Article pubs.acs.org/est
Polyhalogenated Carbazoles in Sediments of Lake Michigan: A New Discovery Jiehong Guo,† Da Chen,*,‡ Dave Potter,§ Karl J. Rockne,∥ Neil C. Sturchio,⊥ John P. Giesy,# and An Li*,† †
School of Public Health, University of Illinois at Chicago, Chicago, Illinois 60612, United States Cooperative Wildlife Research Laboratory and Department of Zoology, Southern Illinois University, Carbondale, Illinois 62901, United States § Research Division, Wellington Laboratories, Guelph, Ontario N1G 3M5, Canada ∥ Department of Civil and Materials Engineering, University of Illinois at Chicago, Chicago, Illinois 60607, United States ⊥ Department of Earth and Environmental Sciences, University of Illinois at Chicago, Chicago, Illinois 60607, United States # Department of Veterinary Biomedical Sciences and Toxicology Centre, University of Saskatchewan, Saskatoon, Saskatchewan S7N 5B3, Canada ‡
S Supporting Information *
ABSTRACT: Previously unknown halogenated compounds were detected during the analysis of halogenated flame retardants in two sediment cores collected from Lake Michigan. Gas chromatography coupled with high- or low-resolution mass spectrometry (MS) was used to determine the chemical structures for a total of 15 novel polyhalogenated carbazoles (PHCs) with the general molecular formula C12H9-x-y-zNClxBryIz. On the basis of the mass spectra generated by electron impact (EI) and electron capture negative ionization (ECNI) MS, eight PHCs were tentatively identified as polybrominated carbazoles, while the others were mixed halogenated carbazoles containing, in addition to bromine, either chlorine or iodine or both. Patterns of halogen substitution of PHCs included Br2 to Br5, ClBr2, ClBr3, ClBr4, ClBr3I, Br4I, and Br3I2. 3,6-Dibromocarbazole and 1,3,6,8-tetrabromocarbazole were also found among the PHCs. Profiles of the concentration versus depth of sediment at the two sites showed various patterns among polybrominated carbazoles. The abundance of mixed halogenated carbazoles peaked at depths of 12−16 cm, remained at relatively constant levels in deeper sediment, but declined markedly in more recently deposited sediments. This is the first study discovering the seven mixed halogenated carbazoles in the environment. Detailed methods for their detection and identification are provided.
■
other aquatic organisms.3 For example, the soil fungus Penicillium sp. produces 2,4-dichlorophenol, and grasshoppers secrete 2,5-dichlorophenol.6 Methoxylated and hydroxylated polybrominated diphenyl ethers (PBDEs) as well as polybrominated phenols found in some marine organisms were proven to be of natural origin.7−10 Some naturally produced organohalogens have molecular structures similar to those of synthetic PBT chemicals, warranting more research on their environmental behavior and impact. A group of halogenated organic compounds, which could be of natural and/or anthropogenic origins, are polyhalogenated carbazoles (PHCs).11−14 Chemical structures of PHCs resemble those of polyhalogenated dibenzofurans, with a NH group replacing the oxygen. A number of studies have identified either chlorinated or brominated carbazoles in riverine or marine sediments as well as soils.11,14−18 In the Great Lake region, 1,3,6,8-tetrachlorocarbazole was discovered in the
INTRODUCTION Numerous anthropogenic organic chemicals contain halogens. According to the 2014 Chemical Data Reporting (CDR) information submitted to the United States Environmental Protection Agency (U.S. EPA), annual production of bromine (Br), chlorine (Cl), and iodine (I) in the United States was 2.1 × 105, 9.9 × 106, and 6.1 × 103 metric tonnes, respectively.1 In North America, 610 chemicals were prioritized for environmental monitoring, about 62% of which are halogenated organic compounds.2 These chemicals have caused global environmental and human health concerns, because of their persistent, bioaccumulative, and potentially toxic (PBT) nature. In addition, more than 3800 naturally occurring organohalogens have been identified from both biogenic and abiogenic sources.3 Natural processes, such as volcanic eruptions and forest fires, are known to produce various organohalogens, including congeners of polychlorinated biphenyls (PCBs).4 Chlorophenols are naturally produced but are also manufactured and used on a large scale in industry and are listed as priority pollutants by the U.S. EPA.5 Natural biogenic organohalogens are usually produced by organisms, such as bacteria, fungi, plants, marine and freshwater algae, and some © 2014 American Chemical Society
Received: Revised: Accepted: Published: 12807
August 12, 2014 September 24, 2014 October 1, 2014 October 1, 2014 dx.doi.org/10.1021/es503936u | Environ. Sci. Technol. 2014, 48, 12807−12815
Environmental Science & Technology
Article
sediment of the Buffalo River in 1984.18 In the sediment of Lake Michigan, a group of tri- to pentabrominated carbazoles were discovered and the most abundant congener was identified as 1,3,6,8-tetrabromocarbazole.16 Recently, a carbazole with both chlorine and bromine substituents was detected in sediments of streams across southern Ontario, Canada.19 In the present study, a number of carbazole-like compounds with a variety of halogen substitution patterns were discovered in sediments from Lake Michigan. The objectives of the study were to (1) identify and characterize structural information on these compounds and (2) reveal their temporal trends in sediments of Lake Michigan. To our knowledge, this is the first report on the presence of this suite of PHCs in the environment.
time of 10 min per cycle. The extract was concentrated by a rotary evaporator and solvent-exchanged into hexane. A glass column (11 mm inner diameter × 40 cm long) was prefilled with dichloromethane (DCM). The column was then filled from the bottom to the top with 1 g of granular anhydrous Na2SO4, 16 g of alumina, 4 g of silica gel, and 4 g of granular anhydrous Na2SO4. After the column was packed, DCM was completely replaced by hexane. Concentrated extract was added to the prepared columns and eluted with 100 mL of hexane (F1), 200 mL of 4:1 hexane/DCM mixture (F-2), and finally, 100 mL of DCM (F-3). F-2 and F-3 containing the target analytes were concentrated and placed in 200 μL glass inserts of GC vials, prior to analysis by gas chromatography (GC)−lowresolution mass spectrometry (LRMS) in electron impact (EI) or electron capture negative ionization (ECNI) mode. Initial analyses were performed on Agilent 6890/5973 GC− MS (Agilent Technologies, Santa Clara, CA) in ECNI mode. It was equipped with a Gerstel programmable temperature vaporization (PTV) injection port, operated in solvent vent mode. The inlet temperature started at 40 °C, held for 2 min, and then increased to 300 °C at 600 °C/min. The injection volume was 60 μL (20 μL × 3) for each run. The carrier gas was helium. The vent flow was 100 mL/min, and the purge flow was 100 mL/min, at 2.75 min. GC was equipped with a Restek Rtx1614 capillary column (15 m × 0.25 mm inner diameter × 0.10 μm film thickness). The initial oven temperature was set at 50 °C, held for 3 min, then increased to 300 °C at 10 °C/min, and kept for 10 min until the run was completed. The carrier gas flow was kept constant at 1.5 mL/ min. To achieve baseline separation of all peaks and ensure the quality of mass spectra of targeted analytes, a 30 m Agilent DB5MS capillary column (0.25 mm inner diameter × 0.25 μm film thickness) was also used. The initial oven temperature was 50 °C, held for 3 min, and then increased to 150 °C at 10 °C/min and further to 300 °C at 5 °C/min. The flow of carrier gas was kept constant at 1.2 mL/min. For both columns, the temperature of the GC−MS interface was kept at 300 °C. Full scan mass spectra were obtained in LRMS with ECNI (m/z 35−800) or EI (m/z 50−800) ionization. The temperature of the ECNI ion source was set at 200 °C, and methane was used as reagent gas. The temperature of the EI ion source was 230 °C. The quadrupole analyzer was maintained at 150 °C during ECNI and EI analyses. To achieve adequate responses from the accurate mass determination using GC−high-resolution MS (HRMS), 16 g of dry composite sediment from Lake Michigan was extracted. The extract was fractionated in the same silica gel/alumina column but with slightly different eluting solvents, i.e., 100 mL of hexane (F-1), 60 mL of hexane/DCM mixture (4:1, v/v) (F2), and 240 mL of hexane/DCM mixture (4:1, v/v) (F-3). F-3 contained target analytes, was concentrated using a rotary evaporator, and was placed in 200 μL glass inserts of GC vials prior to analysis. Accurate mass determination was performed using Agilent 6890 GC coupled to Waters Autospec Ultima MS. The fraction containing target analytes was injected into the GC−HRMS system and analyzed in full-scan mode (m/z 50−1000). Voltage scan experiments were created to scan narrow mass ranges that encompassed molecular fragments of interest. These narrow mass ranges also included one to three reference peaks from the mass calibrant, perfluorokerosene (PFK). The instrument was tuned to a resolution of at least 10 000. A spectrum calibration curve was constructed using the PFK reference mass peaks in a
■
EXPERIMENTAL SECTION Sampling. Two sediment cores were collected in Lake Michigan onboard the U.S. EPA Research Vessel (R/V) Lake Guardian. One was collected using a box corer in September 2010 at site M018 (latitude, 42.7338°; longitude, −86.9995°) in the southern basin. The other was collected by a multi-corer in June 2011 from M041 (latitude, 44.7367°; longitude, −86.7215°) in the deep Chippewa basin. At each site, four subcores (10 cm in diameter) were obtained and sectioned using hydraulic extruders with 1 cm intervals until 10 cm and then 2 cm intervals to a depth of 30 cm. To minimize potential “smearing”, sediment within 2 mm of the wall of the coring tube was trimmed off and discarded. After each segment was cut, all sectioning gear was thoroughly cleaned using tap water, acetone, and deionized water. At each site, the segments at corresponding depths were combined and mixed with stainlesssteel spoons in glass bowls. Well-mixed composite samples were distributed into pre-cleaned 125 mL amber glass jars with Teflon-liner screw caps. The samples were immediately frozen. They were transported in coolers to the laboratory, where they were stored at −20 °C until further processing and analysis. Chemicals and Reagents. Chemical standards 3,6dibromocarbazole (3,6-DiBC) (97%) and 1,3,6,8-tetrabromocarbazole (1,3,6,8-TeBC) were purchased from Sigma-Aldrich (St. Louis, MO) and the Florida Center for Heterocyclic Compounds of the University of Florida (Gainesville, FL), respectively. Surrogates 4′-fluoro-2,3′,4,6-tetrabromodiphenyl ether (F-BDE69) and injection standard decabromobiphenyl (BB209) were purchased from AccuStandard (New Haven, CT). All solvents were high-performance liquid chromatography (HPLC)- or Optima-grade and purchased from Fisher Scientific (Pittsburgh, PA). Silica gel (100−200 mesh, 75−150 μm, grade 644), alumina (neutral, Brockmann I, 50−200 μm by Acros Organics), and granular anhydrous sodium sulfate (Na2SO4) were also from Fisher Scientific. The sorbents silica gel and alumina as well as Na2SO4 were activated at 500 °C for 8 h, stored at 160 °C, and cooled to room temperature in desiccators before use. Qualitative Analysis. Extensive sample preparation and cleanup were conducted to separate target analytes (PHCs) from potentially interfering compounds (mainly PBDEs) and produce high-quality mass spectra of PHCs. In brief, 8 g of freeze-dried, composite sediment from Lake Michigan was extracted with an accelerated solvent extraction system (Dionex ASE350, Thermo Fisher Scientific, Inc.), using a procedure similar to that described in the U.S. EPA Method 3545A. The sediment was extracted with a hexane and acetone mixture (1:1, v/v) at 100 °C for 3 cycles, with heat time of 5 min and static 12808
dx.doi.org/10.1021/es503936u | Environ. Sci. Technol. 2014, 48, 12807−12815
Environmental Science & Technology
Article
Table 1. Elemental Compositions of Halogenated Carbazoles Found in Lake Michigan Sediments and Their Characteristic Ions during the EI and ECNI Mass Spectral Analyses accurate mass (Da) name
formula
UNC-1 UNC-2 UNC-3 3,6-DiBC UNC-4 UNC-5 UNC-6 UNC-7 UNC-8 UNC-9 1,3,6,8-TeBC UNC-10 UNC-11 UNC-12 UNC-13 UNC-14 UNC-15
C12H6NClBr2 C12H6NBr3 C12H7NBr2 C12H7NBr2 C12H6NBr3 C12H5NClBr3 C12H6NBr3 C12H5NClBr3 C12H6NBr3 C12H5NBr4 C12H5NBr4 C12H6NBr3 C12H4NClBr4 C12H4NBr5 C12H4NClBr3I C12H4NBr4I C12H4NBr3I2
measured
434.7661 434.7661 478.7155 478.7155 512.6766 556.6261 560.6603 604.6122
calculateda 356.855575 400.805068 322.894545 322.894545 400.805068 434.766098 400.805068 434.766098 400.805068 478.715591 478.715591 400.805068 512.676621 556.626114 560.662625 604.612118 652.598122
EI ions differenceb
−0.000002 −0.000002 0.000091 0.000091 0.000021 0.000014 0.002325 −0.000082
CI ions
molecular
main fragment
Qc
q1d
q2d
spectrum figure
358.8 404.8 324.8 324.8 404.8 438.7 404.8 438.7 404.8 482.7 482.7 404.8 516.6 560.6 562.6 608.6 654.5
279.9 323.8 245.9 245.9 323.8 357.8 323.8 357.8 323.8 401.7 403.6 323.8 438.6 481.6 438.7 482.7 529.8
79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81 79/81
358.9 402.8 324.9 324.9 402.8 438.8 402.8 438.8 402.8 482.7 482.7 402.8 516.7 560.6 562.7 608.7 654.5
280.9 324.9
S17 S7 S8 S3 and S5 S9 S18 S10 2 S11 S12 S4 and S6 S13 S19 S14 3 S20 S21
324.9 358.8 324.9 358.8 324.9 402.7 402.7 324.9 436.7 480.7 438.7 482.8 527.7
a
Calculations are based on the following exact masses: C, 12.000000; H, 1.007825; Br, 78.918348; Cl, 34.968855; F, 18.998405; N, 14.003074; and I, 126.904352.20 bDifference between calculated and measured masses. cQuantifier ion. dQualifier ion.
Figure 1. Partial GC−ECNI−LRMS chromatograms (30 m DB-5MS column, full scan, m/z 35−800) of deep sediment (26−30 cm) extract from sampling site M041 (A, F-2; B, F-3) and (C) standard mixture of halogenated flame retardants. In panel A, UNC-15 is zoomed in.
by use of the PFK reference peaks. The corrected data file was then opened, and the background was subtracted from the region of interest to generate the final mass spectrum. The
portion of the total ion current (TIC) where no peaks eluted. A secondary reference correction (Masslynx 4.1) was applied to the acquired data file to autocorrect each mass in the spectrum 12809
dx.doi.org/10.1021/es503936u | Environ. Sci. Technol. 2014, 48, 12807−12815
Environmental Science & Technology
Article
Figure 2. (Top) ECNI (scan range m/z 35−800) and (bottom) EI (scan range m/z 50−800) mass spectra (GC−LRMS) of UNC-7 in Lake Michigan sediment samples.
Quantifications of 3,6-DiBC and 1,3,6,8-TeBC were based on calibration curves developed from their corresponding reference standards. Calibration curves were generated from the bromine response ( 79 Br − or 81 Br − ) ratios versus concentration ratios of 3,6-DiBC and 1,3,6,8-TeBC to injection standard BB209. Because of the lack of commercially available reference standards for the 15 unknown compounds (UNCs) (Figure 1), semi-quantification was achieved via SIM for 79Br− and 81Br−, and based on the calibration curves for 3,6-DiBC or 1,3,6,8-TeBC. Specifically, UNC-1, UNC-3, UNC-5, and UNC7 were semi-quantified using the calibration curve of 3,6-DiBC because of the nearness in GC retention times. The calibration curve of 1,3,6,8-TeBC was used to semi-quantify other UNCs, including all tribromocarbazoles, one of which (UNC-8) matches with the impurity in the chemical standard of 1,3,6,8-TeBC. Quality Control. Two laboratory procedural blanks (Na2SO4) were analyzed along with sediment samples from each of the two cores (M018 and M041). Blanks were analyzed using the same laboratory procedures and instrument as used for sediments in quantitative analysis. Concentrations of PHCs in the procedural blanks ranged from no detection (nd) to 0.1 ng/g of dry weight (dw). Surrogate F-BDE69 was added to each sample before extraction, and its average recovery was 121 ± 15% for M018 samples and 80 ± 11% for M041 samples. Two samples (one for each core) were analyzed in duplicate, and the average relative percentage differences (RPDs) of the duplicate analyses for individual PHCs ranged from 0.4 to 26%. In addition, duplicate blanks (Na2SO4) were spiked with 1,3,6,8-TeBC (8 ng), and its average recovery was 95.4 ± 8%. Instrument detection limits (on the basis of 3 times the signal-
Masslynx 4.1 elemental composition program was used to determine the best fitting formula for each isotope signal in the mass spectrum of interest. All injections were made in the splitless mode at a temperature of 280 °C. The oven temperature program was the same as that for GC−LRMS analysis. Quantitative Analysis. Quantitative or semi-quantitative analyses of PHCs were conducted using a procedure similar to that used for qualitative analyses. Approximately 5 g of freezedried sediments were spiked with surrogates F-BDE69 (4 ng), stabilized overnight, and then extracted with accelerated solvent extraction (ASE). The cleanup column was packed with 8 g of alumina and 4 g of silica gel. Concentrated extract was loaded on the column and eluted with 100 mL of hexane (F-1), 100 mL of 4:1 hexane/DCM mixture (F-2), and 100 mL of DCM mixture (F-3). The latter two fractions contained PHCs. They were concentrated to 2 mL, and aliquots were placed in 200 μL glass inserts of GC vials. Injection standard BB209 (0.525 ng) was added to the insets prior to instrumental analysis. Separation and quantification of PHCs were performed on the Agilent 6890/5973 GC−ECNI−MS system with the use of the 15 m Restek Rtx1614 column described above, and MS was operated in the selective ion monitoring (SIM) mode. Other instrumental conditions were the same as those used in qualitative analyses. Table 1 lists the ions used for quantification (79Br− and 81Br−) for each PHC as well as their respective qualifier ions for confirmation during ENCI− LRMS analyses. The molecular ion and an additional fragment ion for each analyte under EI full scan are also included in Table 1. 12810
dx.doi.org/10.1021/es503936u | Environ. Sci. Technol. 2014, 48, 12807−12815
Environmental Science & Technology
Article
Figure 3. (Top) ECNI (scan range m/z 35−800) and (bottom) EI (scan range m/z 50−800) mass spectra (GC−LRMS) of UNC-13 in the extract of Lake Michigan sediment samples.
the identities of UNC-1−UNC-15 as a suite of polyhalogenated carbazoles, including eight polybrominated carbazoles and seven mix-halogenated carbazoles (i.e., containing more than one type of halogen atom) (Table 1). The identification and characterization of these two groups are separately discussed below. Polybrominated Carbazoles. 3,6-DiBC and 1,3,6,8-TeBC were among the initially targeted analytes of this work. The EI and ECNI full-scan mass spectra of their standards are given in Figures S3 and S4 of the Supporting Information, respectively. Figures S5 and S6 of the Supporting Information display the EI and ECNI full-scan spectra of these two compounds in a sediment extract, respectively, which agree well with those of the standards. In the EI−MS spectrum of 1,3,6,8-TeBC, the molecular ion cluster is centered at m/z 482.7, representing the [M+4]+ ion, and no ions were observed above this cluster to the maximum measured m/z value of 800 by the GC−LRMS analysis and 1000 by the GC−HRMS analysis. The major fragment ions included [M−Br]+, [M−2Br]+, [M−3Br]+, and [M−4Br]+ (see Figures S4 and S6 of the Supporting Information). The GC−HRMS analysis of the sediment extract further confirmed the elemental composition of C12H5NBr4 for the peak corresponding to 1,3,6,8-TeBC in standard solution, with the exact mass of molecular ion [M]+ being 478.7155, which matches well with the calculated mass of 478.715591 Da. In addition to 3,6-DiBC and 1,3,6,8-TeBC, eight other brominated carbazoles substituted with different numbers of bromine atoms were identified in the sediment extracts (Table 1, Figure 1, and Figure S1 of the Supporting Information). Their MS spectra are presented in Figures S7−S14 of the Supporting Information. These brominated UNCs included a
to-noise ratio) for 3,6-DiBC and 1,3,6,8-TeBC are 2.3 and 0.6 pg, respectively, when they were analyzed on GC−ECNI− LRMS.
■
RESULTS AND DISCUSSION The extensive cleanup and fractionation of sediment extracts facilitated the acquisition of high-quality GC chromatograms and mass spectra of UNCs. Figure 1 and Figure S1 of the Supporting Information show the chromatograms of optimally isolated fractions of a sediment extract obtained using GC− LRMS with ECNI and EI ion sources, respectively. This sample was obtained from the deepest segments of core M041, where interference from emerging halogenated flame retardants is negligible. A total of 15 halogenated UNCs as well as 3,6-DiBC and 1,3,6,8-TeBC were observed in the sediment extracts. For comparison, GC chromatograms of a mixture of halogenated flame retardant standards generated under the same instrumental conditions are included in Figure 1 and Figure S1 of the Supporting Information. Both the 15 m and 30 m GC columns resulted in the same elution order of targeted analytes, which were separated from each other as well as from most other halogenated compounds in fractionated sample extracts. The mass spectra of individual compounds are shown in Figure 2, Figure 3, and Figures S2−S21 of the Supporting Information. The UNCs differed from the known halogenated analytes in mass spectra. Full-scan EI spectra revealed that the molecular ion of the UNCs appeared at odd m/z values. According to the nitrogen rule of MS, the odd number of nominal mass implies an odd number of nitrogen atoms in the molecule.20 In the absence of chemical reference standards, evidence from both LRMS and HRMS spectra strongly support 12811
dx.doi.org/10.1021/es503936u | Environ. Sci. Technol. 2014, 48, 12807−12815
Environmental Science & Technology
Article
dibrominated (UNC-3), five tribrominated (UNC-2, UNC-4, UNC-6, UNC-8, and UNC-10), a tetrabrominated (UNC-9), and a pentabrominated carbazole (UNC-12). The identification was mainly based on the accurate mass determination of molecular ions (Table 1) and the interpretation of both EI−MS and ECNI−MS spectra. The EI or ENCI mass spectra of these UNCs consistently revealed fragmentation patterns of consecutive loss of bromines, similar to those of 3,6-DiBC or 1,3,6,8TeBC (see Figures S3 and S4 of the Supporting Information). The EI ionization of all polybrominated carbazoles consistently produced a fragment ion cluster centered around m/z 162, which appears to represent the backbone of substituted carbazoles. The GC−HRMS analyses further supported the identity of these UNCs as polybrominated carbazoles. For example, the accurate mass determination suggested an elemental composition of C12H4NBr5 for UNC-12, which has the exact mass of 556.6261 Da for its molecular ion. This agrees well with its calculated mass 556.626114 Da (Table 1). On the basis of the GC retention times and the MS spectra, UNC-8 and UNC-12 appear to be a tribromocarbazole (see Figure S15 of the Supporting Information) and a pentabromocarbazole (see Figure S16 of the Supporting Information) present as impurities in the 1,3,6,8-TeBC chemical standard. The substitution positions of bromine atoms on the carbazole backbone remain unknown. In laboratory syntheses, direct bromination of carbazole appears to favor 3, 3,6, 1,3,6, and 1,3,6,8 substitution patterns for mono-, di-, tri-, and tetrahomologues, respectively.21 Mixed Halogenated Carbazoles. The EI and ECNI fullscan mass spectra of seven mixed halogenated carbazoles (UNC-1, UNC-5, UNC-7, UNC-11, UNC-13, UNC-14, and UNC-15) are presented in Figures 2 and 3 as well as Figures S17−S21 of the Supporting Information. The analyses of both low- and high-resolution mass spectra suggest that these peaks correspond to a suite of compounds structurally similar to brominated carbazoles but containing more complicated compositions of halogen substituents. Because UNC-7 exhibited a high response during the EI analysis, it is used as an example to illustrate the structural identification strategy. The ECNI mass spectrum of UNC-7 reveals high relative abundances of the [Br]− ions at m/z 79 and 81 (top of Figure 2). It also exhibits successive losses of two bromine atoms, resembling the mass spectrum of 1,3,6,8-TeBC. In EI scan (bottom of Figure 2), an ion cluster centered at m/z 438.7 dominates the abundances of fragment ions and appears to be the molecular ion cluster because essentially no ions were observed above the m/z 438.7 cluster to the maximum m/z 800. This molecular ion cluster exhibits an isotopic signature corresponding to the presence of one chlorine and three bromine atoms.20 On the basis of the calculated isotope pattern of ClBr3, [M+4]+ should be the base peak and the calculated relative abundance of [M]+, [M+2]+, [M+6]+, and [M+8]+ to [M+4]+ would be 26.2, 85.3, 48.7, and 7.88%, respectively. For UNC-7, the measured ratios were at 25.5, 84.9, 48.6, and 6.94%, respectively, agreeing well with the calculated values. The GC−HRMS analysis revealed the elemental composition of UNC-7 to be C12H5NClBr3 because the accurate mass [M]+ (12C121H514N35Cl79Br3) was 434.7661 Da and is essentially the same as the calculated mass of 434.766097 (Table 1). Major EI fragment ions include [M−Br]+, [M−2Br]+, and [M−3Br]+, resembling the fragmentation pattern of 1,3,6,8-TeBC. Cleavage of Cl was also observed in the mass spectrum, producing minor fragment ions, such as [M−Cl]+, [M−Cl−
Br]+, [M−Cl−2Br]+, and [M−Cl−3Br]+ (Figure 2). Cleavage of all halogen substituents produces a fragment ion cluster centered at m/z 162.0 (i.e., [M−Cl−3Br]+), which corresponds to a carbazole ion fragment. Using the same strategy, we identified three additional halogenated carbazole congeners that contain both bromine and chlorine atoms, including UNC-1 (C 12 H 6 NClBr2 ), UNC-5 (C 12 H 5 NClBr 3 ), and UNC-11 (C12H4NClBr4). In addition to bromine and chlorine, iodine was also present in some halogenated carbazoles found in this work. The ECNI spectrum of UNC-13 as well as those of UNC-14 and UNC-15 exhibits a major fragment ion cluster centered at m/z 126.9 (Figure 3), which appears to represent the ion of [I]−.22 HRMS analyses suggest molecular compositions of C12H4NClBr3I for UNC-13 (centered at m/z 562.6) and C12H4NBr4I for UNC-14 (centered at m/z 608.6). The EI−LRMS ionization of UNC-13 results in a suite of dehalogenated fragment ions, including [M−Br]+, [M−I]+, [M−Cl−I]+, [M−Br−I]+, [M−Cl−Br−I]+, [M−2Br−I]+, [M−Cl−2Br−I]+, and [M−Cl−3Br−I]+ (Figure 3). The low concentration of UNC-15 in sediment made it difficult to determine the actual mass with HRMS and resulted in a less clear EI−LRMS spectrum compared to those of other UNCs (see Figure S21 of the Supporting Information). Nonetheless, its ECNI−LRMS and high m/z portion of its EI−LRMS spectra (see Figure S21 of the Supporting Information) indicate an ionization fragment pattern similar to those of other halogenated carbazoles. Therefore, UNC-15 is tentatively identified as a mixed halogenated carbazole with the molecular formula of C12H4NBr3I2. A close examination of the EI mass spectra revealed the possible presence of a doubly charged ion of [M]2+ with m/z value of M/2, in some PHCs. For example, in the EI spectra of UNC-7 (Figure 2) and UNC-13 (Figure 3), the ion clusters centered at m/z 219 and 282 may represent their respective doubly charged molecular ion [M]2+. Doubly charged ions were also found in other PHCs, such as tribromocarbazole UNC-2 (see Figure S7 of the Supporting Information), UNC-11 (see Figure S19 of the Supporting Information), pentabromocarbazole UNC-12 (see Figure S14 of the Supporting Information), and UNC-14 (see Figure S20 of the Supporting Information) with m/z values of 202.3, 258.3, 280.1, and 303.2, respectively. For dibromocarbazoles (see Figures S3, S5, and S8 of the Supporting Information) and tetrabromocarbazole (see Figures S4, S6, and S12 of the Supporting Information), the ion clusters of m/z 163−165 and 240−242 may represent doubly charged ions and/or dehalogenated fragment ions. Some PHCs could also produce doubly charged ions after losing two halogens. For example, UNC-12 and UNC-14 both exhibit an EI fragment ion cluster centered at m/z 200, which might represent [M−2Br]2+ and [M−I−Br]2+, respectively. Carbazole itself also exhibits a doubly charged ion with m/z 84 (see Figure S2 of the Supporting Information). Doubly charged ions are typical for polycyclic aromatic hydrocarbons (PAHs)20 and dibenzofuran.23 Characteristic doubly charged ions have been observed in mass spectra of polybrominated diphenoxybenzenes.24 PBDE congeners can also produce doubly charged ions of [M−2Br]2+ upon EI ionization, particularly for those with no ortho bromine or having 2,3- substitution on one ring and no ortho bromines on the other.25 The formation of a dibenzofuran-like fragment ion during EI ionization of PBDEs was proposed.26 In the present study, the doubly charged ion [M]2+ observed in the EI spectra of halogenated carbazoles may reflect their structural similarity to halogenated dibenzofurans. 12812
dx.doi.org/10.1021/es503936u | Environ. Sci. Technol. 2014, 48, 12807−12815
Environmental Science & Technology
Article
Figure 4. Depth profiles of PHCs and BDE209 in Lake Michigan sediment cores M018 (red) and M041 (blue).
Concentrations and Depth Profile. Concentrations of 3,6-DiBC and 1,3,6,8-TeBC ranged from 1 to 41 ng/g of dw and from 7 to 65 ng/g of dw, respectively. The latter is similar to the concentration range (i.e., from nd to 54 ng/g) reported by Zhu and Hites in their sediment samples from Lake Michigan.16 On the basis of semi-quantified results, in both cores, the most abundant PHCs were UNC-7 and 1,3,6,8TeBC, followed by UNC-12 and UNC-14. UNC-4, UNC-6, UNC-8, and UNC-15 had the lowest levels (
