Advances in the Analysis of Dioxin and Environmental Toxicants

Transcription

1 Advances in the Analysis of Dioxin and Environmental Toxicants

2 Co-Authors Eric Reiner, Terry Kolic, Karen MacPherson, Sheng Yang, Alina Muscalu, Adrienne Boden, Karl Jobst, Patrick Crozier, Vin Khurana, Teresa Gobran, Liad Haimovici,, Li Shen Christy Hartley, Rob Brunato, Vince Taguchi, Jeff Scott, Paul Helm, Tony Chen, Vadrana Pantelic, Gerry Ladwig Frank Dorman, Jack Cochran, Peter Gorst-Allman Joe Binkley, David E Alonso Ian Brindle, Chris Marvin 2 2

3 What is Dioxin? Polychlorinated dibenzo-p-dioxins (PCDD) and polychlorinated furans (PCDFs) A group of 210 compounds with chlorines 0 to 8 chlorines 2,3,7,8-tetrachlorodibenzo-p-dioxin 2,3,7,8-tetrachlorodibenzofuran 3

4 What is Dioxin? Most toxic group of chemicals with LD 50 as low as 1 ug/kg body weight Persistent, bioacummulative and toxic Health effects include: immune impairment, reproductive disorders, weight loss, development impairment, cancer Toxicity through the AhR binding 4 LD 50 : Lethal dose (50% test population)

5 5

6 Dioxin Toxicity NOEL = 3g/kg LD 50 = 1ug/kg TCDD: 2,3,7,8-tetrachlorodibenzo-p-dioxin NOEL: No-observable effect-level 6 LD 50 : Lethal dose (50% test population) 6

7 7

8 Dioxin-Like Toxicity Most toxic are planar with 3Å x 10Å dimensions 8

9 9

10 Toxic Equivalent Quantity (TEQ) Results reported in TEQ normalized to 2,3,7,8-TCDD TEQ = Σ[PCDD i TEF i ] + Σ[PCDF j TEF j ] + Σ[PCB k TEF k ] + Σ[DLC l TEF l ] Where: i = 1 7, j = 1 10, k = 1 12, l = 1? 10

11 Dioxin/PCB TEFs Eadon Ontario Germany California USEPA Nordic NATO-I WHO WHO WHO PCDDs 2,3,7,8-TCDD ,2,3,7,8-PeCDD ,2,3,4,7,8-HxCDD ,2,3,6,7,8-HxCDD ,2,3,7,8,9-HxCDD ,2,3,4,6,7,8-HpCDD ,2,3,4,6,7,8,9-OCDD PCDFs 2,3,7,8-TCDF ,2,3,7,8-PeCDF ,3,4,7,8-PeCDF ,2,3,4,7,8-HxCDF ,2,3,6,7,8-HxCDF ,2,3,7,8,9-HxCDF ,3,4,6,7,8-HxCDF ,2,3,4,6,7,8-HpCDF ,2,3,4,7,8,9-HpCDF ,2,3,4,6,7,8,9-OCDF dl-pcbs PCB-077 (3,3',4,4'-TCB) PCB-081 (3,4,4',5-TCB) PCB-105 (2,3,3',4,4'-PeCB) PCB-114 (2,3,4,4',5-PeCB) PCB-118 (2,3',4,4',5-PeCB) PCB-123 (2',3,4,4',5-PeCB) PCB-126 (3,3',4,4',5-PeCB) PCB-156 (2,3,3',4,4',5-HxCB) PCB-157 (2,3,3',4,4',5'-HxCB) PCB-167 (2,3',4,4',5,5'-HxCB) PCB-169 (3,3',4,4',5,5'-HxCB) PCB-189 (2,3,3',4,4',5,5'-HpCB)

12 Agent Orange 3 > miscarriages in sprayed areas 12

13 Seveso, Italy 1976 Chlorophenol plant explosion Normal sex ratio 106 males to 100 females Seveso parents produced only 26 boys to 48 girls Mocarelli et al, Lancet,

14 14 Yuschenko Poisoning 2002

15 15 Yuschenko Poisoning 2009

16 Dioxin Sources Open burning/wood burning Medical and waste incineration Chemical fires Auto exhaust Chemical production Tanneries Metal reclamation Industrial emissions 16 16

17 17 Pulp and paper plant lagoon

18 18 Open Barrel Burning

19 19 Hagarsville Tire Fire 1990

20 20 Hagarsville Tire Fire 1990

21 Plastimet Recycling Plant Fire

22 POPs Food Contamination BB-153 FeedMaster/FireMaster Belgian dairy Dioxin/PCBs1999 Irish Chicken Dioxin 2006 German chicken Dioxin 2009 BB = Brominated Biphenyl 22 22

23 Method Attributes Sensitivity Selectivity (Precision and Accuracy) Speed $ (Cost) Dioxin analysis requires selective/sensitive detection ( 100 fg) which requires concentration factors of 10 5 to

24 Dioxin Guidelines Food: 1 6 pg/g lipid weight (lw) (EU) Feed: ng/kg (EU) Air: 5 pg/m 3 (ON) Drinking Water 15 pg/l (ON) Surface Water: pg/l (ON) Effluent: 60 pg/l 5 pg/g 2,3,7,8-TCDD (ON) Stack Emission: 80 pg/m 3 (CAN) Soils: 7 pg/g (ON) Sediments: 0.85 pg/g (CAN) Fish: 2.3 pg/g (ww) (ON) ON = Ontario, CAN = Canada, EU = European Union, 24 lw = lipid weight ww = wet weight 24

25 Dioxin Daily Intake Guidelines Average Daily intake: 1 to 3 pg/kg/day Guidelines: U.S. EPA: 0.7 pg/kg/day EU: 1 4 pg/kg/day Canada: 10 pg/kg/day 25 25

26 POPs in Environmental Samples: the Analytical Challenge Ensure all labware, reagents and analytical instruments are free of contamination and interferences before beginning analysis Detect analytes at sub picogram level to meet sub picogram detection limits for dioxin, e.g., every piece of labware should be prechecked (<500 fg) or labware segregated for high and low samples Determination of a representative sub-sample including gravimetric or volumetric determination for analysis. May require adjusting sample size or replicate analysis Addition of all internal and surrogate standards to the sample such that they will behave as the natural analytes in the sample during the analysis Quantitative extraction of analytes from matrix Extract may contain a gram or more of coextractable organic material including: Dioxins, Furans, PCBs PCNs, PCDEs, polycyclic aromatic hydrocarbons (PAHs), organochlorine pesticides (OCs), brominated flame retardants (BFRs), and many other organic compounds including lipids, humic material and sulfur 26 26

27 POPs in Environmental Samples: the Analytical Challenge * Cleaning of extracts to remove interfering coextractables to the degree where DQOs and QC limits can be met for dioxin analysis * Not all coextractable compounds need to be removed, but they should not affect the separation or detection systems with respect to the analytes of interest * Separation of target analytes from non-target or non-toxic isomers, congeners or interfering compounds (GC-HRMS) * There are many congeners per analyte group * dioxins/furans: 210; PCBs: 209, PCNs 75 * Separate and accurately quantify all toxic congeners * dioxins/furans: 17, PCBs: 12, PCNs: 8 10 * Ensure method and instrument selectivity and sensitivity meet DQOs and QC limits * Accredited laboratory (to ISO 17025), Quality System, trained/experienced analysts, proper instrument and analytical procedures, validated and documented methods and SOPs, control charting, non conformance and root cause determination 27 27

28 POPs in Environmental Samples: the Analytical Challenge * Ensure quantitative accuracy * Calibration, blanks, spiked samples, CRMs, ILS, Performance Evaluation, surrogates and internal standards, standard validation. * Other Considerations: * Toxicity of compounds can range up to 6 orders of magnitude * T 4 CDD toxicity can range from: NOEL = 3 g/kg to LD 50 = 1 ug/kg must identify correct isomers using proper GC columns and conditions. * Range of concentrations fg/g (10-15 g/g ) to % levels potential lab contamination and instrument issues from carryover. * Range of sample types and complexities biota, air, water, soil, hazardous waste which have different matrix dependent and method requirement issues. Use method fit for purpose that has been validated for specific matrix being analyzed. * Are the patterns of congeners representative of samples being analyzed, e.g., does an ash sample have a combustion source pattern or does a biota sample have only the toxic congeners present? 28 28

29 Analytical Approach Selection of appropriate method Fit for Purpose Quantitative extraction of analytes from matrix Cleanup of sample extract to remove interfering matrix and coextractable compounds Chromatographic separation selective detection 29

30 Analytical Approach Gold standard regulatory compliance Isotope dilution with 13 C labeled internal standards Capillary gas chromatography separation High resolution mass spectrometry detection at a resolution > 10,000 (10% valley definition) using selected ion monitoring Many more compounds in samples and grouped analyses desirable Alternate methods like multidimensional chromatography, tandem mass spectrometry and time-of-flight mass spectrometry can enhance selectivity. These techniques may not have the required sensitivity. 30

31 Horwitz Uncertainty Curve 31 from Eppe, Chemosphere , (2008)

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33 Extraction Classical Method Aqueous: filter particles, Soxhlet extract filter paper, liquid/liquid extract aqueous phase and combine extracts Solid Soxhlet extraction Green Methods Solid phase extraction with particles together Pressurized liquid extraction PLE 33

34 Solid Phase Extraction 34

35 35 Solid Phase Extraction

36 Phase Extraction

37 Solid Phase Extraction 37

38 38 Pressurized Liquid Extraction PLE

39 39 Pressurized Liquid Extraction PLE

40 40 Pressurized Liquid Extraction PLE

41 41 Pressurized Liquid Extraction PLE

42 Extract Cleanup and Fractionation Multi-analyte groups are extracted together (dioxins/furans, PCNs, PCBs, PBDEs, HFRs) but cannot be separated in a single GC column analysis Coeluting / interfering compounds can be separated physically (into fractions) or by using multiple GC column phases (e.g., PCB 77 and 110) Sample extracts are separated using silica, carbon and alumina Dioxins/furans, PCNs and non-ortho PCBs in reverse carbon fraction Ortho substituted PCBs, PBDEs and other HFRs in forward carbon fraction Combining these analyses can save significant time and costs 42 42

43 43 Classical Open Column Sample Preparation

44 Classical Open Column Sample Preparation 44

45 FMS PowerPrep 45 FMS = fluid management systems 45

46 FMS PowerPrep 46

47 FMS WMF replicates PCDD/F 140 DLPCB C-244'-tri BDE 28 13C-22'44'-tetra BDE 47 13C-2378-TCDF 13C PeCDF 13C PeCDF 13C HxCDF 13C HxCDF 13C HxCDF 13C HxCDF 13C HpCDF 13C HpCDF BDEs 13C-22'44'5-penta BDE 99 13C-22'44'56'-hexa BDE C-22'44'55'-hexa BDE 153 CRM = certified reference material 13C-22'344'5'6-hepta BDE C-2378-TCDD 13C PeCDD 13C HxCDD 13C HxCDD 13C HpCDD 13C-OCDD 13C-deca BDE C-23'4'5-tetra pcb (70) 13C-344'5-tetra PCB (81) 13C-33'44'tetra PCB (77) 13C-2'344'5-penta PCB (123) C-233'44'-penta PCB (105) 13C-23'44'5-penta PCB (118) 13C-2344'5-penta PCB (114) 13C-33'44'5-penta PCB (126) PCNs 13C-23'44'55'-hexa PCB (167) 13C-233'44'5hexa PCB (156) 13C-233'44'5'-hexa PCB (157) 13C-33'44'55'-hexa PCB (169) 13C-233'44'55'-hepta PCB (189) 13C-1,3,5,7CN-PCN(42) 13C-1,2,3,4CN-PCN(27) 13C-1,2,3,5,7CN-PCN(52) 13C-1,2,3,4,5,7CN-PCN(64) 13C-octaCN-PCN(75) 47

48 PLE PowerPrep CRMs RM-EDF 2525 RM-EDF 2526 Compound Acceptable range Result Acceptable range Result pg/g pg/g pg/g pg/g PCDD/F 2378-TCDF 24.3 ± ± PeCDF 4.58 ± ± PeCDF 14.5 ± ± HxCDF 5.95 ± ± HxCDF 1.73 ± ± HxCDF 1.04 ± ± HxCDF 0.10 ± ± HpCDF 0.59 ± ± HpCDF 0.16 ± ± OCDF 0.38 ± ± TCDD 17 ± ± PeCDD 3.71 ± ± HxCDD 0.33 ± ± HxCDD 2.03 ± ± HxCDD 0.30 ± ± HpCDD 0.48 ± ± OCDD 1.71 ± ±

49 PLE PowerPrep CRMs RM-EDF 2525 RM-EDF 2526 Compound Acceptable range Result Acceptable range Result pg/g pg/g pg/g pg/g DLPCBs 344'5-tetra PCB (81) 161 ± ± '44'-tetra PCB (77) 1850 ± ± '344'5-penta PCB (123) 3280 ± ± '44'5-penta PCB (118) ± ± '5-penta PCB (114) 3410 ± ± '44'-penta PCB (105) ± ± '44'5-penta PCB (126) 628 ± ± '44'55'-hexa PCB (167) 7060 ± ± '44'5-hexa PCB (156) ± ± '44'5'-hexa PCB (157) 3380 ± ± '44'55'-hexa PCB (169) 52.1 ± ± '44'55'-hepta PCB (189) 1440 ± ±

50 PLE PowerPrep CRMs RM-EDF 2525 Compound Acceptable range Result pg/g pg/g BDEs 244'-tri BDE ± '45'-tetra BDE ± '44'-tetra BDE ± '44'-tetra BDE ± '44'6-penta BDE ± '44'5-penta BDE ± '44'56'-hexa BDE ± '44'55'-hexa BDE ± '344'5'6-hepta BDE ± Deca BDE ±

51 PLE PowerPrep CRMs CRM WMF-01 Wellington Labs Fish Quasimeme Biota Round 50 Target MOE Target MOE BDE ± ± BDE ± ± BDE ± ± BDE ± ± BDE ± ± BDE ± ± BDE BDE N=4 51

52 Cost Comparison Cost ($CDN) Method Solvent Usage (ml) Time (days)* Wages Solvent Column Packing Total Extraction Acid digestion ASE Sample Cleanup Column Chromatography FMS Combined Extraction and Cleanup Acid digestion + Column Chromatography ASE + FMS

53 Cape Sample Prep System 53 53

54 54 Cape Technologies Spiked Surrogates n=10

55 Dioxin Data Comparison WMF- 01-A WMF- 01-B Certified Range EDF A EDF B Certified Range Dioxins 2378-TCDF ± ± PeCDF 1.2 < ± ± PeCDF ± ± HxCDF ± ± HxCDF 1.4 < ± ± HxCDF < ± < ± HxCDF <0.59 < ± 0.4 <0.41 < ± HpCDF <0.50 < ± 1.9 <0.37 < ± HpCDF <0.54 < ± 0.5 <0.34 < ± 0.32 OCDF <0.43 < ± 2.1 <0.39 < ± TCDD ± ± PeCDD ± ± HxCDD <0.35 < ± 0.3 <0.35 < ± HxCDD < ± ± HxCDD <0.46 < ± 0.4 <0.48 < ± HpCDD <0.51 < ± 0.7 <0.37 < ± 0.36 OCDD <0.81 < ± 6.2 <1.4 < ±

56 dlpcb Data Comparison WMF-01- A WMF-01- B Certified Range EDF A EDF B Certified Range dlpcbs 344'5-tetra PCB (81) ± ± '44'-tetra PCB (77) ± ±834 2'344'5-penta PCB (123) ± ± '44'5-penta PCB (118) ± ± '5-penta PCB (114) ± ± '44'-penta PCB (105) ± ± '44'5-penta PCB (126) ± ± '44'55'-hexa PCB (167) ± ± '44'5-hexa PCB (156) ± ± '44'5'-hexa PCB (157) ± ± '44'55'-hexa PCB (169) ± ± '44'55'-hepta PCB (189) ± ±

57 Automated Sample Prep Summary The automated extraction/cleanup method: Is simple, non-labour intensive, time- and cost-saving Decreases the possibility of human health hazards from solvent exposure as well as solvent usage Minimizes background contamination Combines four analyte groups (PCDDs/PCDFs, DLPCBs, PCNs and BDEs) in a single extraction using only 2 or 3 columns for clean-up and fractionation Reduces the time of sample preparation from 15 to 3 working days and increases productivity Is robust, repeatable, accurate, has lower method detection limits and greater analytical precision than the separate classical methods 57

58 Gas Chromatographic Separation Gold standard for dioxins: 60 m, 0.25 mm id, 0.25 um ft, 5% phenyl-methyl column for analysis 30 m, 0.25 mm id, 0.25 um ft, 50% cyanopropyl, 50% phenylmethyl polysiloxane column for confirmation of coeluting congeners Analyte Potential Coeluting Compounds on 5% phenyl 2,3,7,8-T 4 CDF 1,2,4,9- T 4 CDF, 1,2,7,9- T 4 CDF, 2,3,4,6- T 4 CDF, 2,3,4,7- T 4 CDF, 2,3,4,8- T 4 CDF 1,2,3,7,8-P 5 CDF 1,2,3,4,8- P 5 CDF 2,3,4,7,8- P 5 CDF 1,2,4,8,9- P 5 CDF, 1,2,3,6,9- P 5 CDF, 1,2,6,7,9- P 5 CDF 1,2,3,4,7,8-H 6 CDF 1,2,3,4,6,7- H 6 CDF 2,3,7,8-T 4 CDD 1,2,3,7- T 4 CDD*, 1,2,3,8- T 4 CDD*, 1,2,3,9- T 4 CDD* (*nearest neighbors) 1,2,3,7,8- P 5 CDD PCB-169 (fragment ion) 1,2,3,7,8,9- H 6 CDD 1,2,3,4,6,7- H 6 CDD PCB-77 PCB-110 PCB-81 PCB-87 PCB-123 PCB-149 PCB-126 PCB-178, PCB-129 PCB-156 PCB-171 PCB-157 PCB

59 GC Column comparison PCDDs DB-5, Rtx-5MS HP5-MS, Equity-5 CP-Sil 8 CB/MS DB-5MS ZB-5MS Rtx- Dioxin2 ZB5UMS DB-XLB DB-225 SP ,3,7,8-TCDD ,2,3,7,8-PeCDD ,2,3,4,7,8-HxCDD ,2,3,6,7,8-HxCDD ,2,3,7,8,9-HxCDD ,2,3,4,6,7,8-HpCDD ,2,3,4,6,7,8,9-OCDD PCDFs 2,3,7,8-TCDF ,2,3,7,8-PeCDF ,3,4,7,8-PeCDF ,2,3,4,7,8-HxCDF ,2,3,6,7,8-HxCDF ,2,3,7,8,9-HxCDF ,3,4,6,7,8-HxCDF ,2,3,4,6,7,8-HpCDF ,2,3,4,7,8,9-HpCDF ,2,3,4,6,7,8,9-OCDF = Baseline separation, +- = partial cocelution, - - = complete coelution

60 Potential interferences in TCDD analysis 60 from Tondeur, Biomed. Environ. Mass Spectrom (1987)

61 Potentially interfering compounds to consider Compound Ion Resolution Interferent Needed Nonachlorobiphenyl ,236 No Pentachlorobiphenylene ,432 Yes Heptachlorobiphenyl ,480 Yes Hydroxytetrachlorodibenzofuran Infinity Yes removed in cleanup DDE ,044 No Polychloroxanthenes ,089 Yes Tetrachloromethoxybiphenyl ,870 No 61

62 Column Performance win/res mix-f dlp-cs4 back jan10_02std : Voltage SIR 25 Channels EI e7 1237/1238-tcdd \ 2378-tcdd % 1289-tcdd 1234-tcdd Time 62 5% phenyl-methyl column

63 TCDF Ion Channel for Flyash Extract c may12_res Flyash Extract : Voltage SIR 20 Channels EI e % may12_res : Voltage SIR 20 Channels EI e6 13C12-2,3,7,8-tcdf % Time 63

64 Comparison of GC Columns Column Length (m) Inner diameter (mm) Film thickness (µm) Theoretical plates/m 8,600 3,300 5,300 Total plates (for column) 172, , ,000 Calculated no. plates (T 4 CDD) 176, , ,000 Relative Efficiency Relative Anal. Time Need 175,000 plates to separate critical pair 1,2,3,7/1,2,3,8 and 2,3,7,8 TCDD 64 64

65 Fast GC Method Attributes Changes in parameters shorter & narrower columns thinner stationary phase films faster oven temperature programming rates higher pressures, faster carrier gas flow rates Faster analyses increased sample throughput Narrower peak widths place new demands on detection systems 65

66 What is Fast GC? GC ramp rates >50ºC/min. GC column diameters <0.18 mm Stationary phase <0.18 µm GC column head pressures >60 psi Phase ratio: β = r / df r = column diameter df = film thickness 66

67 Comparison of Different Column Dimensions Handbook of GC/MS H.J. Huebschmann 67

68 Fast GC Chromatogram 60m 0.25μm, 0.25mm 5% phenyl RT = % Valley 20m 0.1 μm, 0.1mm 5% phenyl RT = 9.00 RTA = 38% 25% Valley 68

69 69 How many data points are necessary to correctly characterize a chromatographic peak?

70 70 Seven to ten points per peak is optimum

71 Reduction in Analysis Time Using Microbore Columns 60 m (.25/.25) 40 m (.18/.18) 30 m (.25/.25) 20 m (.10/.10) 10 m (.10/.10) Dioxins [50] 28 (44%) 14 (72%) PCB Congeners [90] 18 (80%) PAH [40] 22 (55%) 14 (65%) OC Pesticides [55] 12 * (78%) 71

72 Parallel Columns Most compound classes cannot be uniquely separated on a single chromatographic phase e.g., PCBs, dioxins, PAHs Extracts must be separated or analyzed on 2 phases Parallel columns can be used to analyze multiple fractions simultaneously e.g., dioxins/furans/coplanar PCBs and ortho PCBs Extracts can also be analyzed separately Analysis and confirmation in the same run Both columns must be temperature compatible 72 72

73 Parallel Column Advantages Mono-ortho PCBs elute from 20m column well before PCDD/Fs elute from 40 m column DPEs, and interfering PCBs in mono-ortho DLPCB sample fraction Avoid potential interferences: 1) Furan formation in ion source 2) Reduces coelution of higher chlorinated PCBs with coplanars (i.e., PCB 110 with PCB 77) 73 73

74 GC Configuration HP model GC 2 injection ports and 2 autosamplers 2 GC Columns Dioxin/furan & coplanar PCBs injected on 40 m Rtx5, 0.18 mm x 0.20 µm Mono-ortho PCBs 20 m Rtx5, 0.1 mm x 0.1 µm 74 74

75 The Gas Chromatograph 75 75

76

77 Fraction 2: 40 m Tetra Octa PCDDs 2378-TCDD PCDD HxCDD / HxCDD HxCDD \ / HpCDD / OCDD 81 \ 77 / Fraction 2: 40 m Coplanar PCBs 123 \ 118 / 114 / 105 / \ 157 / 189 / Fraction 1: 20 m Mono Ortho PCBs 77 77

78 Dioxins and WHO PCBs in NIST Tetra to octa dioxins on a 40 m column T 4 CDDs P 5 CDD s H 6 CDD s H 7 CDDs OCDD % PCDFs and coplanar PCBs (77/81/126/169) not shown % % % Time H 7 CBs on 20 m column (PCB189) H 6 CBs on 20 m column (PCB156/157/167) P 5 CBs on 20 m column (PCB105/114/118/123) Time 78 78

79 Dioxin/Furan Results Comparison NIST 1944 MARINE SED. Single (pg/g) Dual (pg/g) Ref. Value (pg/g) 2378-TCDD ± PCDD ± HxCDD ± PCDF ± 4 OCDF ±

80 DLPCB Results Comparison NIST 1944 Single Dual Ref. Value MARINE SED. (pg/g) (pg/g) (pg/g) PCB ,000 26,000 24,500 ± 1100 PCB ,500 62,600 58,000 ± 4300 PCB ± 660 PCB N/A PCB N/A 80

81 Two-Dimensional GC (GC GC) Produces higher peak capacity (more chromatographic peaks per space). Increases peak capacity to = 1000 compounds Eliminates the need for second column confirmation. Can do multiple analyte groups in same run and may eliminate need for extract fractionation Fast analysis requires fast detector e.g., time-of-flight mass spectrometer (TOFMS), ECD Provides structured chromatograms for excellent selectivity Provides much more information Results in increased sensitivity 81 81

82 GC GC Schematic Diagram Injector TOFMS Modulator 1 D 2 D P M X X+Y Y P M = modulation time D = dimension 1 t R Retention Time (t R ) 2 t R X 2 t R Y 82 82

83 Second Dimension Modulation Signal X Y 1 t R X 1 t R X 1 t R Y Y 2 t R Second dimension retention time 2 t R X 2 t R Y 83 83

84 84 Inside GC Oven

85 2 nd Oven 85 Modulator

86 Peak Modulation Original peak is chopped into 3 peaks Sensitivity enhancement occurs through focusing Second dimension peaks are only 400 ms wide Need detector capable of defining peaks Hundreds of spectra/sec ms = millisecond 86 86

87 Comprehensive GC GC Form of 2DGC: Orthogonal Column Setup 1st Dimension (Column) Standard (10 60 m, 0.25 mm, 0.25 um) Non-polar (DB-1, Rtx-5) 2nd Dimension (Column) Very short (1 2 m) Narrow bore, thin film ( mm, um) Polar or shape selective (DB-1701, Rtx-PCB) 87 87

88 PCBs Challenging Separations DB5-10M/0.18/0.18_DB17-2M/0.1/

89 PCB Standard by GC GC-ECD Orthogonal Elution PCB STD (BP-MS) Hexa- Penta- 169 Hepta Tetra * * * Tri * 183* * * 178* * * * 95 Di * * 33 18* Nona- Octa- 206* 209 Deca- *to be confirmed by GCxGC/TOF-MS 89 89

90 Dioxin-Like PCBs vs. Other PCBs Rtx-PCB > m/z Rtx-1 > 90 90

91 Sediments by GC GC-ECD CBz PCNs, PCDEs 3 2 nd Dimension (s) 2 PCA bands Unknown compounds 1 PCBs/OCs Dioxins/Furans st Dimension (s) 91

92 GCxGC-TOFMS of sediment sample C for chlorinated dioxins and furans 1m x 0.15mm x 0.15µm Rxi-17Sil MS 40m x 0.18mm x 0.18µm Rtx- Dioxin2

93 GCxGC-TOFMS of sediment sample C for chlorinated dioxins and furans 1m x 0.15mm x 0.15µm Rxi-17Sil MS Dioxin/furan elution area m/z ions plotted: m x 0.18mm x 0.18µm Rtx- Dioxin2

94 GCxGC-TOFMS of sediment sample C for chlorinated dioxins and furans, zoom 1m x 0.15mm x 0.15µm Rxi-17Sil MS 2378 TCDD Only m/z 322 is plotted. Note isobaric interferences eluting below 2378 TCDD that would interfere in 1D GC. 40m x 0.18mm x 0.18µm Rtx- Dioxin2

95 GCxGC-TOFMS of sediment sample B for chlorinated dioxins and furans, zoom 1m x 0.15mm x 0.15µm Rxi-17Sil MS Cl 6 PCBs While looking for Cl 6 PCBs (m/z 360), we see some interesting latereluting peaks 40m x 0.18mm x 0.18µm Rtx- Dioxin2

96 GCxGC-TOFMS of sediment sample B for chlorinated dioxins and furans, zoom 1m x 0.15mm x 0.15µm Rxi-17Sil MS Dibromopyrenes! 40m x 0.18mm x 0.18µm Rtx- Dioxin2

97 Background Muir & Howard, ES&T, 2006, 40, US TSCA United States Toxic Substances Control Act 97

98 Stockholm Agreement Compounds Original 12 Added 2009 Under Discussion aldrin, chlordane, dieldrin, DDT, endrin, heptachlor, hexachlorobenzene (HCB), mirex, toxaphene, polychlorinated biphenyls (PCBs), polychlorinated dibenzodioxins (PCDD) and dibenzofurans (PCDF) chlordecone, α-hexachlorocyclohexane, β-hexachlorocyclohexane, hexabromobiphenyl, tetra- to heptabromodiphenylether, lindane, (δ-hexachlorocyclohexane), pentachlorobenzene, perfluoroctanesulfonic acid (PFOS), its salts and perfluoroctanesulfonyl fluoride short chain chlorinated paraffins (SCCPs), endosulfan, hexabromocyclododecane (HBCD) 98

99 Sediment CRM Lake Ontario CRM = certified reference material Analysis Time: 7.3 minutes 99

100 Sediment CRM Target Compounds Phenanthrene 2. Anthracene 3. Fluoranthene 4. Pyrene 5. Benzofluoranthenes 6. Benzo[e]pyrene 7. Benzo[a]pyrene 8. Perylene 100

101 Time-of-Flight Mass Spectrometry p,p -DDE Dieldrin Endrin PCB-81 PCB-87 PCB-77 PCB-110 PCB-151 PCB

102 PAHs, PCBs & Organochlorine Pesticides by GC-TOF Pyrene g-chlordane Endosulphan-1 a-chlordane PCB-99 PCB-101 PCB

103 GC- HR TOF of Dioxins CS3WT 4.0e5 3.0e e5 1.0e5 0.0e0 Time (s) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) XIC( ±0.005) R FWHM = 25,

104 GC- HR TOF MS Dioxin Sensitivity 3.5e3 3.0e3 TCDF TCDF TCDD 2.5e3 CS1 (500 fg/µl; 2 µl inj.) 2.0e3 1.5e3 CS0.5 (250 fg/µl; 2 µl inj.) 1.0e3 CSL (100 fg/µl; 2 µl inj.) 5.0e2 0.0e0 Time (s) XIC( ±0.005) CSL PEIS 70eV XIC( ±0.005) CSL PEIS 70eV XIC( ±0.005) CS-0.5 PEIS 70eV XIC( ±0.005) CS-0.5 PEIS 70eV XIC( ±0.005) CS-1 PEIS 70eV XIC( ±0.005) CS-1 PEIS 70eV

105 HR TOF MS Full Scan Data OCDD OCDF

106 GC- HRTOF MS Additional Compounds 1.0e5 8.0e4 6.0e4 4.0e4 2.0e4 0.0e0 Time (s) Name Similarity Formula Mass Accuracy (ppm) 1-Chloropyrene 913 C16H9Cl Phenyldibenzofuran 879 C18H12O 0.35 Benzo[2,1-b:3,4-b']bisbenzofuran 891 C18H10O Pyreno[4,5-b]furan (CAS) 918 C18H10O ,3-Dichlorobenzo[c]phenanthrene 821 C18H10Cl

107 GC- HR TOF MS Additional Compounds 2.7e6 2.4e6 2.1e6 1.8e6 1.5e6 1.2e6 9.0e5 6.0e5 3.0e5 0.0e0 Time (minutes) Name Formula RDBE Mass Accuracy (ppm) Benzo[ghi]fluoranthene C18H Triphenylene C18H Benz[a]anthracene, 7-methyl- C19H Benzo[k]fluoranthene C20H

108 108 Plastimet Fire 1997

109 Fourier Transform Mass Spectrometry R=100,000 ROMS (Regular old mass spectrometer) ,000 FWHM 29,000 FWHM 7,000 FWHM 1,500 FWHM Ultrahigh resolution (The above mass range is 0.3 Da) 2048K data points 512K data points 128K data points 32K data points 109

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114 Kendrick Plot 1 Kendrick plot - vegetation exposed to fallout from the 1997 Plastimet fire Kendrick mass = mass x (35 / ) Kendrick Mass Defect unassigned CH CHCl CHClO CHO Nominal Kendrick Mass 114

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118 Summary of Method Enhancements The analysis of dioxin and related compounds requires the most sensitive and selective instrumentation Most POPs sample preparation procedures can be automated or semi-automated and many can be combined to save money and time Fast GC, Parallel GC and GC GC can significantly reduce sample analysis times and costs while increasing analytical capacity. Kendrick Plots can be used to identify unknown compounds. GC GC can increase selectivity and sensitivity and can also be used for analytical triage. Extract fractionation may not be required. These enhancements can save time, costs and reduce the use of solvents and reagents. 118

119 4th Annual Multidimensional Chromatography Workshop Confirmed Speakers: Toronto, Canada January 8 & 9, 2013 Daniela Cavgnino Dani Jack Cochran Restek John Dimandja Collage Frank Dorman Penn State Taduesz Gorecki Waterloo Teruyo Ieda Gerstel Philip Marriott Monash University Brian McCarry McMaster Jef Focant U Liege John Seeley Oakland University Heather Bean Vermont Catherine Rawlinson Murdoch U No Registration Fee Contact Eric Reiner at : Eric.Reiner@Ontario.ca 119

120 Thank You! References: E.J. Reiner, R.E. Clement, A.B. Okey, C.H. Marvin, Advances in the Analysis of Polychlorinated Dibenzo-pdioxins, Dibenzofurans and Dioxin-like Polychlorinated Biphenyls in Environmental Samples Analytical and Bioanalytical Chemistry, 386, 791 (2006). E.J. Reiner, A.R. Boden, T. Chen, K.A. MacPherson, A.M. Muscalu, Advances in the analysis of persistent halogenated organic compounds, LC GC Europe, February, 23(2), 60 (2010) E.J. Reiner, Analysis of Dioxin and Related Compounds Mass Spectrometry Reviews, 29(4), 526 (2010). V.Y Taguchi, RJ Nieckarz, RE Clement. S Krolik, R Williams, Dioxin Analysis by Gas Chromatography-Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (GC-FTICRMS), J Am Soc Mass Spectrom 21, 1918 (2010) 120