Influence of the molecular structure and exposure concentration on the uptake and elimination kinetics, bioconcentration, and biotransformation of anionic and nonionic surfactants.

In vivo experiments with fish (sole) were performed for pure homologs of linear alkylbenzene sulfonates (LAS) and alcohol ethoxylates (AEO), the most widely used anionic and nonionic surfactants. The surfactant concentration measured in the organism was higher than in the experimental water, and increased with the exposure concentration for both compounds. At the exposure levels tested the bioconcentration factors (BCF) for AEO were one order of magnitude higher than for LAS. Two linear relationships for hydrophobicity were established, one with BCF and the other with uptake rate. The influence of the exposure concentration on the uptake (k(1)) and elimination (k(2)) velocities was researched. The value obtained for k(1) for AEO was higher than for LAS, while k(2) was very similar for both compounds, indicating differences in the incorporation and but not in the depuration rate. For the first time, internal degradation products of LAS were identified and quantified in fish and water and the glucuronic conjugate of AEO was detected in an organism. The predominant biotransformation process for these compounds may be different: the results suggest a phase I biotransformation for LAS and phase II for AEO, due to their different molecular structures.

Anaerobic degradation pathway of linear Alkylbenzene sulfonates (LAS) in sulfate-reducing marine sediments.

Linear alkylbenzene sulfonates (LAS) are among the principal synthetic surfactants used worldwide. Their presence in the environment has been reported in a significant number of studies, and it has been generally assumed that LAS are not biotransformed in the absence of oxygen. However, laboratory experiments performed by our group using anoxic marine sediments have reported LAS degradation percentages that can reach up to 79% in 165 days. Here, we show for the first time the initial reaction metabolites (generated via fumarate addition to the LAS molecules), their biotransformation into sulfophenyl carboxylic acids (SPC), and the progressive degradation of these by successive beta-oxidation reactions. Advanced mass spectrometry has been used to carry out the identification of these compounds. This is the first time that an anaerobic degradation pathway for LAS is described, and these results represent a significant advance in understanding the final fate of these and other similar compounds in anoxic environments.

Sources, transport and reactivity of anionic and non-ionic surfactants in several aquatic ecosystems in SW Spain: a comparative study.

Presence, distribution and transport mechanisms of the four major synthetic surfactants--linear alkylbenzene sulfonates (LAS), alkyl ethoxysulfates (AES), nonylphenol ethoxylates (NPEOs) and alcohol ethoxylates (AEOs)--have been simultaneously studied in different aquatic ecosystems. Urban wastewater discharges and industrial activities were identified as the main sources for these compounds and their metabolites. LAS, AES and carboxylic metabolites remained in the dissolved form (87-99%). However, NPEOs and AEOs were mostly associated with particulate matter (65-86%), so their degradation in the water column was limited due to their lower bioavailability. It was also observed that sorption to the particulate phase was more intense for longer homologs/ethoxymers for all surfactants. With respect to surface sediments, AES levels were considerably below (<0.25 mg/kg) the values detected for LAS and NPEOs. Concentrations of AEOs, however, were occasionally higher (several tens of ppm) than those found for the rest of the target compounds in several sampling stations.

Experimental determination of bioconcentration, biotransformation, and elimination of linear alkylbenzene sulfonates in Solea senegalensis.

The bioconcentration, biotransformation, and elimination of anionic surfactants, linear alkylbenzene sulfonates (LAS), specifically 2ØC(10) and 2ØC(12) LAS, which are pure isomers from two different homologues, have been characterized in Solea senegalensis using an experimental flow-through system with environmental exposure levels. The LAS and their carboxylated metabolites, sulfophenylcarboxylic acids (SPCs), were analyzed by pressurized liquid extraction, followed by solid-phase extraction, liquid chromatography with fluorescence detection, and mass spectrometry. The bioconcentration factors obtained in the steady state were 17.2 +/- 1.2 L/kg for 2ØC(10) LAS and 386.5 +/- 31.5 L/kg for 2ØC(12) LAS. A kinetic study also was performed with 2ØC(12) LAS, and the rate constants of uptake and elimination were K(1) = 177.04 L/kg d and K(2) = 0.92/d, respectively. The formation of the intermediate degradation products was monitored during the 2ØC(10) LAS experiment, thus allowing, to our knowledge for the first time, the identification and quantification of SPCs from 5ØC(6) to 9ØC(10) in marine organisms and in the depuration water as a result of biotransformation by the organisms, diffusive elimination, and excretion. The elimination percentage of the surfactants is both time- and concentration-dependent, and this suggests the existence of a concentration limit for the surfactant in the fish below which the elimination processes are slow. Overall, LAS are not persistent bioaccumulative compounds at environmental exposure levels.

Monitoring the primary biodegradation of linear alkylbenzene sulfonates and their coproducts in anoxic sediments using liquid chromatography-mass spectrometry.

An accompanying article has demonstrated the anaerobic degradation of the surfactant linear alkylbenzene sulfonate (LAS) in microcosms filled with marine sediments through the generation of sulfophenylcarboxylic acids (SPCs). A detailed study shows that this process was uniform in the blanks (non-spiked natural sediments) for every LAS homologue during the complete course of the experiment. However, when sediments were spiked with commercial LAS and, therefore, enriched with short-chain homologues, degradation was enhanced for these homologues until their percentages were close to those for non-spiked sediments. The reason is that short-chain homologues are more bioavailable due to their higher solubility and lower sorption capacity. Thus, sorption on sediments was found to be increased with the length of the alkyl chain for LAS homologues, following a linear Freundlich isotherm, whereas the metabolites generated were predominant in solution due to their much higher polarity. Intermediate-chain SPC homologues (C7-C9 SPCs) were the most abundant during the experiment, but a significant increase in the concentration of shorter-chain SPC homologues (C4-C6 SPCs) was detected toward the end. In the case of isomers, the steric effect of the aromatic group implies that LAS primary degradation took place preferentially over external isomers. Therefore, the generation of external isomers of SPCs was predominant during the complete experiment although internal isomers of SPCs became more evident when the degradation process had advanced and external isomers of LAS became scarce. The identity of both types of SPC isomer was confirmed by tandem mass spectrometry. With respect to LAS coproducts, the relative percentage of iso-LAS increased during the complete experiment and removal percentages for dialkyl tetralinsulfonates (<30%) were typically lower that those for LAS (66-79%), although a similar behavior was observed for their homologues in both cases.

Anaerobic degradation of linear alkylbenzene sulfonates in coastal marine sediments.

This research shows for the first time the degradation of linear alkylbenzene sulfonates (LAS) under anaerobic conditions, together with the presence of metabolites and the identification of microorganisms involved in this process. This compound is the most widely used surfactant and its main environmental concern is related to its persistence in the absence of oxygen as LAS accumulates in anaerobic sediments and sewage sludges. Laboratory experiments performed with anoxic marine sediments spiked with 10-50 ppm of LAS demonstrated, however, that its degradation reached 79% in 165 days via the generation of sulfophenyl carboxylic acids (SPCs). Almost all of the added LAS (>99%) was found to be attached to the sediment while the less hydrophobic SPCs were predominant in solution, as their concentration increased progressively up to 3 ppm during the full course of the experiment. Average half-life for LAS has been estimated to be 90 days, although higher values should be expected when the LAS concentration exceeds 20 ppm, due to inhibition of the microbial community. Sulfate-reducing and methanogenic activities proved to be intense during the experiment. Several sulfate-reducing bacteria and firmicutes/clostridia have been identified as possible candidates for effecting this degradation. Our results imply that the persistence of LAS in anoxic compartments, such as marine sediments, should be reconsidered when evaluating its environmental risk.

Biological adverse effects on bivalves associated with trace metals under estuarine environments.

Toxic effects of pollutants on marine organisms can be studied both by performing field measurements, and by undertaking laboratory simulation experiments. Here is described the effect of trace metals Zn, Cd, Pb and Cu on the clam Scrobicularia plana along a salinity gradient simulated in a hypothetical estuary using simulation experiments. The simulator produces a continuous entry of trace metals into the estuary through injection in the lower salinity tank of the system. The clams were exposed during two weeks to different concentration of trace metals to assess the bioaccumulation process along a salinity gradient. Bivalves were analysed for body tissue residue to determine the bioaccumulation factors related to each metal and the salinity influence was addressed. Differences among tanks were observed as a result of the salinity gradient. In the achieved assays, the mechanism of bioaccumulation of Zn and Cd in organisms was more efficient at high salinity values. Bioaccumulation factors for both metals showed a linear increase with the increase of salinity values. It seems that the mechanism of bioaccumulation of Pb and Cu in organisms was dependent on two simultaneous processes: the proximity to the input point of metals and the low salinity values.

Development of a method for the simultaneous analysis of anionic and non-ionic surfactants and their carboxylated metabolites in environmental samples by mixed-mode liquid chromatography-mass spectrometry.

A new methodology capable of performing the simultaneous analysis of the main surfactants--linear alkylbenzene sulfonates (LAS), alkyl ethoxysulfates (AES), alkyl sulfates (AS), nonylphenol polyethoxylates (NPEOs) and alcohol polyethoxylates (AEOs)--and their carboxylated metabolites--sulfophenyl carboxylic acids (SPCs) and alkylphenol ethoxycarboxylates (APECs)--in environmental samples has been developed for the first time. Extraction is carried out by solid-phase extraction (SPE) and pressurized liquid extraction (PLE) from water and sediment, respectively. Identification and quantification of the target compounds is performed using a liquid chromatography-mass spectrometry (LC-MS) system equipped with an electrospray interface (ESI) operating in mixed-mode. Optimization of parameters such as pH, ionic strength, temperature and solvents has been carried out in order to obtain recoveries in the range from 70 to 107% for most homologs, while the limits of detection are 0.05-0.5 ng mL(-1) in water and 1-10 ng g(-1) in sediment. The proposed methodology has been applied for the simultaneous determination of all the target compounds in samples taken from aquatic ecosystems in the SW of Spain. Values for LAS, AS, AES, NPEOs and AEOs are up to 38.7, 3.0, 2.9, 5.0 and 1.2 microg L(-1) in waters, and in the ranges of 1.73-12.80, 0.11-0.24, 0.02-0.59, 1.94-2.70 and 0.64-3.64 mg kg(-1) in sediments, respectively. The highest concentrations of metabolites found in water are 149.6 microg L(-1) of SPCs and 3.9 microg L(-1) of APECs.

Simultaneous extraction and determination of anionic surfactants in waters and sediments.

A new method has been developed for the simultaneous determination of the most frequently used anionic surfactants - linear alkylbenzene sulfonates (LAS), alkyl ethoxysulfates (AES) and alkyl sulfates (AS) - in aqueous and sediment samples. Preconcentration and purification of water samples are carried out by means of solid-phase extraction (SPE). The efficiency of two different extraction methods for the analysis of sediments - Soxhlet extraction and pressurized liquid extraction (PLE) - has been compared. Identification and quantification of the target compounds is performed using a liquid chromatography - mass spectrometry (LC-MS) system equipped with an electrospray interface (ESI) in negative ion-mode. Homologue recoveries are 85-123% for SPE, 94-112% for Soxhlet extraction and 81-125% for PLE in the case of LAS, and 60-94% for SPE, 61-109% for Soxhlet extraction and 55-99% for PLE in the case of AES, whereas the limits of detection are 0.1-0.5 ngml(-1) in water and 1-5 ngg(-1) in sediment. This method has been applied to the determination of anionic surfactants in the Guadalete estuary (SW Spain), and LAS concentration levels from 538 to 1014 ngg(-1) in sediments and from 25.1 to 64.4 ngml(-1) in waters have been found. AES values from 168 to 536 ngg(-1) in sediments and from 4.5 to 11.9 ngml(-1) in waters are reported for the first time in European rivers.

Determination and distribution of alkyl ethoxysulfates and linear alkylbenzene sulfonates in coastal marine sediments from the Bay of Cadiz (southwest of Spain).

The distribution of the two main anionic surfactants manufactured and used in the world, alkyl ethoxysulfates (AES) and linear alkylbenzene sulfonates (LAS), has been studied in sediments from a salt marsh and an estuary of the Bay of Cadiz (southwest of Spain). The identification and quantification of AES and LAS was carried out after automated Soxhlet extraction with methanol, followed by solid-phase extraction and liquid chromatography coupled to electrospray mass spectrometry. The latter procedure permitted the unequivocal identification of every LAS homologue as well as the AES homologues of up to 16 carbon atoms in their alkyl chain and of up to 12 ethylene oxide groups. Recoveries were in the range of 51% to 84% and limits of detection from 1 to 5 microg/kg. We have focused our attention particularly on AES because, in spite of their great use, these compounds have received less attention than LAS and their occurrence has not been described in marine environmental samples. Alkyl ethoxysulfates concentration values range between 100 and 400 microg/kg in the topmost layer of sediments at the sampling areas. The relative distribution of AES homologues shows higher percentages for the longer alkyl chain homologues in sediments as well as for the shorter ethoxymers. A decrease in LAS concentrations has been found relative to past studies in one of the sampling areas as a consequence of the reduction of urban wastewater discharges.

Sediment quality in littoral regions of the Gulf of Cádiz: a triad approach to address the influence of mining activities.

To evaluate sediment quality in different areas in the Gulf of Cádiz affected by mining activities, an integrative approach to assessment was used: the Sediment Quality Triad (SQT). Sediment samples were collected at six stations and subjected to replicated sediment acute and chronic toxicity tests, and comprehensive sediment chemistry analyses. Organisms collected synoptically at the same stations were analyzed for histopathological lesions and chemical concentrations in their tissues to determine 'in situ' alteration and bioavailability, in place of benthic community structure. The results obtained for each component were linked to the SQT using different methodologies of interpretation and expression of the integrated approach: (a) multivariate analysis, (b) significant statistical differences compared to the reference stations, (c) ANOVA-based pie charts, and (d) classical methods using triangles and the SQT index of pollution. SQT results indicated that the highest pollution was mainly associated with metals from mining activities and polyaromatic hydrocarbons in the Ría of Huelva. The use of histopathological measures and bioaccumulation of metals, improved the characterization of 'gray areas' of pollution and in the determination of the bioavailability of metals.

The influence of pH and salinity on the toxicity of heavy metals in sediment to the estuarine clam Ruditapes philippinarum.

An approach is presented for determining the influence of two key variables, pH and salinity (S), on the toxicity of four common heavy metals bound to sediments in estuaries. Two samples of environmental sediment taken from two estuaries in southern Spain (the Huelva estuary and the Guadalquivir River estuary), together with a dilution of toxic mud from the Aznalcóllar (Spain) mining spill (April 1998) were used to determine their toxicity at different values of pH (6.5, 7.5, and 8.5) and salinity (10, 20, and 30) on the estuarine clam Ruditapes philippinarum. Two different endpoints, sublethal, indicated by clam reburial (median effective burial time [ET50]), and relative mortality (median lethal concentration [LC50]), were used to quantify the toxicity associated with the heavy metals. Neither salinity nor pH was found to influence the toxic responses measured by the behavioral endpoint (ET50). However, a strong effect on the LC50 related to pH and salinity was detected, with the toxicity of the heavy metals being increased at low values of both variables (pH = 6.5 and S = 10). The mechanism of heavy metals uptake through water may explain this influence of pH and salinity on the lethal toxicity detected. The results show differences in the toxicity of these heavy metals bound to sediments depending on whether the origin of metal contamination is chronic or acute.

Biodegradation of linear alkylbenzene sulfonates and their degradation intermediates in seawater.

A study has been made of the aerobic biodegradation of linear alkylbenzene sulfonates (C12 and C11 homologues) and sulfophenylcarboxylic acids (C5 and C11 homologues) in seawater at concentrations of the same order as those detected ones in coastal waters influenced by wastewater effluents, at different temperatures, and both with and without the addition of an inoculum adapted to the presence of linear alkylbenzene sulfonate (LAS). The biodegradation of C12LAS, C11LAS, C5SPC, and C11SPC exceeds 99% in all tests performed and can be satisfactorily fitted to a second-degree polynomial without an independent term. The kinetic of degradation of LAS presents a clear seasonal component, since the process is considerably inhibited at lower temperatures; it is also kinetically enhanced by the presence of the inoculum. The intermediates detected for all the cases are sulfophenylcarboxylic acids (SPCs), the most abundant being those intermediates produced by the omega- and beta-oxidations of the parent compound, although intermediates produced by the alpha-oxidation have also been detected. The kinetic of the SPCs generated can be described using a model composed of two terms that represent the formation and the degradation of these intermediates. The total disappearance of the SPCs in all cases indicates that the degradation of LAS in seawater at the tested concentrations in aerobic conditions is complete.

Speciation of heavy metals in recent sediments of three coastal ecosystems in the Gulf of Cádiz, southwest Iberian Peninsula.

A live-step sequential extraction technique was used to determine the partitioning of Cr, Mn, Fe, Cu, Zn, Cd, and Pb among the operative sedimentary phases (exchangeable ions, carbonates, manganese and iron oxides, sulfides and organic matter. and residual minerals) in coastal sediment from three locations in the southwest Iberian Peninsula. Two sites are located close to industrial areas, the salt marshes of the Odiel River and Bay of Cádiz, and one in a nonindustrial area, the Barbate River salt marshes. The Odiel River salt marshes also receive the drainage from mining activities in the Huelva region. In the sediments from the Bay of Cádiz and Barbate River salt marshes, Cr, Cu, Fe, and Zn were extracted from the residual fraction at percentages higher than 60%. In the sediments from the Odiel River salt marshes, concentrations of all the metals, except Cu. Zn, and Cd, exceeded 60% in the residual fraction as well. In the sediments from the Bay of Cádiz and Barbate River salt marshes, the main bioavailable metals were Mn and Cd; in those from the Odiel River salt marshes, the main bioavailable metals were Zn and Cd, respectively. The environmental risk was determined by employing the environmental risk factor (ERF), defined as ERF = (CSQV - Ci/CSQV), where Ci is the heavy metal concentration in the first four fractions and CSQV is concentration sediment quality value (the highest concentration with no associated biological effect). Our results showed that the sediments from the Cádiz Bay and Barbate River salt marshes do not constitute any environmental risk under the current natural conditions. In contrast, in the Odiel River salt marshes, Cu, Zn, and Pb yielded ERFs of less than zero at several sampling stations and, consequently, pose a potential threat for the organisms in the area. This is a consequence of the high levels of metals in the area derived from the mining activity (pyrite) and industrial activities and the association of these heavy metals with more labile fractions of the sediments.

Seasonality of contamination, toxicity, and quality values in sediments from littoral ecosystems in the Gulf of Cádiz (SW Spain).

To seasonally evaluate littoral contamination, toxicity and quality values of sediments from the Gulf of Cádiz, we measured chemical concentrations and conducted toxicity tests in winter and summer and linked these results by means of multivariate analysis. Sediment samples were subjected to two separate, replicated sediment toxicity tests (Microdeutopus gryllotalpa amphipod survival, and Ruditapes philippinarum clam reburial), and to comprehensive sediment chemistry analyses (grain size, organic carbon, 14 heavy metals, and the surfactant linear alkylbenzenesulfonate (LAS)). Only sediments associated with an untreated urban discharge were toxic and related to high levels of surfactant LAS, Ag, and Pb. Multivariate analysis indicated that variables and chemicals associated with geochemical matrix and background levels (specific surface, Fe, Zn, Cu, V, Ni, and Co), chemicals associated with untreated urban discharge sources, and toxicity effects showed no seasonal variability. Only copper concentrations showed seasonal differences, being toxic during the winter and not toxic during summer. Multivariate analysis permits us to derive sediment quality values (SQVs); in terms of concentrations at or below which biological effects were not measured (mg kg(-1) dry sediment), are: LAS, 2.6; lead, 66.8; silver, 0.78; copper, 69.6.