8 × 104 1.9 × 102 5.6 × 104 4.7 × 102
III VI* VI*
3.4 × 104 9.4 × 102 2.3 × 103
Sediments Long Island Sound Long Island Sound Flax Pond
DCF/Kd
a Mean surface area (SA), volume (vol), and surface to volume ratio of algal cells are also shown. An asterisk (*) indicates extraction measurements, which showed that Cr(VI) had been reduced to Cr(III) in the sediments.
P. minimum, T. levis, and T. pseudonana, respectively. Mean cell surface areas and volumes were determined for aliquots of cells with a Coulter Multisizer (Table 1). Each flask then received 7.4 kBq of 51Cr(III) (corresponding to 0.1 nM) or 27.8 kBq of 51Cr(VI) (corresponding to 0.3 nM) additions. The Cr isotopes, obtained from NEN, were 51Cr(III) (in 0.1 N HCl, specific activity 7.7 kBq ng-1) and 51Cr(VI), as Na2CrO4 (in distilled water, specific activity 9.1 kBq ng-1). The algae were then incubated at 15 °C on a 14:10 light/dark cycle, as described elsewhere (31). Periodically, the fraction of radioisotope associated with the algae was measured (32), and the cell growth was monitored by counting cells microscopically. The dry weight concentration factor (DCF, essentially a Kd for phytoplankton) was calculated as
DCF )
dpm 51Cr g-1 dry wt cells dpm 51Cr mL-1 dissolved
(1)
Assimilation of Cr in Mussels Fed Phytoplankton and Sediments Labeled with Cr(III) and Cr(VI). A recent study presented assimilation efficiencies of Cr(III) from seven species of phytoplankton (25) in Mytilus edulis; however, no studies have determined the assimilation of Cr(VI) from ingested food in mussels. We therefore radiolabeled C. autotrophica, P. minimum, T. levis, and T. pseudonana with 51Cr(VI) in 200 mL of 0.2-µm filtered seawater, as described above. The radioisotope addition was 74 kBq (corresponding to 0.8 nM) for each species. After 3-6 d growth (2-6 cell divisions), cells were removed from their radioactive water using Nuclepore membranes and resuspended into 30 mL of filtered unlabeled seawater. Surface-oxidized natural sediments were collected from Long Island Sound, or LIS (2% loss on ignition, ashed at 450 °C for 4 h) and Flax Pond (10% loss on ignition). After sieving through a 120-µm nylon mesh, the sediments were centrifuged and then frozen (1 year) until use. Sediments were thawed and equilibrated in oxygenated seawater at 20 °C for 3 d; 10 mg was then radiolabeled with either 51Cr(III) (18 kBq) or 51Cr(VI) (74 kBq) in 10 mL of 0.2 µm filtered seawater for 5 d (pH 7.8-8.0). Sediments were swirled periodically to prevent development of reduced microenvironments. The partitioning of Cr between the sediments and the dissolved phase at the end of 5 d radiolabeling was measured, and partition coefficients (Kd) were determined (32). The radiolabeled sediments were then collected by centrifugation and resuspended into 5 mL of filtered unlabeled seawater. The Cr species in the labeled sediments were determined using a modification of the method of Cranston and Murray (33).
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Briefly, Cr(III) and Cr(VI) were extracted from the sediment with 1.2 N HCl for 1 min (with an efficiency >90% for both Cr species), the 0.2 µm filtrate was collected, 100 nM Fe(III) (FeCl3 in 0.1 N HCl) was added, and the pH was adjusted to 8.0 with NaOH. Coprecipitation of Cr(III) with the Fe(III) was complete in 1 min. This solution was filtered, and particulate and dissolved fractions, corresponding to Cr(III) and Cr(VI) fractions, were γ-counted (see below). Mussels (Mytilus edulis) of 3.5 cm shell length were collected from LIS, New York, and acclimated to laboratory conditions (28‰ seawater, 15 °C) for 1 week prior to experiments. During this period, mussels were fed the diatom Thalassiosira pseudonana. For the assimilation experiments, radiolabeled phytoplankton and sediments were pulse-fed to the mussels for 0.5 h, as described elsewhere (25, 29). Briefly, six mussels were placed in 700 mL of 0.2 µm filtered seawater. The radiolabeled phytoplankton and sediments (1.5 mg) were added every 5 min. At these food concentrations, no pseudofeces production was observed. After 0.5 h radioactive feeding, mussels were rinsed with filtered seawater, and the radioactivity of each individual was counted. To monitor the release of the radioisotopes from the mussels, five mussels from each treatment were then placed individually into depuration chambers held within an enclosed recirculating seawater system (34). To purge the guts of boluses of undigested food, log-phase cells of the diatom T. pseudonana (unlabeled) were continuously pumped into the aquarium with a peristaltic pump to provide a food ration of about 2% of their tissue dry wt d-1. The radioactivity of mussels was monitored over a 4-d period. Fecal pellets were collected frequently to minimize desorption of Cr from feces into the water and subsequent recycling. The assimilation efficiency (AE) of ingested Cr was calculated as the percentage of ingested Cr retained in the mussels after 3 d of depuration; mussels complete metal digestion and assimilation within 3 d (34). Influx Rates of Cr in Mussels from the Dissolved Phase. 51Cr(III) or 51Cr(VI) was added into 28‰ 0.2 µm filtered seawater (collected from the Atlantic Ocean 8 km off Southampton Long Island and diluted with Milli-Q water) and equilibrated with stable Cr overnight prior to experiments. The added dissolved concentrations of Cr [either Cr(III) (CrCl3) or Cr(VI) (Na2CrO4)] totaled (radioactive plus stable) 2, 10, 40, or 200 nM. The background Cr concentration of Atlantic surface waters was assumed to be 3.3 nM [mostly Cr(VI)] (17). Thus, the total dissolved Cr(VI) concentration was calculated using its background concentration and the dilution with Milli-Q water for salinity adjustment to 28‰. Two individual mussels were placed into 500 mL of seawater containing stable Cr and 51Cr radiotracer for 2 h. There were eight replicate individuals for each concentration treatment. The short-term exposure minimized the decline in mussels’ ventilation rates and the Cr complexation by mussel metabolites in the dissolved phase; both can lead to an underestimation of metal uptake rates from the dissolved phase (29). After 2-h exposure, mussels were dissected, and the radioactivity and dry weight of soft tissues were determined. The Cr uptake rate constants from the dissolved phase were then calculated from the experimentally determined relationship between the metal influx rate and dissolved metal concentration. Mussels were also acclimated to different salinities (15, 20, 28, and 35‰) for a period of 10 d. After this acclimation, it was presumed that there was no significant difference in ventilation rates among different salinity treatments (35). The dissolved uptake rates of Cr(III) and Cr(VI) were then determined at 10 nM, as described above. Cr Efflux from Mussels following Uptake from Dissolved Phase and Ingested Food. Ten individual mussels were exposed to dissolved 51Cr(III) or 51Cr(VI) for 7 d or to 51Cr(III)-labeled diatoms for 8 d. In the experiments assessing
uptake from the dissolved phase, mussels were exposed to 51Cr(III) (3.7 kBq in 2 L of 0.2-µm filtered seawater, corresponding to 4.6 pM) or 51Cr(VI) (6.4 kBq in 2 L of 0.2-µm filtered seawater, 6.6 pM) for a period of 16-18 h each day and then transferred to unlabeled seawater and fed with the diatoms T. pseudonana for another 6-8 h. In the food exposure experiments, mussels were fed with 51Cr(III)-labeled log-phase diatoms (T. pseudonana) for 6-8 h each day (6-7 additions of radiolabeled diatoms), and then transferred to unlabeled seawater for another 16-18 h. The diatoms were radiolabeled as described above. An average daily ingestion rate of 0.6 mg was calculated for each individual mussel. The seawater for radiolabeled exposure was changed each day. On day 7 (dissolved exposure) or day 8 (food exposure), mussels were rinsed with unlabeled seawater and analyzed for their radioactivity. In addition, mussels were further rinsed with 0.1 mM EDTA, but this treatment only removed a small fraction of shell-bound Cr(III) ( T. levis > T. pseudonana. Algal size had no apparent effect on Cr(VI) assimilation in mussels. Influx Rates of Cr(III) and Cr(VI) in Mussels from Dissolved Phase. Relationships of influx rate (Iw) of Cr(III) and Cr(VI) from the dissolved phase with the dissolved concentration of Cr (Cw) are shown in Table 3 and Figure 3. The slope describing the log-log relationship between Iw and Cw is close to 1, suggesting that dissolved uptake conformed with a Freundlich adsorption isotherm. The intercept of this log-log relationship represents the dissolved uptake rate constant (ku or Iw/Cw). Values of ku can be compared among different species and metals and under various physicochemical conditions and can be considered an index of metal bioavailability from the dissolved phase. Thus, the uptake by mussels of dissolved Cr(III) is about 3 times lower than that of Cr(VI). The DCFs of Cr for mussels, calculated by dividing the dissolved uptake rate constant (ku) by the efflux rate constant (ke) (Tables 3 and 4), are 2800 for Cr(III) and 9100 for Cr(VI). The influx rates of dissolved Cr(VI) in mussels remained relatively constant over the salinity range of 20-35‰ (Figure 4). When the salinity was 15‰, however, the influx rate was 4.2 times higher than at 35‰. The influx rates of Cr(III) decreased slightly with increasing salinity, but there was no significant difference among the different salinity treatments (P > 0.05) (Figure 4). Efflux Rates of Cr from Mussels. Depuration of Cr following 7 d uptake from the dissolved phase or 8 d food ingestion conformed with a two- or three-compartment exponential loss pattern (Figure 5). Compartmental analysis of each pool is shown in Table 4. The efflux from the slowest exchanging compartment represented the long-term physiological turnover of Cr from the mussels. There was no significant difference in Cr efflux rate between Cr(III) and Cr(VI) following uptake from the dissolved phase (P > 0.05). Most of the Cr(III) (85.4 ( 2.0%) was on the shells after 7 d uptake from the dissolved phase, and by the end of depuration this had increased to 93.0 ( 3.5%. For Cr(VI), only 22.2 ( 5.3% and 41.3 ( 10.6% were on the shells after 7 d dissolved uptake and at the end of depuration, respectively. Concurrent measurements of Cr(III) depuration from empty shells (dissected after 7 d dissolved uptake) indicated that the efflux from shells (0.012 d-1) was identical to whole mussels (shell plus soft tissues) (0.012 d-1, Table 4). The depuration rate of Cr(VI) from mussel shells was very
TABLE 3. Relationships Describing Cr Influx Rate (Iw, µg g-1 d-1) in Mussels and Dissolved Cr Concentration (Cw, µg L-1) in Seawatera absorption efficiency (%) 2
Cr
equation
r
III VI
Iw ) 0.034*[Cw]0.885 ( 0.097 (SE) Iw ) 0.100*[Cw]0.948 ( 0.086 (SE)
0.976 0.984
ku range (L
g-1
d-1)
0.025-0.048 0.078-0.128
mean
range
0.029 0.085
0.017-0.055 0.051-0.192
a The ranges of dissolved uptake rate constants (k ), calculated using regression of log-transformed influx rates and dissolved concentrations, u are also shown. Absorption efficiencies were calculated by dividing ku by the mussel’s filtration rate (mean 117 L g-1 d-1, range 52-196 L g-1 d-1) (56).
FIGURE 3. Influx rates of Cr(III) and Cr(VI) in mussels from the dissolved phase at different ambient Cr concentrations. Mean (SD (n ) 8). low after 7 d, so the efflux rates calculated for whole mussels probably represented tissue depuration of Cr(VI). The efflux rate (0.010 d-1, from slowest exchanging pool) of Cr(III) following 8 d food ingestion (Table 4) did not differ significantly from the efflux rate following 7 d dissolved uptake (P > 0.05). In this experiment, 89.2 ( 1.2% of Cr(III) was in the soft tissues after 8 d food ingestion, and the efflux rate measured for whole mussels therefore represented depuration from soft tissues. On day 16 of depuration, dissection of one mussel showed that 78.5% of Cr(III) was still in the soft tissues, and this remained essentially constant until the end of the depuration period (79.2 ( 5.9%). Modeling Analysis of Cr Bioaccumulation in Mussels. Kinetic modeling of Cr bioaccumulation in mussels described in eq 5 requires measurements of ku, AE, IR, kew, kef, and g. Because the growth rate constant g is generally an order of magnitude lower than the efflux rate constant and cannot be accurately predicted (this parameter depends greatly on season, size, and diverse ecological conditions), it was ignored in our model calculations. When total suspended solid loads are >5 mg L-1 (e.g., in San Francisco Bay, or SFB), a maximum ingestion rate of 0.27 g g-1 dry wt d-1 is assumed for mussels (29). AEs of Cr(III) from different particle types (phytoplankton and sediments) range between 0.2 and 1.3% (25, this study). Intermediate AEs of 0.5% and 1% were therefore included in the model calculations. Values of ku, kew, and kef were taken from Tables 3 and 4. The kinetic model also requires measurements of geochemical factors such as the Cr concentrations in the dissolved (Cw) and particulate (Cf) phases. Few reliable measurements of Cw and Cf are available from estuarine and coastal waters. For our model calculations, we used data for SFB where sampling and measurements were done using trace metal clean techniques (20). Dissolved Cr(VI) concentrations in south SFB are relatively constant, with a mean concentration of 2.1 nM, but great uncertainty exists for Cf. The Cr concentration in food particles using a weak acid digestion may be more realistic for modeling Cr accumulation in bivalves because trace metal assimilation in marine bivalves
is affected by metal desorption within the acidic gut (pH 5-6) (34, 37). Cr concentrations in south SFB surface sediments, measured using a 24-h 0.5 N HCl extraction, range between 12 and 24 µg g-1, with an average concentration of 17 µg g-1 (38). These values were used as Cf in our modeling. The Cr concentrations in SFB mussels have been monitored by the NOAA’s National Status and Trends Program annually since 1986. The Cr(III) concentration in the dissolved phase was consistently undetectable at salinities >18% in SFB (20). Thus, Cr(III) uptake from the dissolved phase contributes little to Cr accumulation in mussels in south SFB. The contribution of Cr(VI) from the particulate phase should also be negligible because Cr(VI) is not particle-reactive (DCF ) 200-500 for phytoplankton, Table 1). In addition, Cr(VI) can be reduced to Cr(III) when it is adsorbed onto sedimentary particles. It was therefore assumed that all Cr measured on suspended particles was Cr(III). Consequently, the model calculation only considered two source terms, Cr(III) via food ingestion and Cr(VI) via uptake from the dissolved phase; Cr(III) from the dissolved phase and Cr(VI) from ingested food were assumed to be negligible. The Cr concentrations calculated with the model (Css, 2.67.5 µg g-1) using two AEs and Cf that are likely encountered by mussels in south SFB are remarkably close to the Cr concentrations in mussels measured in south SFB (Cmeasured, 3.0-5.1 µg g-1) (Table 5). The predicted contributions of Cr(VI) from the dissolved phase to the total Cr body burden in SFB mussels range between 13 and 38%; ingested Cr(III) on food is calculated to contribute the remaining 62-87%. We also analyzed the effects of variables (AE, Cf, total suspended solids [TSS] loads, salinity) on the Css in SFB, assuming a dissolved Cr(VI) concentration of 0.11 µg L-1 (Figure 6). In analyzing each variable, intermediate values of other parameters were used (AE ) 0.5%, Cf ) 17 µg g-1, IR ) 0.27 g g-1 d-1, ku ) 0.100 L g-1 d-1). The measured Cr influx rates at different salinities (Figure 4) were considered in this analysis. These calculations show that both AE and Cf are critical in affecting the Css in mussels and the contributions of Cr(VI) from the dissolved phase. The slopes describing the relationship between Css and AE, or between Css and Cf, were 4.6 and 0.14, respectively, suggesting that even a slight change in AE would result in a sharp change in Css and the fraction of Css derived from dissolved Cr(VI). For example, an increase in Cr(III) AE from 0.2% to 1.0%, within the range measured from various food particles, would lead to a Css increase from 1.9 to 5.6 µg g-1, and the fraction of this body burden coming from the dissolved phase (as Cr(VI)) would decrease from 52% to 18%. The TSS load appears to have no major effect on Css. In this analysis, we assume that TSS loads only affect mussel ingestion rates and that mussels maintain a maximum ingestion rate of 0.27 g g-1 d-1 at TSS loads > 5 mg L-1 (29). Both the Css and the fraction of body burden of Cr coming from the dissolved phase increase inversely with salinity due to a significant increase in the influx rate of Cr(VI) from the dissolved phase. At 15‰, the Css would increase by 2.5 times as compared to 35‰, and the fraction of body burden of Cr coming from the dissolved phase as Cr(VI) is calculated to increase from 31% to 79%.
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TABLE 4. Compartmental Analysis of Cr Depuration in Mussels following 7 d Dissolved Uptake or 8 d Ingestion of Radiolabeled Diatoms (T. pseudonana)a Cr
period (d)
% in compartment
k (d-1)
tb1/2 (d)
r2
Cr(III) dissolved
0-3 3-20 0-3 3-8 8-20 0-1 1-16 16-25 0-20 0-7 7-20
19.2 ( 6.3 80.8 ( 6.3 28.6 ( 5.3 14.9 ( 6.5 56.5 ( 10.2 17.8 ( 11.5 60.1 ( 15.8 25.9 ( 5.2 100 15.9 ( 4.6 84.1 ( 4.6
0.054 ( 0.019 0.012 ( 0.003 0.096 ( 0.020 0.028 ( 0.014 0.011 ( 0.004 0.164 ( 0.078 0.076 ( 0.006 0.010 ( 0.006 0.012 ( 0.000 0.020 ( 0.007 0.006 ( 0.004
14.3 ( 4.5 58.9 ( 14.1 7.5 ( 1.5 33.2 ( 17.2 74.8 ( 41.5 5.8 ( 3.6 9.2 ( 0.8 65.6 ( 23.2 56.2 ( 2.2 41.5 ( 16.8 271 ( 279
0.856 0.846 0.936 0.904 0.715 0.842 0.987 0.717 0.944 0.916 0.426
Cr(VI) dissolved Cr(III) food Cr(III) shell Cr(VI) shell
a Each compartment assumed a first-order depuration, conforming to A ) A exp(-kt), where A is the % retained at time t, A is the initial % o o retained, and k is the depuration rate constant. The biological half-life (tb1/2) of each compartment is also shown. Data are mean ( SD, n ) 7, for whole mussels and n ) 3 for shells. The depuration rate constant measured in the slowest compartment reflects physiological efflux. The data for Cr(III) release from dissected shells conformed with a one-compartment depuration model.
FIGURE 5. Depuration of Cr(III) and Cr(VI) in mussels following 7-d uptake from the dissolved phase or 8-d food feeding on radiolabeled diatoms (Thalassiosira pseudonana). Values represent % of Cr content in mussels immediately following bioaccumulation. Mean ( SD (n ) 7).
TABLE 5. Model-Predicted Cr Concentration in Mussel Tissues (Css) Using a Range of Measured Cf and AE Values and Comparison with Field-Measured Concentrations (Cmeasured) in Mussels Collected from South San Francisco Bay (Dumbarton Bridge Station, San Mateo Station, and Emeryville Station, 1991-1992)a
FIGURE 4. Influx rates of Cr(III) and Cr(VI) in mussels from the dissolved phase at different salinities. Mean ( SD (n ) 8).
Discussion Our measurements of Cr(III) uptake by four species of phytoplankton are directly comparable to earlier results for Cr(III) in marine phytoplankton (DCF ) 9 × 104) (39). Cellular fractionation in algae has shown that nearly all (>98%) of the cellular Cr(III) is on the cell wall/membrane (25). The cellular fractionation of Cr(VI) in the algae was not measured. The Kd of Cr(III) for sediments is comparable to the DCFs for phytoplankton, but the higher Kd of Cr(VI) for sediments may be due to its reduction to Cr(III). Previous studies showed that Cr(VI) is removed from the water column by particles due to its reduction to Cr(III) (40, 41). Possible mechanisms for Cr(VI) reduction include Fe(II)/organic matter oxidation and bacterial metabolic processes. The high Kd of Cr in organic-rich sediments suggests that organic matter plays a
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Cw Cr(VI) (µg L-1)
Cf Cr(III) (µg g-1)
AE (%)
Css (µg g-1)
Cmeasured (µg g-1)
0.11 0.11 0.11 0.11
12 24 12 24
0.5 0.5 1.0 1.0
2.6 4.2 4.2 7.5
3.0-5.1
% dissolved 38 24 24 13
a Data from National Status and Trends Program (T. P. O’Connor, personal communication). Dissolved and particulate (sediments) Cr concentrations were taken from refs 20 and 38, measured in April 1992. Only ingested Cr(III) and dissolved Cr(VI) are considered to contribute to Cr accumulation in mussels. Also shown are the proportion of total Cr body burden predicted to come from the dissolved phase. See text for detailed discussion of parameters.
role in Cr(VI) reduction, either directly or by facilitating its reduction by reducing Fe(III) to Fe(II) in surface mineral phases (11, 42). Reduction of Cr(VI) by Fe(II) occurs within seconds to minutes (11), whereas transformation by other reductants takes hours to months (42). The rapid increase in Cr reduction when sediments were thawed and warmed to 20 °C in oxygenated water is consistent with the idea that bacterial activity was at least partially involved in this process. Similarly, Cr(VI) may have been reduced to Cr(III) once it
FIGURE 6. Predicted Cr concentration in mussel tissues (Css) in San Francisco Bay and the relative contribution of dissolved Cr(VI) to the overall Cr accumulation in mussels as a function of variations in (A) assimilation efficiency (AE); (B) Cr concentration in food particles (Cf); (C) total suspended solids loads (TSS); and (D) salinity. The solid line represents the Css (scale on left ordinate) and the dotted line represents the percent of body burden from dissolved Cr(VI) (scale on right ordinate). The measured Cr concentrations in mussels in South San Francisco Bay between 1991 and 1992 were 3.0-5.1 µg g-1. was accumulated in the mussels in our study, as noted for other organisms (5); this was not checked. Assimilation efficiencies of ingested Cr(III) in mussels from sediment was similar to values for algal food (AEs in the range of 0.2-1.3%) (25). Most unassimilated Cr(III) is rapidly egested by mussels within the first 10 h, probably via intestinal digestion within the stomach (34). Very little Cr undergoes glandular digestion in the digestive gland. The sharp decline in Cr retention within the first few hours suggests that Cr desorption within the mussel’s acidic gut (leading to longer gut retention) does not affect Cr assimilation. Gut residence time, which can significantly affect the assimilation of some metals in mussels, is not correlated with Cr assimilation from phytoplankton food (25). Similarly, unlike other trace elements (e.g., Se, Zn), Cr shows no relationship to C assimilation in mussels from algal food (25). Although assimilation of ingested Cr(VI) was somewhat higher (1-10%) than Cr(III), these AEs are still low, and there is so little particulate Cr(VI) in natural waters, this is unlikely to be a significant source of Cr for mussels. The lower uptake rate of dissolved Cr(III) than Cr(VI) in mussels is consistent with studies showing that the cellular uptake of Cr(III) is at least 10 times lower than the Cr(VI) from equimolar solutions (5, 43). Thus, Cr(III) from both ingested food and the dissolved phase crosses biological membranes much less readily than Cr(VI). The mechanisms underlying this difference may be related to their different chemical behavior in seawater, where Cr(OH)30 and Cr(OH)2+ dominate the inorganic species of Cr(III) (44) but organically
complexed Cr(III) is also present in natural waters (10-60%) (19, 45). Cr(VI) exists as the chromate anion in seawater (44) and can be taken up through anionic membrane channels as phosphate or sulfate analogues (46). In fact, Cr(VI) toxicity to the diatom T. pseudonana is a function of the Cr to sulfate ratio in seawater (47). The inverse relationship between Cr influx rate and salinity, consistent with earlier studies (29, 48), is probably due to physiological responses to salinity, such as increased cell volume and membrane permeability at lower salinities (29). The pronounced differences between influx rates of Cr(III) and Cr(VI) at 15‰ further indicate that the reduced form is less able to cross membranes than Cr(VI). Both Cr(VI) and Cr(III) speciation are probably independent of salinity within the salinity range examined here. The measured efflux rate constants of Cr are comparable to values (0.019 d-1) for Cr(III) in Mytilus galloprovincialis (49) and to other trace elements in M. edulis, which are typically in the range of 0.01-0.03 d-1 and independent of the uptake route and duration of exposure (29, 50). Because mussels are chronically exposed to metals, metal loss from mussels may be dominated by the physiological turnover of metal in the slowest exchanging pools. After 2 weeks exposure to Cr(III), Cr in M. edulis can be found in the lysosomes where it is associated with P and S and trapped in an insoluble form, leading to very slow turnover rates (51). Our measurements suggest that Cr is accumulated by mussels from two major sources, Cr(VI) in the dissolved phase and Cr(III) in the particulate phase. Due to its very slow
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uptake and often undetectable dissolved concentration in many aquatic environments, we conclude that the uptake of dissolved Cr(III) contributes little to Cr accumulation in mussels. Similarly, although mussels have higher AEs of Cr(VI) than Cr(III) from food, its lower particle reactivity also leads to an insignificant contribution of Cr(VI) from the food pathway. The importance of the Cr(III) food pathway is largely due to its high concentration in suspended particles, despite its very low AE in mussels (25). Marine sediments are often enriched in Cr and may serve as a direct source of Cr for marine organisms. Therefore, any water quality criteria based on the concentration of dissolved Cr(III) would be inadequate because most Cr(III) uptake is from ingested food. Cr concentrations in the clam Scrobicularia plana and the polychaete Nereis diversicolor are directly proportional to Cr concentrations in sediment, which is likely the main Cr source for these species (52). Cr(III) in food was also found to be a more important source of Cr for sea urchin larvae than dissolved Cr(III) (53), although these studies did not determine the uptake of Cr(VI). Comparison of the model-predicted Cr concentrations with field measurements indicates that the kinetic model describes well the bioaccumulation of Cr in mussels and is probably applicable over spatial and temporal variations in the field. The kinetic approach in separating the relative contribution of Cr(III) and Cr(VI) to the total body burden can also help predict the toxicity of accumulated Cr, which is mostly related to Cr(VI) due to its strong oxidizing potential. In our model, dissolved Cr(VI) uptake contributed 13-38% of the total Cr body burden in mussels in SFB. However, at low salinities, especially at 15‰, most of a mussel’s Cr (>70%) is predicted to be from dissolved Cr(VI), which may increase the toxicity of the accumulated Cr to the mussel. In the kinetic model, Css is extremely sensitive to a small change in AE, underscoring the importance of determining AE in assessing metal accumulation in aquatic animals. Reports of AE values in the literature are rare, and most bioaccumulation studies fail to determine them. In our studies, Cr AEs were consistently
