Application of bioassays in toxicological hazard, risk and impact assessments of dredged sediments.

Given the potential environmental consequences of dumped dredged harbour sediments it is vital to establish the potential risks from exposure before disposal at sea. Currently, European legislation for disposal of contaminated sediments at sea is based on chemical analysis of a limited number of well-known contaminants for which maximum acceptable concentrations, action levels (ALs), have been set. The present paper addresses the issue of the applicability of in vitro and in vivo bioassays for hazard, risk and local impact assessment of dredged polluted sediments to be disposed of at sea. It discusses how and to what extent selected bioassays can fill in the gaps left open by chemical analysis and the way in which the bioassays may contribute to the present licensing system for disposal. Three different purposes for application were distinguished: the most basic application (A) is a rapid determination of the hazard (potential toxicity) of dredged sediments which is then compared to ALs in a licensing system. As with chemical analysis on whole sediment extracts, the bioavailability of the chemicals is not taken into account. As in vitro assays with sediment extracts are not sensitive to matrix effects, a selection of specific in vitro bioassays can be suitable fast and standardized additions for the licensing system. When the outcome of (A) does not convincingly demonstrate whether the sediment is clean enough or too polluted, further bioanalysis can help the decision making process (B). More aspects of the mostly unknown complex chemical mixtures are taken into account, including the bioavailability and chronic toxicity focusing on ecologically relevant endpoints. The ecotoxicological pressure imposed by the dredged sediments can be quantified as the potentially affected fraction (PAF) based on chemical or biological analysis of levels of contaminants in sediment or biota. To validate the predicted risk, the actual impact of dumped harbour sediments on local ecosystems (C) can be determined using a dedicated set of in vitro and in vivo bioassays as well as bio-indicators selected based on the information obtained from (A) and (B) and on the characteristics of the local ecosystem. Conversely, the local sediment impact assessment (C) can direct fine-tuning of the selection of chemical and bioassay analyses and for setting safe levels in the licensing system. It is concluded that in vitro and in vivo bioassays and biological indicators are useful tools in the process of hazard, ecotoxicological risk and impact assessment of dredged harbour sediments, provided they are consciously chosen and quality criteria for assay performance are defined.

Use of zeolite for removing ammonia and ammonia-caused toxicity in marine toxicity identification evaluations.

Ammonia occurs in marine waters including effluents, receiving waters, and sediment interstitial waters. At sufficiently high concentrations, ammonia can be toxic to aquatic species. Toxicity identification evaluation (TIE) methods provide researchers with tools for identifying aquatic toxicants. For identifying ammonia toxicity, there are several possible methods including pH alteration and volatilization, Ulva lactuca addition, microbial degradation, and zeolite addition. Zeolite addition has been used successfully in freshwater systems to decrease ammonia concentrations and toxicity for several decades. However, zeolite in marine systems has been used less because ions in the seawater interfere with zeolite's ability to adsorb ammonia. The objective of this study was to develop a zeolite method for removing ammonia from marine waters. To accomplish this objective, we performed a series of zeolite slurry and column chromatography studies to determine uptake rate and capacity and to evaluate the effects of salinity and pH on ammonia removal. We also assessed the interaction of zeolite with several toxic metals. Success of the methods was also evaluated by measuring toxicity to two marine species: the mysid Americamysis bahia and the amphipod Ampelisca abdita. Column chromatography proved to be effective at removing a wide range of ammonia concentrations under several experimental conditions. Conversely, the slurry method was inconsistent and variable in its overall performance in removing ammonia and cannot be recommended. The metals copper, lead, and zinc were removed by zeolite in both the slurry and column treatments. The zeolite column was successful in removing ammonia toxicity for both the mysid and the amphipod, whereas the slurry was less effective. This study demonstrated that zeolite column chromatography is a useful tool for conducting marine water TIEs to decrease ammonia concentrations and characterize toxicity.

Precision of dialysis (peeper) sampling of cadmium in marine sediment interstitial water.

Isolating and analyzing interstitial water (IW) during sediment toxicity tests enables researchers to relate concentrations of contaminants to responses of organisms, particularly when IW is a primary route of exposure to bioavailable contaminants by benthic dwelling organisms. We evaluate here the precision of sampling IW with the dialysis or 'peeper' method using sediments spiked with five different concentrations of cadmium. This method is one of several that are commonly used for collecting IW. Seven consecutive ten-day toxicity tests were conducted on these sediments and IW samples were collected at the end of each of these tests. Prior to each test initiation and insertion of IW samplers, sediments were allowed to equilibrate for seven days under flow-through conditions with filtered seawater. At the end of each ten-day testing period, peepers were retrieved, and IW cadmium measured. Data sets were organized by treatment and test number. Coefficients of variation (CV) for the six replicates for each sediment and testing period and for each sediment across testing periods (42 replicates) was used as a measure of sampling precision. CVs ranged from 25 to 206% when individual testing periods were considered, but ranged from 39 to 104% when concentrations for all testing periods were combined. However, after removal of outliers using Dixon's Criteria, the CVs improved and ranged from 6 to 88%. These levels of variability are comparable to those reported by others. The variability shown is partially explained by artifacts associated with the dialysis procedure, primarily sample contamination. Further experiments were conducted that support our hypothesis that contamination of the peeper causes much of the variability observed. If method artifacts, especially contamination, are avoided the dialysis procedure can be a more effective means for sampling IW metal.

Removal of ammonia toxicity in marine sediment TIEs: a comparison of Ulva lactuca, zeolite and aeration methods.

Toxicity Identification Evaluations (TIEs) can be used to determine the specific toxicant(s), including ammonia, causing toxicity observed in marine sediments. Two primary TIE manipulations are available for characterizing and identifying ammonia in marine sediments: Ulva lactuca addition and zeolite addition. In this study, we compared the efficacy of these methods to (1) remove NH(x) and NH(3) from overlying and interstitial waters and (2) reduce toxicity to the amphipod Ampelisca abdita and mysid Americamysis bahia using both spiked and environmentally contaminated sediments. The utility of aeration for removing NH(x) and NH(3) during a marine sediment TIE was also evaluated preliminarily. In general, the U. lactuca and zeolite addition methods performed similarly well at removing spiked NH(x) and NH(3) from overlying and interstitial waters compared to an unmanipulated sediment. Toxicity to the amphipod was reduced approximately the same by both methods. However, toxicity to the mysid was most effectively reduced by the U. lactuca addition indicating this method functions best with epibenthic species exposed to ammonia in the water column. Aeration removed NH(x) and NH(3) from seawater when the pH was adjusted to 10; however, very little ammonia was removed at ambient pHs ( approximately 8.0). This comparison demonstrates both U. lactuca and zeolite addition methods are effective TIE tools for reducing the concentrations and toxicity of ammonia in whole sediment toxicity tests.

Metal-colloid partitioning in artificial interstitial waters of marine sediments: influences of salinity, pH, and colloidal organic carbon concentration.

For decades, heavy metals have been deposited into marine sediments as a result of anthropogenic activities. Depending on their bioavailability, these metals may represent a risk to benthic organisms. Dissolved interstitial water metal concentrations have been shown to be better predictors of bioavailability than sediment metal concentrations. In order to improve our understanding of metals bioavailability in sediments, it is essential to fully comprehend metal speciation. Colloidal organic carbon is ubiquitous in marine interstitial water and readily forms complexes with numerous dissolved metals, greatly reducing their bioavailability. Methods were applied to isolate dissolved and colloidal cadmium, copper, nickel, lead, and zinc from interstitial waters using centrifugation and stirred cell ultrafiltration. Influences of salinity, pH, and colloidal carbon concentration on partitioning behavior were investigated. In the pH study, colloidal-dissolved partitioning of each metal (Kp) was observed to increase as pH rose. Conversely, in the salinity experiment, metal Kps declined as salinity increased. For the metal-organic colloid study, linear isotherms were calculated for each of the metals, resulting in log Kcocs ranging from 5.90 for zinc to 7.50 for copper. Kp values calculated from metal-spiked field sediments were in good agreement with those measured in the salinity study. These results provide insight into the behavior of metals associated with colloidal organic carbon in marine sediments under the conditions occurring in estuarine systems.

Use of Ulva lactuca to identify ammonia toxicity in marine and estuarine sediments.

Toxicity identification evaluation (TIE) methods are being developed for use with whole sediments. Although a phase I TIE method has been developed to characterize ammonia toxicity in aqueous samples using the marine macroalga Ulva lactuca, the relationship between amphipod and mysid mortality and uptake by U. lactuca of bedded sediment ammonia had not been explored. Additionally, it was not known how interactions in whole sediments between metal and organic contaminants with U. lactuca and ammonia would affect TIE interpretation. The current study showed that ammonia toxicity to amphipods and mysids was reduced significantly in marine sediments in the presence of U. lactuca. The alga slightly affected metal concentrations but did not alter the concentration of organic contaminants in either overlying or interstitial waters. Conversely, ammonia uptake by U. lactuca was not affected by the presence of metal or organic contaminants in the sediments. When used with other TIE manipulations, U. lactuca can be utilized in a whole-sediment, phase I TIE to remove toxicity due to ammonia.

Exploratory analysis of the effects of particulate characteristics on the variation in partitioning of nonpolar organic contaminants to marine sediments.

The partitioning of nonpolar organic contaminants to marine sediments is considered to be controlled by the amount of organic carbon present. However, several studies propose that other characteristics of sediments may affect the partitioning of contaminants. For this exploratory analysis, we measured 19 sediment characteristics from five marine sediments and 11 characteristics of humic acids extracted from the sediments. These characteristics included elemental composition, grain size, soot carbon, polarity indices and molar ratios. Each individual characteristic and combinations of these characteristics were then used to normalize partition coefficients (Kp) generated for three organic contaminants: lindane, fluoranthene and a tetrachlorinated biphenyl (PCB). A coefficient of variation (CV) was then calculated for each contaminant to determine which normalization characteristic (individually or in combination) resulted in the lowest variability in partitioning between study sediments. For lindane and the PCB. normalization by the amount of sediment organic carbon resulted in the lowest variability in partition coefficients with CVs of 16.2% and 37.7%. respectively. However, normalization of fluoranthene by silt content resulted in lower CVs than those generated by organic carbon normalization: 31.0% vs. 37.6%. Normalization of contaminants Kp's by combined values of sediment characteristics resulted in lower CVs but only by a few percent. Using humic acid characteristics, humic organic carbon reduced variability between sediments most effectively. But only the normalized fluoranthene values had a CV (i.e., 25.4%) lower than the one based on normalization by sediment characteristics. When combined, humic acid characteristics resulted in lower CVs than normalization by individual or combinations of sediment characteristics for fluoranthene and the PCB with CVs of 19.3% and 28.7%, respectively. This analysis indicates variability associated with the partitioning of some organic contaminants to marine sediments can be further reduced when normalization by sediment characteristics other than organic carbon are utilized.

Sediment toxicity assessment: comparison of standard and new testing designs.

Standard methods of sediment toxicity testing are fairly well accepted; however, as with all else, evolution of these methods is inevitable. We compared a standard ASTM 10-day amphipod toxicity testing method with smaller, 48- and 96-h test methods using very toxic and reference sediments. In addition we compared parallel exposures of single species, either the amphipod Ampelisca abdita or the mysid Americamysis bahia, to multiple species, mysid, and amphipod, cohabiting the same types of chambers. These comparisons were performed for both water-only and sediment-water tests. Results of the comparison of the standard ASTM 10-day amphipod test with the smaller, 48- and 96-h test chambers indicate that survival was high in both test designs using the reference sediment. With toxic sediments, complete mortality occurred in less than 48 h using the smaller experimental chambers and only after 96 h in the larger experimental chambers. We concluded that although time to death is shorter in the smaller, shorter exposure chambers, there was no overall change in mortality for the organisms, and that the smaller chambers were predictive of the results obtained with larger chambers and longer exposures. For multiple species testing in whole sediment exposures there was no change in toxicity to either the amphipod or the mysid when they cohabited the same chamber. In contrast, for water-only exposures, A. bahia demonstrated less sensitivity when cohabiting the same chamber as A. abdita. Therefore, during whole sediment testing we can add A. bahia and A. abdita to the same test chamber without changing the toxicity to either species; however, in our 10-ml water-only exposures, the species should be tested separately.

Ratio of the concentration of anthraquinone to anthracene in coastal marine sediments.

The ratio of the concentration of the oxidation product anthraquinone to that of its parent polycyclic aromatic hydrocarbon anthracene is reported for several coastal marine sediments. The ratio ranges from 0.317 in a highly contaminated industrialized harbor to 2.81 in a remote, less contaminated site. We hypothesize that differences in this ratio result from the input source of PAHs, with input from atmospheric deposition at remote sites resulting in a predominance of anthraquinone (ratio > 1), and direct discharge to highly contaminated industrialized harbors resulting in a predominance of anthracene (ratio < 1). To support this hypothesis, the fate of anthracene in the marine environment was investigated with respect to conversion to its oxidation product, anthraquinone. Once associated with sediments, anthracene is believed to be relatively persistent; however, it can potentially be subjected to oxidation via biological (microbial degradation) and chemical (chemical oxidation and photooxidation) processes. An assessment of the extent of oxidation of anthracene associated with sediments was conducted both under conditions simulating those found in the marine environment and under rigorous conditions by exposure to UV radiation. Results of this study show that while anthracene associated with marine sediments does not readily undergo oxidation to anthraquinone under conditions normally encountered in the marine environment, under extreme conditions anthracene is photooxidized by exposure to UV radiation. The extent of oxidation is influenced by sediment characteristics such as percent organic carbon, humic acid content and sediment surface area. The relative stability of anthracene under normal conditions may help to validate the use of the anthraquinone to anthracene ratio in marine sediments as an environmental marker of contaminant source.

Effects of sediment homogenization on interstitial water PCB geochemistry.

Sediment homogenization is a common practice in many contaminated sediment toxicity testing and chemical analysis protocols. A primary goal of sediment homogenization is to reduce inter-replicate variability. In this study, the geochemical effects of sediment homogenization were evaluated by measuring the concentration and distribution of polychlorinated biphenyls (PCBs) in environmentally contaminated marine sediment interstitial waters. Sediment homogenization, prior to isolation of interstitial waters, was found to significantly increase the concentration of PCBs in the dissolved and colloidal phases-generally by a factor of two. Long-term storage (i.e., several months) of sediments following mixing appeared to allow interstitial water distributions of PCBs to return to "normal," although a storage artifact may also be present. This study indicates that homogenization results in significant changes in the concentration of PCBs in environmentally contaminated sediment interstitial waters. Consequences of these changes on inferences made based on toxicity tests or chemical analyses using homogenized sediments need to be considered and studied further.

A comparison of the isoelectric focusing patterns of red cell phosphoglucomutase separated on an ampholine, ampholine/separator and immobilised pH gradient.

The isoelectric focusing patterns of red cell phosphoglucomutase (PGM1) separated on Ampholine, Ampholine/separator and rehydratable immobilised pH gradients have been compared. The sharpest and most intense zymograms have been observed on immobilised pH gradients provided that Ampholine was added during the rehydration of the gel. The addition of ampholine in the rehydration of Immobiline plates has been shown to improve the sharpness and intensity of the zymogram obtained and the potential of immobilised pH gradients for PGM typing has been demonstrated.

An improved means of enzyme typing of hair roots using isoelectric focusing.

An improved method of grouping hair, based on the alleles of PGM observed by isoelectric focusing, has been described. The increased discriminating power of this system (0.77) compared to that obtained by the starch gel technique (0.55) provides a new and more sensitive means of typing hair.