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Oct 9, 2009 - Polybrominated dibenzo-p-dioxins (PBDDs) have recently been found in the Baltic Sea at concentrations 1000 times above that of the ...
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Environ. Sci. Technol. 2009, 43, 8245–8250

Polybrominated and Mixed Brominated/Chlorinated Dibenzo-p-Dioxins in Sponge (Ephydatia fluviatilis) from the Baltic Sea M A R I A U N G E R , †,‡ L I L L E M O R A S P L U N D , * ,† ¨ RN,† PETER HAGLUND,§ ANNA MALMVA KRISTINA ARNOLDSSON,§ AND O ¨ RJAN GUSTAFSSON† Department of Applied Environmental Science (ITM), Stockholm University, Department of Environmental Chemistry, Stockholm University, and Department of Chemistry, Umeå University

Received June 10, 2009. Revised manuscript received September 17, 2009. Accepted September 24, 2009.

Polybrominated dibenzo-p-dioxins (PBDDs) have recently been found in the Baltic Sea at concentrations 1000 times above that of the chlorinated analogs (PCDDs), yet their sources are undefined. Marine production of organobrominated compounds by sponges is well documented. The objective of the current study was to investigate the potential for an aquatic sponge (Ephydatia fluviatilis), common to the Baltic Sea, to produce PBDDs and other organobromine compounds in the field. Mono- to pentaBDDs as well as several mixed brominated/ chlorinated dibenzo-p-dioxins (Br/Cl-DDs), PCDDs and methoxylated polybrominated diphenyl ethers (MeO-PBDEs) were quantified in sponge from the SW Baltic. Concentrations of individual PBDDs in the range 1-80 ng per g extractable organic matter were similar as in blue mussels from the Baltic Sea and about 25 000 times higher than 2,3,7,8-tetraCDD. To the best of our knowledge, this is the first time Br/Cl-DDs are reported in biota from a background environment. While this study does not point out sponges as a dominant source, the concentrations of PBDDs in sponge relative to related anthropogenic compounds such as PBDEs and PCDDs as well as the relative abundance of brominated dioxins and furans strengthens the idea of natural production.

Introduction Polybrominated dibenzo-p-dioxins (PBDDs) have recently been found in several biological matrices in the Baltic Sea such as blue mussels, fish (1, 2) red algae, and cyanobacteria (3). Reported environmental PBDD concentrations are about 1000 times above that of their chlorinated counterparts (PCDDs) (2, 4), which have many well-known anthropogenic sources (5) but also some natural sources (6, 7). Mixed brominated/chlorinated dibenzo-p-dioxins (Br/Cl-DDs) have mainly been reported for sites in the immediate vicinity of * Corresponding author phone: +46-8-6747165; fax: +46-86747638; e-mail: [email protected]. † Department of Applied Environmental Science (ITM), Stockholm University. ‡ Department of Environmental Chemistry, Stockholm University. § Department of Chemistry, Umeå University. 10.1021/es901705r CCC: $40.75

Published on Web 10/09/2009

 2009 American Chemical Society

anthropogenic combustion (8, 9). The toxicological effects of PCDDs are well-known and include effects such as chloracne, altered liver function, and impairment of the immune system. Limited studies on the toxicological effects of PBDDs and Br/Cl-DDs indicate similar effects as for PCDDs (10-12). Hence, the high environmental occurrence of PBDDs calls for intensified studies on their sources and environmental behavior. PBDDs and the closely related polybrominated dibenzofurans (PBDFs) are formed anthropogenically during combustion of bromine-containing materials such as brominated flame retardants (BFRs) (9, 11, 13). Some production through photolytical reactions has also been reported from experiments with hydroxylated polybrominated diphenyl ethers (OH-PBDEs) (14). There is no intentional production of PBDDs. Since PBDDs are combustion products of BFRs such as polybrominated diphenyl ethers (PBDEs), their release into the environment can be expected to be significantly lower than that of PBDEs. However, concentrations of PBDDs in blue mussels from the Baltic Sea are higher than the PBDE level (2, 15). This has led to the suggestion that PBDDs found in Baltic biota, in contrast to their chlorinated counterparts, originate largely from natural sources (2). However, these sources of the ubiquitous PBDDs in the Baltic Sea ecosystem remain elusive. This study seeks to test the hypothesis that aquatic sponges, or their associated microorganisms (hereafter jointly referred to as sponges), are producers of PBDDs in the Baltic Sea. Aquatic sponges are well-known producers of a wide array of brominated organic compounds (16-19). Brominated metabolites have been found to represent as much as 12% of the dry weight of sponge (20). Both methoxylated polybrominated diphenyl ethers (MeO-PBDEs) and OH-PBDEs (20, 21) as well as some hydroxylated PBDDs are among the brominated compounds believed to be produced by sponges (17). Here we identify and quantify several classes of organobromine compounds, with a focus on PBDDs, in the sponge Ephydatia fluviatilis, a common freshwater species, in the Baltic Sea.

Materials and Methods Chemicals. A description of all chemicals and standards used for sample treatment can be found in the Supporting Information (SI). Extraction and Purification. The sample extraction and purification procedure is provided as SI. Samples and Instrumentation. Sample Collection. Sponges (Ephydatia fluviatilis) and blue mussels (Mytilus edulis) were collected in September 2007 in the Borgholm ¨ land off the Swedish east coast town harbor on the island O (N56°52′50”, E16°38′48”). E. fluviatilis is reported to be a freshwater species found throughout Europe and North America (22-24). Although plant like, sponges belong to phylum Porifera that consists of sessile filter feeding animals with porous bodies. Several kilograms of E. fluviatilis were readily recovered from constructions in the harbor. Sponges and blue mussels were scraped off and collected in string bags. On site cleaning consisted of pulling the bags through the water to rinse off loose parts followed by separation of mussels and sponges by hand. The cleaned samples were transported in coolers to the Stockholm-based laboratories where they were kept frozen at -18 °C until extraction. Mussels were also collected outside Mo¨nsterås on the Swedish mainland (N57°6′48”, E16°34′38”) in September as ¨ land (N57°04′13”, well as at a different location on O E16°51′18”) in November the same year. Sponges were VOL. 43, NO. 21, 2009 / ENVIRONMENTAL SCIENCE & TECHNOLOGY

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present also at the latter location although to a lesser extent and none were sampled. At the Mo¨nsterås site no traces of sponge could be detected. Instrumentation. Gas chromatography-mass spectrometry (GCMS) analysis of PBDEs, MeO-PBDEs and blue mussel PBDDs was performed on a Themoquest SSQ 7000 using electron capture negative ionization (ECNI) at 70 eV and an ion source temperature of 180 °C. Ammonia was used as reaction gas. The instrument was set to scan from 33 to 1000 m/z or in selected ion monitoring (SIM) mode at m/z 79 and 81 for identification and quantification purposes respectively. A Hewlett-Packard 5890A gas chromatograph with a nonpolar DB5 MS column (15 m, i.d. 0.25 mm, film thickness 0.1 µm) was used for separation. The oven temperature was programmed from 80 °C (2 min), 20 °C min-1 to 200 °C, 5.5 °C min-1 to 315 °C (5 min). Analysis of polybrominated dibenzo-p-dioxins and furans (PBDD/Fs), PCDD/Fs and Br/Cl-DD/Fs in sponge was performed using a Waters-Micromass Autospec Ultra gas chromatography-high-resolution mass spectrometry system (GC-HRMS) operated in the electron ionization (EI) mode at 36 eV and an ion source temperature of 250 °C. SIM was used to enhance the sensitivity. Two of the most intense ions of each molecular ion isotope distribution cluster were monitored, one as a quantification ion, the other as a qualifier ion. To further enhance the sensitivity, the run was timesegmented and the SIM descriptors changed over time to monitor only those compounds that were eluted in a given time segment. A nonpolar J&W DB5MS column (15 m, 0.25 mm i.d., 0.10 mm film thickness) were used to separate the penta- through hepta-BDD/F and a polar Supelco SP-2330 column (60 m, 0.32 mm i.d., 0.25 µm film thickness) to separate mono- through tetra-BDD/F, mono- through octaCDD/F, and Br/Cl-DD/Fs. All injections were made in the splitless mode (3 µL). The nonpolar column was operated at a constant head pressure of 80 kPa and with an oven temperature program of 190 °C (1 min), 10 °C min-1 to 320 °C (8 min). Similarly, the polar column was fed a constant flow of 1.0 mL min-1 helium and the oven was temperature programmed from 190 °C (2 min), 3 °C min-1 to 274 °C (11 min). In total, five injections had to be made on the latter

(polar) column, one for each of the following compound classes: PBDD/F, PCDD/F, monoCl-, diCl-, and triClBDD/F. In addition, accurate mass experiments were performed to verify the presence of Br/Cl-DDs in the purified sponge extracts. For each target, a SIM descriptor was constructed that allowed monitoring of the ion intensities at and around the expected (theoretical) m/z ratio (-24, -14, -6, -2, 0, +2, +6, +14, +24 milli mass units (mmu)). The resulting ion intensities (peak areas) were then plotted versus the m/z values and the apex of the curve was determined, which corresponds to the experimentally derived molecular weight of the ion. The quantifications were performed using the internal standard technique. Compounds lacking a matched reference standard were quantified vs a homologue with the same number and type of substitution. This was not possible in the two cases with less chlorinated DD/Fs and BrCl-DD/Fs,. Mono- through tri-CDD/Fs were therefore quantified vs 2,3,7,8-tetraCDD and Br/Cl-DD/Fs vs a PBDD/F with the same number of bromine substituents, assuming the same molar response factors. Validation. Laboratory blanks were prepared in parallel to the samples. For the main target compound class the PBDDs, the blanks were below limit of detection (LOD) for most congeners while three of the PBDDs were found in the blanks at low amounts relative to the samples (0.10%). Reported PBDF congeners were also mostly below LOD with two exceptions of 17 and 23%, respectively. The blank level of the BDE-47 was 22%. Reported values of all compounds present in the blanks have thus been corrected. No Br/ClDDs (LOD 3 pg g-1) or MeO-PBDEs (LOD 1 pg g-1) were detected in the blank. The reported concentrations were corrected for recovery yields of 80% ((4% n ) 5) for MeOPBDE and PBDE and 69% ((5% n ) 3) for PCDD/Fs and PBDD/F. Due to the lack of suitable 13C-labeled PBDD/F standards, chlorinated 13C-labeled congeners were used as internal standards for both compound classes. Results from earlier method development experiments have shown that the PCDD/Fs and PBDD/Fs behave similarly during the sample cleanup and no discrimination was observed.

FIGURE 1. Chromatogram (GC-HRMS SIM) of dioxin fraction from sponge showing (A) Br/Cl-DDs (m/z 295.9239; 377.8304; 455.7409) (B) mono- to tetraBDFs (m/z 245.9680; 341.8714; 419.7809; 499.6904) (pentachlorinated biphenyls (PeCBs) interfere with diBDFs) and (C) mono- to tetraBDDs (m/z 261.9629; 325.8765; 403.7869; 483.6954). Numbers refer to the position of the bromine substituents and full names are presented in Table 1 and SI Table S2. 8246

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TABLE 1. List of Compound Names, Abbreviations Used in the Text and Concentrations (ng gEOM-1 ± std. dev.) of the Most Abundant Brominated Dibenzo-P-Dioxins Found in Sponge from the Baltic Sea. Two MeO-PBDE Congeners and BDE-47 Are Also Shown. Concentrations Are Corrected for Recoveries and Blank Levels compound name

abbreviation

PBDDs 2,7-dibromo-dibenzo-p-dioxin/ 2,8-dibromo-dibenzo-p-dioxin 1,8-tribromo-dibenzo-p-dioxin 1,3,7-tribromo-dibenzo-p-dioxin 1,3,8-tribromo-dibenzo-p-dioxin tribromo-dibenzo-p-dioxin with unknown substitution pattern #1 2,3,7-tribromo-dibenzo-p-dioxin 1,2,4,7-tetrabromo-dibenzo-p-dioxin/ 1,2,4,8-tetrabromo-dibenzo-p-dioxin 1,2,3,7-tetrabromo-dibenzo-p-dioxin 1,2,3,8-tetrabromo-dibenzo-p-dioxin

2,7/2,8-diBDD 1,8-diBDD 1,3,7-triBDD 1,3,8-triBDD

concentration(ng gEOM-1) 3.2 ((0.18 n ) 3) 0.87 ((0.05 n ) 3) 43 ((6.1 n ) 3) 84 ((10 n ) 3)

3U1

1.8 ((0.14 n ) 3)

2,3,7-triBDD

4.5 ((0.44 n ) 3)

1,2,4,7/1,2,4,8-tetraBDD

3.1 ((0.70 n ) 3)

1,2,3,7-tetraBDD 1,2,3,8-tetraBDD

3.2 ((0.97 n ) 3) 1.7 ((0.41 n ) 3)

Br/Cl-DDs monochloro-dibromo-dibenzo-p-dioxin with unknown substitution pattern no. 2 monochloro-dibromo-dibenzo-p-dioxin with unknown substitution pattern no. 3 monochloro-dibromo-dibenzo-p-dioxin with unknown substitution pattern no. 4 monochloro-tribromo-dibenzo-p-dioxin with unknown substitution pattern #6 2′-methoxy-2,3′,4,5′-tetrabromodiphenyl ether 6-methoxy-2,2′,4,4′-tetrabromodiphenyl ether 2,2′,4,4′-tetrabromodiphenyl ether

ClBr2DD-1

0.76 ((0.12 n ) 3)

ClBr2DD-2

2.3 ((0.24 n ) 3)

ClBr2DD-3

1.3 ((0.22 n ) 3)

ClBr3DD-6

1.9 ((0.45 n ) 3)

MeO-PBDEs and PBDEs 2′-MeO-BDE68 6-MeO-BDE47 BDE-47

55 ((12 n ) 4) 93 ((11 n ) 4) 4.7 ((13 n ) 4)

Results and Discussion Identification of Organobromines in Sponge. Mono- to penta-BDDs, mono- to hexa-BDFs as well as some Br/ClDDs and MeO-PBDEs were identified in the sponge sample. PCDD/Fs were also analyzed for comparison. Figure 1 shows gas chromatogram of mono- to tetra-BDDs, mono- to tetraBDFs and monochloro mono- to tri-BDDs. Additional chromatograms showing MeO-PBDEs as well as a list of all identified compounds is available in SI Figure S1 and Tables S1 and S2. The most abundant PBDDs were 1,3,8-triBDD and 1,3,7-triBDD whereas the most abundant MeO-PBDEs were 6-MeO-2,2′,4,4′-tetrabromodiphenyl ether (6-MeOBDE47) and 2′-MeO-2,3′,4,5′-tetrabromodiphenyl ether (2′MeO-BDE68) (Table 1). MeO-PBDEs have been reported in biological matrices from many geographically separated locations. The most abundant congeners are generally 6-MeO-BDE47 and 2′MeO-BDE68 although their relative amounts vary (25-27). These two compounds were isolated from North Atlantic whale samples and were shown to be produced naturally through measurements of their natural abundance 14C signal (26). PBDDs have to our knowledge been reported in much higher concentrations in environmental samples from the Baltic Sea (2) than in samples from elsewhere (28, 29). While other studies have implied natural production of brominated dibenzo-p-dioxins in aquatic sponges, the reported compounds have been substituted not only with bromine but also with hydroxyl or methoxyl groups (17, 30). Mixed halogenated Br/Cl-DDs have mainly been reported in combustion products such as fly ash (31). Some monoBr-PCDDs have also been detected in sediments from Japan (32), Hong Kong, and Korea (33). Although other studies have looked for Br/Cl-DDs in biological matrices (29, 34) they have not previously been found in ambient biota. Here, ClBr3DDs were first tentatively identified in sponge from the Baltic Sea by their ECNI mass spectra (Figure 2). Subsequent GCEI-HRMS analysis revealed a series of compounds with retention time windows consistent with ClBrx-DDs. The

FIGURE 2. Mass spectrum (ECNI) of ClBr3-DD in sponge. Marked peak (*) in SI Figure S1A corresponds to this spectrum. presence of ClBrDDs, ClBr2-DDs, and ClBr3-DDs were also supported by the GC-HRMS accurate mass experiments, which indicated a m/z of 295.924, 375.832, and 453.743 amu for the M+, [M+2]+ and [M+2]+ ions, respectively, of these compounds. All these values are within (1 mmu of the corresponding theoretical value. Since there are several possible congeners for the ClBrx-DDs it has not been possible to determine the positions of the halogen substituents. Quantification and Relative Abundance of Organobromines. Among the brominated dibenzo-p-dioxins, three diBDD, four triBDD, four tetraBDD, and four Br/Cl-DDs were present in the sponge at concentrations higher than 0.5 ng gEOM-1. The extractable organic matter (EOM) metric is commonly used as a proxy for the bulk lipid and lipid-like matrices that are amenable to accumulate persistent lipophilic compounds (e.g., ref 35). The concentrations of these congeners are listed in Table 1 together with those of 2′MeO-BDE68, 6-MeO-BDE47 and BDE-47. The concentrations of several other PBDDs and Br/Cl-DDs are available in the SI Table S2 and Figure 3 gives an overview of the concentrations of all halogenated dibenzo-p-dioxins quantified in the sponge. Selected PBDDs, MeO-PBDEs, and BDE-47 were also quantified in the blue mussel samples to allow a comparison with organobromine contents in sponge (SI Table S3). VOL. 43, NO. 21, 2009 / ENVIRONMENTAL SCIENCE & TECHNOLOGY

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FIGURE 3. Overview of all dibenzo-p-dioxins/furans quantified in sponge. Observe that the z-axis is truncated for visibility of compounds present in low concentrations. The level of triBDDs given in the figure is for the sum of four isomers and should not be confused with the slightly lower value provide in Table 2 and in the text, which is for the sum of just the two most abundant isomers. The two most abundant PBDD congeners, 1,3,7-triBDD and 1,3,8-triBDD (hereafter jointly referred to as TriBDD) are often reported together and their total concentration (127 ng gEOM-1) was 26 times higher than the concentration of the most abundant flame retardant, BDE-47 (4.7 ng gEOM-1). 6-MeO-BDE47 (93 ng gEOM-1) was present at 20 times the concentration of the anthropogenic BDE-47. The concentrations of the other brominated dibenzo-pdioxins listed in Table 1 were between 0.76 ng gEOM-1 and 3.2 ng gEOM-1. The concentrations of diBDDs and tetraBDDs found here for aquatic sponge were in the same range as those previously reported for blue mussels in the Baltic Sea (2). For Br/Cl-DDs, these are the first reported concentrations in biota from the background environment. The TriBDD concentration is at least 25 000 times higher than the concentration of the toxic 2,3,7,8-tetrachloro dibenzo-p-dioxin (TCDD) in sponge, as calculated from the limit of detection of the TCDD (LOD 5 pg gEOM-1). Nevertheless, the levels of TCDD and other PCDD/Fs in fatty fish from the Baltic Sea frequently exceeds the European Union maximum residue limits for food (36). Based on limited reports of TCDD in blue mussels, a TriBDD/TCDD ratio between 9000 and 65 000, which is similar to the ratio found in the aquatic sponge, can be estimated (37, 38). The mean TriBDD concentrations in blue mussels for the Baltic Sea collected from the literature is 162 ng gEOM-1 ((77, n ) 7). This includes blue mussels collected for reference as part of this study in the same year and general area as the sponge samples. The TriBDD level found in sponge was in the same range as in blue mussels (Figure 4a). The 6-MeO-BDE47 is always more abundant than the BDE-47 although there are large variations (Figure 4b). The mean 6-MeO-BDE47/ BDE-47 ratio in blue mussels from the Baltic Sea was 106 ((73, n ) 6). The ratio in sponge was 20 ((2.6, n ) 4), which is more similar to the ratio found in red algae (Figure 4b). However, blue mussels from sites with sponge also showed a lower ratio. The much higher levels of 6-MeO-BDE47 than the anthropogenic BDE-47 found in blue mussels point to a natural source of MeO-PBDEs. Further, the large variations in the concentration ratio are also consistent with a local in situ source. Similarly, the TriBDD to BDE-47 ratios in biota largely follow the same pattern as the 6-MeO-BDE47 to BDE-47 ratio (Figure 4c). As for the MeO-PBDEs, the variations in the ratio 8248

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FIGURE 4. TriBDD (sum of 1,3,7-triBDD and 1,3,8-triBDD), MeO-BDE47 and BDE-47 in the Baltic Sea. (A) Concentration of TriBDD (ng gEOM-1); (B) ratio of 6-MeO-BDE47 to BDE-47 concentrations; (C) ratio of TriBDD to BDE-47 concentrations. Comparison of sponge (black square) to blue mussels from the same year (white diamond) and previously published blue mussel (black diamond) (2) and algae (triangle) (3) data. Concentrations of TriBDD in blue mussels from 2007 as well as all BDE-47 and 6-MeO-BDE47 concentrations are available as SI Table S3. Circled white diamond represents blue mussel sample from the mainland site. show that the major sources of PBDDs and PBDEs are unrelated and the relative concentrations indicate a natural production of PBDDs. Taken together, the levels of MeO-PBDE and PBDDs relative to anthropogenic BDE-47 are quite similar for blue mussels and sponges, which are both filter-feeding organisms. The relative concentration seems to be slightly lower in the red algae (Figure 4c), which is a primary producer. Octanol-Water Partition Coefficients and Bioaccumulation Potentials. Bioaccumulation of organohalogens is partly regulated by their lipid solubility, which can be approximated by the octanol-water partition coefficient (log Kow). Regrettably, there is a shortage of empirical measurements of Kow for the MeO-PBDEs and PBDDs targeted in this study. This physicochemical property can however be estimated for any compound of known structure with the fragment contribution method (39, 40). As a starting compound for the PBDDs we used TCDD, for which a log Kow

TABLE 2. Octanol-Water Partitioning Coefficients (Log Kow) for Selected Brominated Compounds Calculated with the Fragment Contribution Model (39) Using TCDD (Log Kow 6.88) and BDE-47 (Log Kow 6.81) As Starting Compounds for the Halogenated Dibenzo-P-Dioxins and MeO-PBDEs, Respectively compound/compound class

log Kow

DiBDD TriBDD TetraBDD ClBr2DD ClBr3DD 6-MeO-BDE47/2′-MeO-BDE68

6.08 6.97 7.86 6.73 7.62 6.89

value of 6.88 has been established (41). This yield log Kow values for diBDD of 6.08 and tetraBDD of 7.86, with the triBDD and Br/Cl-DDs in between (Table 2). Estimating the log Kow from all individual fragments of the molecules yielded, for these halogenated dioxins, values that agreed very well (within 0.1 log unit) with the preferred method of “rebuilding” from the empirical TCDD value. For the MeO-PBDEs, we used as a starting compound the BDE-47 molecule for which Braekeveld and co-workers have established a log Kow value using long-term slow-stirring (42). The calculation suggests a log Kow of 6.89 for 6-MeO-BDE47 (Table 2) compared to 6.81 for BDE-47. There is a recently published log Kow value for MeO-BDE47 in Yu et al. (43) of 7.19, derived using a reverse-phase high-pressure liquid chromatography method. Unfortunately, that study employed a broad suite of calibration compounds representing different functional groups (e.g., PAHs) and thus interaction mechanisms with the stationary and mobile phases that are different than for the target MeO-PBDEs. Further, the stationary phase was an alkyl silane whereas, e.g., an octadecyl alcohol would have been a better choice to mimic molecular interactions with octanol. Hence, we recommend that the herewith estimated log Kow value for 6-MeO-BDE47 and 2′MeO-BDE68 should be used. The log Kow values for PBDDs and MeO-PBDEs indicate that they have the potential to bioaccumulate. Potential Sources and Implications. PBDDs have never been intentionally produced but are formed during combustion of bromine-containing materials such as PBDEs. During combustion of PBDEs generally more PBDF than PBDD is formed (44). The ratio varies depending on combustion temperature and degree of bromination where higher brominated PBDEs yield a higher ratio of PBDFs over PBDDs. However, when PBDE is mixed with a polymer the amount of PBDDs become negligible and almost 100% PBDFs is formed, most likely since the polymer consumes the oxygen needed for dioxin formation (44). However, in the sponge, several PBDD but no PBDF congeners were present in concentrations higher than 0.5 ng gEOM-1 (SI Table S2). This is a strong indication that the major source of PBDDs in the Baltic Sea is not anthropogenic combustion of bromine-containing materials. Further support to the idea of natural production of PBDDs is found in the bromine substitution pattern. All PBDDs found in high concentrations in the sponge have bromine substitution patterns consistent with condensation of brominated phenols along the path suggested by Haglund et al. (2). Alternatively, PBDDs may form through internal cyclization (with loss of HBr) from OH-PBDEs. Through this reaction the two major PBDD congeners, 1,3,7-triBDD and 1,3,8-triBDD, could form from naturally occurring 6-OHBDE47 and 2′-OH-BDE68, respectively. Also, two monochloro OH-tetraBDEs (OH-ClBr4-DEs) have been found in Baltic blue mussel and algae (45) that could be precursors to the Br/Cl-DDs found in the sponge. The present evaluation suggests that the high levels of PBDDs in the Baltic Sea stem largely from natural production. Since the concentrations of both PBDD and MeO-PBDEs in sponge were similar to what is found in blue mussels, it does not seem that the E. Fluviatilis is a major natural source. Biological and microbiological studies of this aspect of the Baltic Sea environment could increase our knowledge and the rapid development of compound specific isotope analysis (CSIA) may also shed further light on the origins of these types of compounds. The documented high levels of bioaccumulated brominated and mixed brominated/chlorinated compounds in Baltic biota underscores the importance for further studies of both source apportionment and environmental behavior, including ecotoxicological effects.

Acknowledgments Ulla Eriksson is gratefully acknowledged for technical assistance. Henrik Petersson at Stenninge Bygg & Dyk is acknowledged for the sampling of sponge and mussels. KarlGustav Ottosson is acknowledged for mussel sampling and Laura Sanchez-Garcia also assisted with sample collection. Roland Engkvist at the University of Kalmar identified the Ephydatia fluviatilis. This project was financially supported by the Swedish Research Council for Environment, Agricultural Sciences and Spatial Planning (FORMAS project 216¨ .G.) and the Stockholm Marine Research Center 2006-565 to O ¨ .G also acknowledges support as an Academy (SMF). O Researcher from the Swedish Royal Academy of Science through a grant from the Knut and Alice Wallenberg Foundation.

Supporting Information Available Materials and methods, additional chromatograms, and tables with all quantified compounds (46), (47), (48). This material is available free of charge via the Internet at http:// pubs.acs.org.

Literature Cited (1) Malmvarn, A.; Zebuhr, Y.; Jensen, S.; Kautsky, L.; Greyerz, E.; Nakano, T.; Asplund, L. Identification of polybrominated dibenzo-p-dioxins in blue mussels (Mytillus edulis) from the Baltic Sea. Environ. Sci. Technol. 2005, 39, 8235–8242. (2) Haglund, P.; Malmvarn, A.; Bergek, S.; Bignert, A.; Kautsky, L.; Nakano, T.; Wiberg, K.; Asplund, L. Brominated dibenzo-pdioxins: A new class of marine toxins. Environ. Sci. Technol. 2007, 41, 3069–3074. (3) Malmvarn, A.; Zebuhr, Y.; Kautsky, L.; Bergman, A.; Asplund, L. Hydroxylated and methoxylated polybrominated diphenyl ethers and polybrominated dibenzo-p-dioxins in red algae and cyanobacteria living in the Baltic Sea. Chemosphere 2008, 72, 910–916. (4) Domingo, J. L.; Bocio, A. Levels of PCDD/PCDFs and PCBs in edible marine species and human intake: A literature review. Environ. Int. 2007, 33, 397–405. (5) Baker, J. I.; Hites, R. A. Is combustion the major source of polychlorinated dibenzo-p-dioxins and dibenzofurans to the environment? A mass balance investigation. Environ. Sci. Technol. 2000, 34, 2879–2886. (6) Holmstrand, H.; Gadomski, D.; Mandalakis, M.; Tysklind, M.; Irvine, R.; Andersson, P.; Gustafsson, O. Origin of PCDDs in ball clay assessed with compound-specific chlorine isotope analysis and radiocarbon dating. Environ. Sci. Technol. 2006, 40, 3730– 3735. (7) Hoekstra, E. J.; De Weerd, H.; De Leer, E. W. B.; Brinkman, U. A. T. Natural formation of chlorinated phenols, dibenzop-dioxins, and dibenzofurans in soil of a Douglas fir forest. Environ. Sci. Technol. 1999, 33, 2543–2549. (8) Weber, R.; Kuch, B. Relevance of BFRs and thermal conditions on the formation pathways of brominated and brominatedchlorinated dibenzodioxins and dibenzofurans. Environ. Int. 2003, 29, 699–710. (9) Sovocool, G. W.; Mitchum, R. K.; Tondeur, Y.; Munslow, W. D.; Vonnahme, T. L.; Donnelly, J. R. Bromo-polynuclear and bromochloro-polynuclear aromatic-hydrocarbons, dioxins and dibenzofurans in municipal incinerator fly-ash. Biomed. Environ. Mass. 1988, 15, 669–676. (10) Birnbaum, L. S.; Staskal, D. F.; Diliberto, J. J. Health effects of polybrominated dibenzo-p-dioxins (PBDDs) and dibenzofurans (PBDFs). Environ. Int. 2003, 29, 855–860. (11) World Health Organization. Polybrominated Dibenzo-P-Dioxins and Dibenzofurans; Environmental Health Criteria (205): Geneva, Switzerland, 1998. (12) Olsman, H.; Engwall, M.; Kammann, U.; Klempt, M.; Otte, J.; van Bavel, B.; Hollert, H. Relative differences in aryl hydrocarbon receptor-mediated response for 18 polybrominated and mixed halogenated dibenzo-p-dioxins and -furans in cell lines from four different species. Environ. Toxicol. Chem. 2007, 26, 2448– 2454. (13) Buser, H. R. Polybrominated dibenzofurans and dibenzo-ParadioxinssThermal-reaction products of polybrominated diphenyl ether flame retardants. Environ. Sci. Technol. 1986, 20, 404–408. VOL. 43, NO. 21, 2009 / ENVIRONMENTAL SCIENCE & TECHNOLOGY

9

8249

(14) Steen, P. O.; Grandbois, M.; McNeill, K.; Arnold, W. A. Photochemical formation of halogenated dioxins from hydroxylated plybrominated diphenyl ethers (OH-PBDEs) and chlorinated derivatives (OH-PBCDEs). Environ. Sci. Technol. 2009, 43, 4405–4411. (15) Asplund, L.; Nylund, K.; Eriksson, U.; Bignert, A. Monitoring of PBDE, metoxylated PBDE and PCB in blue mussels from the swedish coast line. Organohal. Comp. 2007, 69, 1713–1716. (16) Gribble, G. W. Naturally occurring organohalogen compounds. Acc. Chem. Res. 1998, 31, 141–152. (17) Utkina, N. K.; Denisenko, V. A.; Scholokova, O. V.; Virovaya, M. V.; Gerasimenko, A. V.; Popov, D. Y.; Krasokhin, V. B.; Popov, A. M. Spongiadioxins A and B, Two new polybrominated dibenzo-p-dioxins from an Australian marine spongeDysidea dendyi. J. Nat. Prod. 2001, 64, 151–153. (18) Melcher, J.; Janussen, D.; Garson, M. J.; Hiebl, J.; Vetter, W. Polybrominated hexahydroxanthene derivatives (PBHDs) and other halogenated natural products from the mediterranean sponge Scalarispongia scalaris in marine biota. Arch. Environ. Contam. Toxicol. 2007, 52, 512–518. (19) Thompson, J. E.; Barrow, K. D.; Faulkner, D. J. Localization of 2 brominated metabolites, aerothionin and homoaerothionin, in spherulous cells of the marine sponge Aplysina-fistularis ()verongia-thiona). Acta Zool. 1983, 64, 199–210. (20) Unson, M. D.; Holland, N. D.; Faulkner, D. J. A brominated secondary metabolite synthesized by the cyanobacterial symbiont of a marine sponge and accumulation of the crystalline metabolite in the sponge tissue. Mar. Biol. 1994, 119, 1–11. (21) Cameron, G. M.; Stapleton, B. L.; Simonsen, S. M.; Brecknell, D. J.; Garson, M. J. New sesquiterpene and brominated metabolites from the tropical marine sponge Dysidea sp. Tetrahedron 2000, 56, 5247–5252. (22) Gaino, E.; Rebora, M.; Corallini, C.; Lancioni, T. The life-cycle of the sponge Ephydatia fluviatilis (L.) living on the reed Phragmites australis in an artificially regulated lake. Hydrobiologia 2003, 495, 127–142. (23) Okland, K. A.; Okland, J. Freshwater sponges (Porifera: Spongillidae) of Norway: Distribution and ecology. Hydrobiologia 1996, 330, 1–30. (24) Lauer, T. E.; Spacie, A. New records of freshwater sponges (Porifera) for southern Lake Michigan. J. Great Lakes Res. 1996, 22, 77–82. (25) Melcher, J.; Olbrich, D.; Marsh, G.; Nikiforov, V.; Gaus, C.; Gaul, S.; Vetter, W. Tetra- and tribromopbenoxyanisoles in marine samples from Oceania. Environ. Sci. Technol. 2005, 39, 7784– 7789. (26) Teuten, E. L.; Xu, L.; Reddy, C. M. Two abundant bioaccumulated halogenated compounds are natural products. Science 2005, 307, 917–920. (27) Covaci, A.; Losada, S.; Roosens, L.; Vetter, W.; Santos, F. J.; Neels, H.; Storelli, A.; Storelli, M. M. Anthropogenic and naturally occurring organobrominated compounds in two deep-sea fish species from the mediterranean sea. Environ. Sci. Technol. 2008, 42, 8654–8660. (28) Fernandes, A.; Dicks, P.; Mortimer, D.; Gem, M.; Smith, F.; Ield, M. D.; White, S.; Rose, M. Brominated and chlorinated dioxins, PCBs and brominated flame retardants in Scottish shellfish: Methodology, occurrence and human dietary exposure. Mol. Nutr. Food Res. 2008, 52, 238–249. (29) Ashizuka, Y.; Nakagawa, R.; Hori, T.; Yasutake, D.; Tobiishi, K.; Sasaki, K. Determination of brominated flame retardants and brominated dioxins in fish collected from three regions of Japan. Mol. Nutr. Food Res. 2008, 52, 273–283. (30) Utkina, N. K.; Denisenko, V. A.; Virovaya, M. V.; Scholokova, O. V.; Prokof’eva, N. G. Two new minor polybrominated dibenzop-dioxins from the marine sponge Dysidea dendyi. J. Nat. Prod. 2002, 65, 1213–1215. (31) Chatkittikunwong, W.; Creaser, C. S. Bromo-dibenzo-p-dioxins, bromochloro-dibenzo-p-dioxins and chloro-dibenzo-p-dioxins and dibenzofurans in incinerator fly-ash. Chemosphere 1994, 29, 559–566.

8250

9

ENVIRONMENTAL SCIENCE & TECHNOLOGY / VOL. 43, NO. 21, 2009

(32) Takahashi, S.; Sakai, S.; Watanabe, I. An intercalibration study on organobromine compounds: Results on polybrominated diphenylethers and related dioxin-like compounds. Chemosphere 2006, 64, 234–244. (33) Terauchi, H.; Takahashi, S.; Lam, P. K. S.; Min, B. Y.; Tanabe, S. Polybrominated, polychlorinated and monobromo-polychlorinated dibenzo-p-dioxins/dibenzofurans and dioxin-like polychlorinated biphenyls in marine surface sediments from Hong Kong and Korea. Environ. Poll. 2009, 157, 724–730. (34) Dejong, A. P. J. M.; Vanderheeft, E.; Marsman, J. A.; Liem, A. K. D. Investigation on the occurrence of polyhalogenated (Br/Cl) dibenzodioxins and dibenzofurans in cows milk and fish tissue. Chemosphere 1992, 25, 1551–1557. (35) Sobek, A.; Reigstad, M.; Gustafsson, O. Partitioning of polychlorinated biphenyls between arctic seawater and sizefractionated zooplankton. Environ. Toxicol. Chem. 2006, 25, 1720–1728. (36) The Council of the European Union. Commission regulation (EC) No 1881/2006 of 19 December 2006 Setting Maximum Levels for Certain Contaminants in Foodstuffs; Official Journal of the European Union: Geneva, Switzerland, 2006. (37) Broman, D.; Naf, C.; Rolff, C.; Zebuhr, Y.; Fry, B.; Hobbie, J. Using ratios of stable nitrogen isotopes to estimate bioaccumulation and flux of polychlorinated dibenzo-p-dioxins (Pcdds) and dibenzofurans (Pcdfs) in 2 food-chains from the northern Baltic. Environ. Toxicol. Chem. 1992, 11, 331–345. (38) Ostfeldt, P.; Gustavson, K.; Jansson, B.; Jonsson, P.; Miettinen, V.; Ringstad, O.; Wese´n, C. Halogenated Organic Compounds in the Marine Environment 1989-1990; TemaNord: Copenhagen, 1994; p 591. (39) Meylan, W. M.; Howard, P. H. Atom fragment contribution method for estimating octanol-water partition-coefficients. J. Pharm. Sci. 1995, 84, 83–92. (40) Schwarzenbach, R. P.; Gschwend, P. M.; Imboden, D. M. Organic Liquid - Water partitioning: The n-Octanol - Water Partition Constant; Environmental Organic Chemistry., Wiley-Interscience: Hoboken, NJ, 2003; 223-235. (41) Aberg, A.; MacLeod, M.; Wiberg, K. Physical-Chemical property data for dibenzo-p-dioxin (DD), dibenzofuran (DF), and chlorinated DD/Fs: A critical review and recommended values. J. Phys. Chem. Ref. Data 2008, 37, 1997–2008. (42) Braekevelt, E.; Tittlemier, S. A.; Tomy, G. T. Direct measurement of octanol-water partition coefficients of some environmentally relevant brominated diphenyl ether congeners. Chemosphere 2003, 51, 563–567. (43) Yu, Y. J.; Yang, W. H.; Gao, Z. S.; Lam, M. H. W.; Liu, X. H.; Wang, L. S.; Yu, H. X. RP-HPLC measurement and quantitative structure-property relationship analysis of the n-octanol-water partitioning coefficients of selected metabolites of polybrominated diphenyl ethers. Environ. Chem. 2008, 5, 332–339. (44) Thoma, H.; Hauschulz, G.; Knorr, E.; Hutzinger, O. Polybrominated dibenzofurans (Pbdf) and dibenzodioxins (Pbdd) from the pyrolysis of neat brominated diphenylethers, biphenyls and plastic mixtures of these compounds. Chemosphere 1987, 16, 277–285. (45) Malmvarn, A.; Marsh, G.; Kautsky, L.; Athanasiadou, M.; Bergman, A.; Asplund, L. Hydroxylated and methoxylated brominated diphenyl ethers in the red algae Ceramium tenuicorne and blue mussels from the Baltic Sea. Environ. Sci. Technol. 2005, 39, 2990–2997. (46) Marsh, G.; Stenutz, R.; Bergman, A. Synthesis of hydroxylated and methoxylated polybrominated diphenyl ethers - Natural products and potential polybrominated diphenyl ether metabolites. Eur. J. Org. Chem. 2003, 2566–2576. (47) Jensen, S.; Reutergårdh, L.; Jansson, B. Analytical methods for measuring organochlorines and methyl mercury by gas chromatography. FAO Fish Tech. Pap. 1983, 212, 21–33. (48) Bignert, A.; Nyberg, E.; Asplund, L.; Eriksson, U.; Wilander, A. Comments Concerning the National Swedish Contaminant Monitoring Programme in Marine Biota; Swedish Museum of Natural History: Stockholm, 2006; www.nrm.se/download/18. 3f4924f310cc9243848000546/Marina_programmet2006.pdf.

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