Comparison of Atmospheric Pressure Ionization Gas Chromatography

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Comparison of Atmospheric Pressure Ionization Gas Chromatography-Triple Quadrupole Mass Spectrometry to Traditional High-Resolution Mass Spectrometry for the Identification and Quantification of Halogenated Dioxins and Furans Kari L. Organtini,† Liad Haimovici,‡ Karl J. Jobst,‡,§ Eric J. Reiner,‡,∥ Adam Ladak,⊥ Douglas Stevens,⊥ Jack W. Cochran,#,○ and Frank L. Dorman*,†,○ †

Biochemistry, Microbiology, and Molecular Biology Department, The Pennsylvania State University, 107 Althouse Laboratory, University Park, Pennsylvania 16802, United States ‡ Ontario Ministry of the Environment, 125 Resources Road, Toronto, Ontario, Canada, M9P 3 V6 § Department of Chemistry, McMaster University, 1280 Main Street West, Hamilton, Ontario, Canada, L8S 4M1 ∥ Department of Chemistry, University of Toronto, 80 St. George Street, Toronto, Ontario, Canada, M5S 3H6 ⊥ Waters Corporation, 34 Maple Street, Milford, Massachusetts 01757, United States # Restek Corporation, 110 Benner Circle, Bellefonte, Pennsylvania 16823, United States ○ Forensic Science Program, The Pennsylvania State University, 107 Whitmore Laboratory, University Park, Pennsylvania 16802, United States S Supporting Information *

ABSTRACT: The goal of this study was to qualify gas chromatography coupled to atmospheric pressure ionization tandem mass spectrometry (APGC-MS/MS) as a reliable and valid technique for analysis of halogenated dioxins and furans that could be used in place of more traditional gas chromatography coupled to highresolution mass spectrometry (GC-HRMS) analysis. A direct comparison of the two instrumental techniques was performed. APGC-MS/MS system sensitivity was demonstrated to be on the single femtogram level. The APGC-MS/MS analysis also demonstrated method detection limits (MDLs) in both sediment and fish that were 2−18 times lower than those determined for the GC-HRMS. Inlet conditions were established to prevent issues with sample carry-over, due largely to the enhanced sensitivity of this technique. Additionally, this work utilized direct injection for sample introduction through the split/splittless inlet. Finally, quantification of both sediment and fish certified reference materials were directly compared between the APGC-MS/MS and GC-HRMS. The APGC-MS/ MS performed similarly to, if not better than, the GC-HRMS instrument in the analysis of these samples. This data is intended to substantiate APGC-MS/MS as a comparable technique to GC-HRMS for the analysis of dioxins and furans.

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clean up to remove potential interferences, which prove to be time-consuming. Chromatographically, structure similarity poses a challenge when it comes to peak resolution of so many congeners. Advances have been made in GC column technology to overcome coelution of important congeners through the development of dioxin specific columns that resolve the 2,3,7,8- substituted congeners considered most toxic from the non 2,3,7,8- substituted congeners.5 The universally accepted detection technique for this analysis typically requires the use of a mass spectrometer that is both highly selective and sensitive. A minimum resolving power of

olychlorinated dibenzo-p-dioxins (PCDD) and dibenzofurans (PCDF) have been compounds of importance for decades, with the first publications describing analytical techniques for their identification appearing in the 1970s.1−3 PCDDs and PCDFs are environmental contaminants that are generated as industrial and combustion byproducts. Of the 210 potential isomers, only 17 are regulated by various organizations around the globe and the World Health Organization (WHO) has assigned these 17 toxicity ratios termed “Toxic Equivalence Factors” or TEFs relative to 2,3,7,8TCDD.4 Frequently referred to as “dioxin analysis”, it is considered a challenging analytical field for several reasons. Environmental samples typically are challenging when it comes to sample preparation methods. There are generally accepted methods for dioxin sample prep that involve extensive sample © XXXX American Chemical Society

Received: May 5, 2015 Accepted: July 3, 2015

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DOI: 10.1021/acs.analchem.5b01705 Anal. Chem. XXXX, XXX, XXX−XXX

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etry (MS/MS) has been applied to dioxin analysis using both triple quadrupole and quadrupole ion trap instruments and has proven to be valid a technique for the analysis.23,24 In 2014, the European Union created EU 589 as a validated method for the analysis of dioxins in food, which included GC-MS/MS as a confirmatory method.25 This is a big step in the acceptance of non-HRMS methodology for the analysis of dioxins but is strictly restricted to the application of food and feed products. The goal of this research is to directly compare the performance of APGC-MS/MS to GC-HRMS in an effort to validate the technology as one that is equal in performance for the analysis of dioxins in environmental samples.

10 000 is generally needed for mass spectral resolution of the dioxin compounds from other halogenated organic pollutants that are common interferences.6 The mass spectrometer also needs to be highly sensitive to be able to detect trace levels of dioxins in samples, although concentrations can range over large orders of magnitude in a single sample. Because of these requirements, high-resolution mass spectrometers, typically magnetic sector based, have long been considered the gold standard for dioxin analysis. Most validated and accepted regulatory methods require GC-HRMS as the analytical technique used for dioxin analysis, including U.S. EPA methods 1613 and 8290, EN 1948, MOE 3418, and JIS methods K0311 and K0312.7−12 While HRMS analysis has proven to have the sensitivity and selectivity required for dioxin analysis, it also has some disadvantages. The instruments are fairly expensive to upkeep and require a skilled user to operate, making dioxin analysis impractical for most laboratories. Additionally, when operating HRMS systems in selected ion recording (SIR) mode, the number of masses monitored is inversely related to sensitivity. This effectively limits the number of different compounds that can be monitored for at any given time. Increasing the list of target compounds analyzed in a single run to include other potentially toxic halogenated dioxins (polybrominated, mixed halogenated Br/Cl) is highly desirable but would significantly reduce the sensitivity of the instrument. A new approach to dioxin analysis is proposed through this research that does not utilize a high resolution-mass spectrometer. The new approach utilizes a triple quadrupole mass spectrometer coupled with an atmospheric pressure ionization GC source. Atmospheric pressure ionization (API) is an ionization technique that has been around since the 1970s.13,14 These early applications of the technique used 63 Ni foil as an electron emission source to induce charge transfer ionization. The current instrumentation utilizes a plasma discharge from a corona pin to induce ionization under atmospheric pressure. The largest benefit to using this ionization technique is that it is a soft ionization process creating more molecular ion than would be present from the more classically used electron ionization. Increased molecular ion allows for enhanced sensitivity when using multiple reaction monitoring (MRM) transitions for detection. Added benefits from the use of an atmospheric pressure source include less GC method restrictions in the case of flow rate since the column effluent is not exiting into a vacuum outlet. Coupling to a triple quadrupole also increases sensitivity by using MRMs to monitor for specific precursor and product ions of interest. Switching between MRM transitions is fast, making it possible to monitor for upward of hundreds of MRMs in a single run without sacrificing sensitivity.15 This is extremely useful for dioxin analysis with the increasing desire to expand methods to include additional compounds. Retention time overlap of homologue groups often restricts method expansion on a GCHRMS system due to the limited number of ions that can be monitored at a time without significant loss in sensitivity. Overall, APGC-MS/MS systems promise increased ease of operation, easier maintenance and operation, and increased ability to analyze for many additional compounds without a significant decrease in sensitivity compared to GC-HRMS systems. APGC has been used for varying applications in the more recent years, including pesticide analysis,16,17 food analysis,18 environmental pollutants,19 metabolic profiling,20,21 and pharmaceuticals analysis.22 In addition, tandem mass spectrom-



EXPERIMENTAL SECTION Standards and Chemicals. All standards used were obtained from Wellington Laboratories, Inc. (Guelph, Ontario, Canada). Polychlorinated dibenzo-p-dioxin and dibenzofuran (PCDD/F) identification was performed using a mixture of 17 regulated tetra- through octachloro dioxins and furans (EPA1613CVS). This standard is a five point calibration set used to create the calibration curves used for quantification. A low level tetrachloro dibenzo-p-dioxin mix containing six different TCDD isomers ranging from 2 to 100 fg/μL (TF-TCDDMXB) was used for determining the sensitivity of the APGCMS/MS instrument. A mix of 13-C labeled PCDD/Fs (EPA1613LCS) was spiked into samples prior to extraction to use for isotope dilution quantification. Toluene was obtained from Avantor Performance materials (formerly JT Baker, Center Valley, PA) and was ultra resianalyzed grade. Nonane was obtained from Acros Organics (New Jersey) and was 99% pure. Reference Samples. Reference sample extracts were provided by the Ontario Ministry of the Environment and Climate Change (MOECC). These were samples that had previously been extracted at the MOECC for analysis on a GCHRMS instrument.10 Samples used for the determination of method detection limit (MDL) included two matrixes, sediment and fish. A variety of reference sample extracts were tested for instrumental comparison including WMS-01 Reference Lake Sediment for Organic Contaminant Analysis (Wellington Laboratories Inc.), WMF-01 Reference Freeze-dried Fish Tissue for Organic Contaminant Analysis (Wellington Laboratories Inc.), EDF-2524 Clean Fish Reference Material (Cambridge Isotope Laboratories Inc., Tewksbury, MA), EDF2525 Contaminated Fish Reference Material (Cambridge Isotope Laboratories Inc.), and NIST 1944 New York/New Jersey Waterway Sediment. Sample extracts were reconstituted with 20 μL of injection standard containing 2000 pg of 13C-labeled 1,2,3,4-TCDD and 13C-labeled 1,2,3,7,8,9-HxCDD in order to determine the recoveries of the labeled internal standards. For this study, 7 PCDD and 10 PCDF compounds were quantified using isotope dilution. For GC-HRMS analysis, no further sample preparation was performed. For APGC-MS/MS analysis, sample extracts were diluted prior to injection due to instrumental sensitivity. All samples, excluding NIST 1944, were diluted 20fold in toluene. The NIST 1944 samples were diluted 10-fold in toluene. GC-HRMS Analysis. High-resolution mass spectrometry (HRMS) experiments were performed using a Micromass Autospec magnetic deflection instrument (Waters Corporation, Milford, MA) coupled to an HP 7890A gas chromatograph B

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analysis and then maintained at a flow rate of 215 L/h until the end of the analysis. Corona voltage was initially 20 μA for the first 8 min of the analysis and then maintained at 4.0 μA until the end of the analysis. Cone voltage was maintained at 30 V for all compounds. The APGC source was heated to 150 °C and the mass spectrometer transfer line was run at 360 °C. The mass spectrometer was operated using the multiple reaction monitoring (MRM) mode. Two MRM transitions were utilized for each compound monitored, one serving as a quantification ion, and the second as a qualifier ion. Each MRM accounted for the loss of the −COCl fragment (−13COCl in the case of the 13C labeled standards and −CO37Cl in the case of the 37 Cl labeled TCDD standard). Specific MRM information, including retention time windows, collision energies, and dwell times utilized can be found in Table S2 in the Supporting Information. Chlorinated dioxins and furans were separated into functions dependent on chlorination level and within each function an additional MRM was monitored for a common polychlorinated diphenyl ether interference (Table S2 in the Supporting Information). Reduction of Carry-Over. A series of experiments were performed to reduce issues initially observed with carry-over in the APGC-MS/MS system. The source of carry-over was isolated to the inlet of the GC. Different inlet liners and syringes were tested as a result. Inlet liners used during these experiments included 4.0 mm Single Taper IP deactivated with wool, 4.0 mm Single Taper IP deactivated with no wool, and 4.0 mm drilled hole Uniliner (Restek Corporation, Bellefonte, PA). Syringes used during these experiments included Hamilton 1700 series 10 μL nongas tight, 10 μL gastight, and 5 μL nongas tight (Hamilton Company, Reno, NV) and SGE 10 μL nongas tight and 10 μL gastight syringes (SGE Analytical Science, Victoria, Australia).

equipped with an Agilent 7693B autosampler (Agilent Technologies, Santa Clara, CA). The HRMS was operated in the electron ionization (EI) mode at a resolution of 10 000 (10% valley definition) across the entire mass range. Detection was achieved by selected ion monitoring (SIM) of the PCDD/ PCDF molecular ions. A list of target masses monitored in SIM mode can be found in Table S1 in the Supporting Information. Perfluorokerosene (PFK) was introduced via a heated septum inlet system to generate lock mass ions. Helium carrier gas was used, and the GC was operated in splitless mode at a constant flow rate of 0.8 mL/min. The injector was maintained at a temperature of 280 °C and utilized a 4.0 mm double gooseneck splitless liner (Restek, Bellefonte, PA). Samples were injected under these conditions at a volume of 1.0 μL. A 40 m × 0.18 mm, 0.18 μm DB-5 column (Agilent Technologies, Santa Clara, CA) column was used for the separation. The GC oven temperature program was as follows: initial oven temperature 140 °C hold 1 min, 40 °C/min to 200 °C no hold, 3 °C/min to 235 °C no hold, 2.9 °C/min to 300 °C hold until OCDD eluted. The injector, ion source, and transfer line temperatures were 280 °C. Instrumental performance was evaluated in terms of HRMS sensitivity, GC performance, system stability, and instrumental accuracy. HRMS sensitivity was demonstrated before each analytical run by injecting 0.5 pg of 2,3,7,8-TCDD on-column and ensuring that the resulting peak is detected with a signal-tonoise ratio of 3:1 or greater. To check the chromatographic resolution of the stationary phase, a column performance mixture containing 1,2,3,4-, 1,2,3,7-, 1,2,3,8-, 2,3,7,8-, and 1,2,3,9-TCDD isomers (Column Performance isomers) was analyzed prior to each run in order to confirm that the 2,3,7,8TCDD peak is separated from its closest neighbors by a valley that is 30% or lower. To assess instrumental stability and accuracy, a midlevel calibration standard was run before and after each set and quantified to ensure that the native concentrations were within 20% of the expected value and that the 13C-labeled surrogate concentrations were within 30% of the expected values. APGC-MS/MS Analysis. Sample analysis was performed using a Xevo TQ-S equipped with atmospheric pressure ionization source (Waters Corporation, Milford, MA), an Agilent 7890A gas chromatograph, and Agilent 7693 autosampler (Agilent Technologies, Santa Clara, CA). A 60 m × 0.18 mm × 0.10 μm Rtx Dioxin-2 (Restek, Bellefonte, PA) column was used for the analysis. Approximately 1.0 m × 0.32 mm stainless steel Sulfinert tubing (Restek, Bellefonte, PA) was coupled to the end of the column to act as a transfer line into the ion source. Helium carrier gas was used, and the GC was operated in splitless mode at a flow rate of 1.1 mL/min. The injector was maintained at a temperature of 290 °C and utilized a 4.0 mm drilled hole Uniliner (Restek, Bellefonte, PA). Samples were injected under these conditions at a volume of 0.5 μL. The GC oven temperature program was as follows: initial oven temperature 120 °C hold 1 min, 35 °C/min to 200 °C no hold, 4.5 °C/min to 280 °C hold 8 min, 20 °C/min to 330 °C hold 15 min. The Xevo TQS triple quadrupole mass spectrometer source was run under dry conditions to promote charge transfer ionization. Nitrogen was supplied by an NM32LA nitrogen generator from Peak Scientific (Billerica, MA) and was used as the auxiliary gas, maintained at a flow rate of 400 L/h. Argon was utilized as the collision gas at a flow rate of 0.18 mL/min. Cone gas flow was initially turned off for the first 8 min of the



RESULTS AND DISCUSSION Reduction of Carry-Over. Because of the enhanced sensitivity of the APGC-MS/MS system, a systematic approach to reduce carry-over between samples had to be developed. Carry-over was influenced by the parameters and conditions of the inlet/injector. A series of inlet liners, syringes, and wash solvents were tested to determine conditions that eliminated carry-over of the dioxin compounds. The original inlet conditions used a single gooseneck split/splitless liner, a 10 μL Hamilton syringe, A and B wash solvents (three washes in each) were both nonane, and three samples pumps were performed prior to injection. The parameters listed in Table 1 Table 1. Results of Carry-Over on Inlet Parametersa parameter tested original conditions no sample pumps no glass wool in liner SGE syringe SGE gastight syringe Hamilton gastight syringe toluene wash solvents (× 5 each) Uniliner 1.0 μL injection Uniliner 0.5 μL injection

standard (peak area)

blank (peak area)

percent carryover

657 113 1 756 220 555 436 604 579 549 098 602 524 158 821

1 056 1 461 155 68 76 102 31

0.161 0.083 0.028 0.011 0.014 0.017 0.020

350 458 106 102

4 10

0.001 0.009

a

Standard and blank (analyzed immediately following the standard) values are measured as the peak area of 1,2,3,7,8-PeCDF.

C

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Figure 1. Chromatogram demonstrating the sensitivity of the APGC-MS/MS. Congener identifications are, from left to right, 1,3,6,8-, 1,3,7,9-, 1,3,7,8-, 1,4,7,8-, 1,2,3,4-, and 2,3,7,8-tetrachlorodibenzo-p-dioxin.

Table 2. Comparison of the Soil and Fish Matrix MDL and % RSD Values Determined for Both the APGC-MS/MS and GCHRMS Instruments fish matrix n = 10

soil matrix n = 10 cmpd

units

APGC-MS/ MS MDL

APGC-MS/MS % RSD

GC-HRMS MDL

GC-HRMS % RSD

APGC-MS/ MS MDL

APGC-MS/MS % RSD

GC-HRMS MDL

GC-HRMS % RSD

2,3,7,8-TCDF 2,3,7,8-TCDD 1,2,3,7,8-PeCDF 2,3,4,7,8-PeCDF 1,2,3,7,8-PeCDD 1,2,3,4,7,8-HxCDF 1,2,3,6,7,8-HxCDF 1,2,3,7,8,9-HxCDF 2,3,4,6,7,8-HxCDF 1,2,3,4,7,8-HxCDD 1,2,3,6,7,8-HxCDD 1,2,3,7,8,9-HxCDD 1,2,3,4,6,7,8-HpCDF 1,2,3,4,7,8,9-HpCDF 1,2,3,4,6,7,8-HpCDD OCDF OCDD

pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g pg/g

0.17 0.15 1.3 0.48 0.39 0.78 0.54 0.41 0.37 0.62 0.40 0.35 0.28 0.56 0.41 0.74 1.4

9.7 11 15 6.6 5.2 11 7.0 6.3 5.1 8.7 5.9 3.9 3.9 6.6 5.5 5.5 9.0

0.68 0.80 2.6 2.2 3.9 2.3 1.0 2.2 2.3 3.8 3.0 4.3 3.4 4.9 1.6 4.9 4.5

2.5 2.9 2.0 1.7 2.8 1.6 0.76 1.7 1.8 2.7 2.1 2.9 2.7 3.5 1.2 1.7 1.6

0.21 0.23 2.0 0.31 0.55 0.53 0.30 0.51 0.54 0.70 0.40 0.76 0.50 0.84 1.2 2.0 1.3

18 23 33 5.5 9.2 9.9 5.2 9.6 10 13 7.3 10 9.2 14 22 17 9.9

0.77 4.3 3.5 2.6 1.8 3.0 2.7 2.7 1.9 6.0 0.82 4.1 5.8 2.9 4.8 3.7 7.5

2.2 2.4 2.0 1.5 1.0 1.7 1.6 1.6 1.0 1.8 2.3 2.3 3.4 1.6 2.5 2.1 2.2

were altered one at a time to determine which parameters were the largest contributors to sample carry-over. Level of carryover was assessed by monitoring peak area of the 1,2,3,7,8pentachloro dibenzofuran (1,2,3,7,8-PeCDF) peak. The pentachloro furan was chosen because it exhibited the largest carryover of all the chlorinated dioxins and furans monitored. Table 1 outlines the results displayed as peak areas of the 1,2,3,7,8PeCDF peak from an injection of the highest calibration standard as well as the same peak present in a nonane solvent blank injection run immediately after the standard injection. The original set of inlet and injection conditions resulted in a 0.161% carry-over. On a percentage basis, this seems fairly small, but if it is considered that the on-column concentration

of 1,2,3,7,8-PeCDF was 100 pg, 0.161% carry-over is 0.161 pg or 161 fg. This is significant since this is in the concentration range of the lowest calibration standard. As seen by the results in Table 1, the initial step of simply removing sample pumps prior to injection was able to reduce carry-over by approximately a factor of 2. Removing the glass wool in the inlet liner allowed for a further reduction of the carry-over, a total of approximately a factor of 6. For the remaining carryover tests, the glass wool was removed from the inlet liner and no sample pumps were performed prior to injection. The testing of different manufacturer’s syringes and different types of syringes (gastight versus conventional) did not significantly differ from one another. Therefore, a certain syringe was not D

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Figure 2. Graphical representation of the comparison of quantitative analysis of NIST 1944 Waterway Sediment reference material on both the APGC-MS/MS and GC-HRMS instruments.

calculated by using the equation MDL = SD × t, where SD is the standard deviation of data measurement and t is the 98% confidence interval t-value for n − 1 samples.10 The results for the MDL study on both instruments are tabulated in Table 2. The MDL values are considerably lower on the APGC-MS/ MS instrument for both the soil and fish matrix. MDL values determined on the APGC-MS/MS were 2 through 18-fold lower as compared to the GC-HRMS. On the other hand, the % RSD varied more overall on the APGC-MS/MS than the GC-HRMS, but it should be noted that the samples were run on the APGC-MS/MS instrument using a 20-fold dilution. The increased RSD values could be attributed to detection of a much smaller concentration of each compound. APGC-MS/MS vs GC-HRMS: Reference Samples. Sediment Matrix Reference Materials Comparison. Two different sediment reference materials were chosen for comparison between the two analytical systems, WMS-01 Lake Sediment and NIST 1944 Waterway Sediment. Figure 2 demonstrates a graphical representation of the quantitative comparison of the NIST 1944 Waterway Sediment reference material. Both sediment reference materials demonstrated similar trends and comparisons. The quantified values in pg/g for each of the 17 chlorinated dioxins and furans present in the samples as well as the certified reference value reported for each sample can be found in Table S3 in the Supporting Information. The APGC-MS/MS values compare very well to the certified reference value for both sediment reference materials. In cases where the APGC-MS/MS quantification was outside of the standard error range of the certified value, the value corresponded well to the GC-HRMS quantification, indicating this was most likely due to sample preparation instead of an instrumental quantification error. For example, in the WMS-01 sample analysis for 1,2,3,4,6,7,8-HpCDD, the certified range is 456−760 pg/g. The APGC-MS/MS and GC-HRMS analyses quantified at 275 ± 7 and 297 ± 5 pg/g, respectively. Both values are outside the certified range but very similar to each other. The same situation is observed in the NIST 1944 sample for 2,3,7,8-TCDD. Furthermore, in the NIST 1944 sample,

preferred over another. Using toluene, as opposed to nonane, as the wash A solvent and washing five times with both wash A and wash B, pre and post injection, also exhibited a decrease in the percentage of carry-over by approximately 8-fold. Finally, the most significant reduction of carry-over was seen by installing a Uniliner as the inlet liner. The GC column makes a pressfit connection directly into the Uniliner, forcing the entire injection volume onto the column. This minimizes the opportunity for analytes to become trapped in the inlet system and available for carry-over in subsequent injections because it comes into contact with less surface area of the injection port. An added benefit to using the Uniliner was the reduction of injection volume to 0.5 μL. The injection of 1.0 μL onto the Uniliner created peak fronting or column overload. Therefore, by using a 0.5 μL injection, the carry-over level is reduced to the single femtogram level, less sample volume is used, and sensitivity is equivalent, or greater, than for splitless injection. In summary, the inlet method and injection process that resulted in the least amount of carry over utilized no sample pumps prior to injection of 0.5 μL of sample into a Uniliner containing no glass wool. Toluene must be used as the pre and post wash solvents. Sensitivity and Method Detection Limits. Sensitivity of the APGC-MS/MS instrument was tested by running a low level tetra-chlorinated dioxin mix containing six TCDD congeners ranging in concentration from 2 to 100 fg (on column). Figure 1 demonstrates the resulting chromatogram from an injection of this standard. The 2.0 fg peak, corresponding to 1,3,6,8-TCDD has a peak-to-peak S:N value of 5. The sensitivity threshold established for the GC-HRMS instrument requires a S:N of 3:1 for an on-column concentration of 500 fg. The concentration detected on the APGC-MS/MS is 250-fold less, with a S:N that exceeds this requirement. Method detection limits (MDL) were determined for each instrument in both a soil and fish matrix. The MDL for each matrix was calculated from 10 replicates of the same set of sample extracts analyzed on both instruments. MDL was E

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Figure 3. Graphical representation of the comparison of quantitative analysis of EDF-2525 Contaminated Fish reference material on both the APGC-MS/MS and GC-HRMS instruments.

APGC-MS/MS system performed well, reporting values below 1.0 pg/g for all relevant compounds except OCDF. The GCHRMS quantification of OCDF was extremely high as well, correlating well to the APGC-MS/MS value, indicating a likely interference. Again, the APGC-MS/MS detected and quantified values lower than most of the < values reported for the GCHRMS instrument. Comparable results were seen also with the EDF-2525 Contaminated Fish reference material. There were some compounds that were quantified higher on the APGCMS/MS instrument relative to the GC-HRMS, but these either fell within, or were close to, the certified range. These typically correlated to the compounds in the reference material with the highest variance in certified reference values. Similarly to the previous fish reference material, two of the five values reported as a < quantification on the GC-HRMS were quantified at a significantly lower concentration on the APGC-MS/MS

there are some compounds where the APGC-MS/MS quantification compares well with the certified range and the GC-HRMS quantification does not. It should be mentioned that the 2,3,7,8-TCDF quantification in the NIST 1944 sample using the GC-HRMS was approximately 3 times higher than the certified value. This is most likely due to coelutions of 2,3,7,8-TCDF with other TCDF congeners from using a DB-5 column. In a study of all tetra- through octa-dioxin and furan congeners on various stationary phases, Ryan et al. has documented coelutions of 2,3,7,8-TCDF with 2,3,4,8-, 2,3,4,7-, 2,3,4,6-, 1,2,4,9-, and 1,2,7,9- TCDF.26 By utilizing the Rtx-Dioxin2 column for this separation in the APGC-MS/ MS analysis, 2,3,7,8-TCDF was effectively resolved from the potential coeluting congeners. Fish Tissue Matrix Reference Materials Comparison. Three different fish tissue reference materials were chosen for comparison between the two analytical systems; WMF-01 Freeze-dried Fish, EDF-2524 Clean Fish Tissue, and EDF-2525 Contaminated Fish Tissue. Figure 3 demonstrates a graphical representation of the quantitative comparison of the EDF 2525 Contaminated Fish reference material. All fish reference materials demonstrated similar trends and comparisons. The quantified values in pg/g for each of the 17 chlorinated dioxins and furans present in the samples, as well as the certified reference value reported for each sample can be found in Table S4 in the Supporting Information. The fish reference comparisons are more difficult to identify distinct trends from due to large variability of standard deviation in the certified values. For the WMF-01 reference material, the APGC-MS/MS quantified values either correlated well with the GC-HRMS quantified values or fell within the range of the certified values. In some cases where a value below the detection limit of the GC-HRMS was reported, denoted as a < value, the APGC-MS/MS system was able to detect and quantify a lower value. The EDF-2524 reference sample provided an interesting test of system sensitivity as it was a clean fish tissue reference material with all concentrations well below 1.0 pg/g, with the exception of 2,3,7,8-TCDF. The



CONCLUSIONS A method was successfully developed for the analysis of polychlorinated dioxins and furans using an APGC-MS/MS and was directly compared to a validated methodology using a GC-HRMS instrument. Utilizing MRM transitions for the 17 regulated dioxins and furans demonstrated accurate and extremely sensitive identification of these compounds. The APGC-MS/MS was able to detect as low as 2 fg of TCDD with a very acceptable S:N value of approximately 5:1. Having an instrument available that is reliably accurate at such low levels is extremely important for analysis of environmentally important compounds in challenging matrixes, such as dioxins. Additionally, a method detection limit study (MDL) resulted in significantly lower MDL values for the APGC-MS/MS compared to those established on the GC-HRMS system, in both sediment and fish matrixes. In addition to the instrument being highly sensitive, it is generally easy to use and maintain, making it a valid addition to commercial laboratories that otherwise may not be able to perform these types of analyses. The high sensitivity of the APGC-MS/MS instrument posed initial issues with sample carry-over. This was resolved by F

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Article

Analytical Chemistry

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establishing a set of inlet conditions that eliminated sample carry-over. Important to resolving carry-over was the use of a Uniliner inlet liner, in which the column is sealed into the glass liner, eliminating major pathways for compound interaction with the injection port. Following the institution of these inlet conditions, carry-over was effectively eliminated from the system. Further comparisons between the APGC-MS/MS and GC-HRMS instrumentations were made using certified reference materials. Again, the APGC-MS/MS instrument performed equally to the GC-HRMS in the reference material comparison. The work outlined above demonstrates the validity of using APGC-MS/MS as an instrument for dioxin analysis. Not only is it possible to monitor for currently regulated compounds, such as the 17 PCDDs and PCDFs, but with the selectivity, sensitivity, and speed of MRM acquisition mode, these methods can be expanded upon to monitor for a wider range of environmental contaminants that are becoming of increasing concern. This work demonstrates that APGC-MS/MS can be applied to analysis of halogenated dioxins and furans.



ASSOCIATED CONTENT

S Supporting Information *

Target masses monitored in SIM mode, dwell times, delay times, and isotope ratios for GC-HRMS analysis; MRM transitions monitored, retention time windows, dwell times cone voltages, and collision energies for APGC-MS/MS analysis; and quantified values for all soil and fish reference materials calculated for APGC-MS/MS and GC-HRMS analysis. The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/ acs.analchem.5b01705.



AUTHOR INFORMATION

Corresponding Author

*E-mail: fl[email protected]. Phone: 814-863-6805. Notes

The authors declare no competing financial interest.



ACKNOWLEDGMENTS The authors would like to acknowledge the following (1) Waters Corporation for the Xevo APGC-TQS system, (2) Brock Chittim, Alex Konstantinov, and Jeff Klein at Wellington Laboratories for technical assistance, (3) Terry Kolic from the Ontario Ministry of the Environment and Climate Change (MOECC) for assistance with the GC-HRMS data, and (4) Restek Corporation for various chromatographic consumables and columns.



REFERENCES

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DOI: 10.1021/acs.analchem.5b01705 Anal. Chem. XXXX, XXX, XXX−XXX