Tandem Mass Spectrometry of Polychlorodibenzo-p-Dioxin and

Roger S. Mercer .... were performed on a Varian (Walnut Creek, CA) Saturn 4D GC/MS equipped with a Varian waveform generator, autosampler (Model 8200)...
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Anal. Chem. 1996, 68, 2345-2352

Tandem Mass Spectrometry of Polychlorodibenzo-p-Dioxin and Polychlorodibenzofuran in a Quadrupole Ion Trap. 1. Comparison of Single-Frequency, Secular Frequency Modulation, and Multifrequency Resonant Excitation Modes Jeffry B. Plomley† and Raymond E. March*

Department of Chemistry, Trent University, Peterborough, Ontario, Canada K9J 7B8 Roger S. Mercer

Dioxin/Furan Unit, Laboratory Services Branch, Ontario Ministry of the Environment and Energy, Etobicoke, Ontario, Canada M9P 3V6

An investigation has been carried out on three modes of resonant excitation in the development of an analytical protocol for the determination of polychlorodibenzo-pdioxins (PCDDs) and polychlorodibenzofurans (PCDFs) by ion trap tandem mass spectrometry. The resonant excitation modes investigated are single-frequency irradiation (SFI), secular frequency modulation (SFM), and multifrequency irradiation (MFI) of isolated molecular ion species. Each excitation mode was examined for fragmentation channel selectivity so as to optimize instrument sensitivity. Collision-activated dissociation mass spectra obtained with each excitation mode are compared with those obtained by triple-stage quadrupole mass spectrometry (TSQMS). While the same reaction channels (corresponding to losses of Cl•, COCl•, and 2COCl• for PCDDs and Cl•, COCl•, and COCl2 and COCl3• for PCDFs) were observed for each excitation mode, the fragment ion relative abundances differed among the resonant excitation modes and showed that internal energy deposition in the precursor ion was in the order MFI > SFI > SFM. In each resonant excitation mode, the observed conversion efficiency for loss of COCl• exceeded that observed with TSQMS. The procedure of tuning SFI to ion secular frequencies was laborious, while the required duration of irradiation with SFM was excessively long compared to the gas chromatographic time scale. The tuning requirements of MFI using 1 and 2 kHz bandwidth pulses were less rigorous than those for SFI, and the duration of irradiation was compatible with the gas chromatographic time scale. Polychlorodibenzo-p-dioxins (PCDDs) and polychlorodibenzofurans (PCDFs) are two classes of compounds which are of environmental concern because of the high toxicity of those † Present address: Thermo Instruments (Canada), 5716 Coopers Ave., Unit 1, Mississauga, ON, Canada L4Z 2E8.

S0003-2700(95)01160-7 CCC: $12.00

© 1996 American Chemical Society

isomers with 2,3,7,8-tetrachloror substitution.1 In the United States alone, some 500 kg of PCDDs/PCDFs are released annually into the environment2 from municipal and industrial waste incinerators,3,4 automobile exhaust,5 pulp and paper mill effluents,6,7 and the manufacture of chlorophenol products.8 The potential threat to human health posed by PCDDs/PCDFs in the environment is confirmed in the recent reassessment of 2,3,7,8TCDD and related compounds,9 carried out by the United States Environmental Protection Agency (U.S. EPA). In view of adverse health effects and the widespread and persistent presence of PCDDs/PCDFs in the environment, these compounds are monitored in air, rain, effluents, soil, and biota matrices. Currently, both the U.S. EPA and the Ontario Ministry of the Environment and Energy (MOEE) monitor the total concentration of all PCDD/PCDF congener groups (i.e., total tetrachlorinated dioxins, total pentachlorinated dioxins, etc.) and the concentrations of each of the 17 2,3,7,8-substituted toxic isomers. Such determinations are costly since PCDD/PCDF determinations require extensive sample preparation, the use of 16 expensive 13C12-2,3,7,8-substituted internal standards,10 and mass spectrometers of high capital cost. High-resolution mass spectrometry (HRMS)11,12 or triple-stage quadrupole mass spec(1) Clement, R. E. Anal. Chem. 1991, 63, 1130A-1139A. (2) Silbergeld, E. K. Organohalogen Compd. 1995, 26, 1-6. (3) Olie, K.; Vermeulen, P. L.; Hutzinger, O. Chemosphere 1977, 6, 455. (4) Markland, S.; Kjeller, L. O.; Hansson, M., Tysklind, C.; Rappe, C.; Collazo, H.; Dougherty, R. In Chlorinated Dioxins and Dibenzofurans in Perspective; Rappe, C., Choudhary, G., Keith, L., Eds.; Lewis: Chelsea, MI, 1986; p 72. (5) Markland, S.; Andersson, R.; Tysklind, M.; Rappe, C.; Egeback, K.; Bjorkman, E.; Grigoriadis, V. Chemosphere 1990, 20, 553. (6) Swanson, S. E.; Rappe, C.; Malstrom, A.; Kringstad, K. P. Chemosphere 1988, 17, 681. (7) Clement, R. E.; Tashiro, C.; Suter, S.; Reiner, E. J.; Hollinger, D. Chemosphere 1989, 18, 1189. (8) Rappe, C.; Andersson, R.; Bergqvist, P. A.; Brohede, C.; Hansson, M. Chemosphere 1987, 16, 1603. (9) U.S. Environmental Protection Agency. Health Assessment for 2,3,7,8-TCDD and Related Compounds; External review draft EPA/600/BP-92/001a-c, 1992. (10) U.S. EPA. Method 1613. Tetra-through Octa-chlorinated Dioxins and Furans by Isotope Dilution HRGC/HRMS, Revision A; United States Environmental Protection Agency: Washington, DC, 1990.

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trometry (TSQMS)13-15 is necessary in order to differentiate between (a) PCDDs/PCDFs and interferents, such as polychlorinated biphenyls,16 and (b) 13C12-PCDFs and native PCDDs, whose isotopic clusters overlap when analyzed by low-resolution mass spectrometry (LRMS). Recently, we reported a rapid screening technique for the detection and quantitation of 2,3,7,8-TCDD using a quadrupole ion trap operated tandem mass spectrometrically (MS/MS).17,18 While the sensitivity of the ion trap MS/MS technique (500 fg/ µL instrumental detection limit with a S/N of 5:1) was shown to be comparable to that of TSQMS, only one scan function could be applied to the determination of a single congener group (i.e., tetra) in each chromatographic run. A further impediment, imposed by the software at that time, was the inability to perform quantitation using internal standards which coeluted chromatographically with their native analytes. These impediments have been overcome by software advances such that it is possible now to deconvolute mass spectra generated from analytes which coelute chromatographically.19,20 When operated in MS/MS mode, the ion trap is now capable of multiple-reaction monitoring (MRM) daughter ions from tetra- to octa-PCDDs/PCDFs in a single chromatographic acquisition. Thus, we have attempted to develop an MS/MS method for the ultratrace detection and quantitation of the tetra- to octa-PCDDs/PCDFs using isotopic dilution techniques. The first phase of this investigation involves both optimization of molecular ion isolation and directed fragmentation of the tetra- to octa-PCDDs/PCDFs and is reported here. The second phase is concerned with the analytical aspects of PCDDs/PCDFs analysis by GC/MS/MS (i.e., linear dynamic range, instrumental and method detection limits, and quantitation in a number of environmental matrices) and will be reported later. Essential to the development of an analytical method for the analysis of PCDDs/PCDFs by MS/MS with an ion trap is optimization of the collision-activated dissociation (CAD) process by variation of the experimental parameters: the amplitude of the resonant potential, the duration of irradiation and ion cooling periods, and the mode of resonant excitation. The major fragmentation channel observed for these compounds when analyzed by MS/MS with the TSQMS is the loss of COCl•.16 Three modes of resonant excitation of ions isolated within an ion trap have been examined in an attempt to optimize this major dissociative channel: single-frequency irradiation (SFI), secular frequency modulation (SFM), and multifrequency irradiation (MFI). The latter two modes obviate the laborious procedure of SFI, wherein (11) Clement, R. E.; Tosine, H. M. Mass Spectrom. Rev. 1988, 7, 593. (12) Taguchi, V. Y.; Reiner, E. J.; Wang, D. T.; Meresz, O.; Hallas, B. Anal. Chem. 1988, 60, 1429. (13) Tondeur, Y.; Niederhut, W. N.; Campana, J. E.; Missler, S. R. Biol. Environ. Mass Spectrom. 1987, 14, 449. (14) Chess, E. K.; Gross, M. L. Anal. Chem. 1980, 52, 2057. (15) Reiner, E. J.; Shellenberg, D. H.; Taguchi, V. Y. Environ. Sci. Technol. 1991, 25, 110. (16) Method for the Determination of Polychlorinated Dibenzo-p-Dioxins and Polychlorinated Dibenzofurans in Fish; Ontario Ministry of the Environment and Energy: Etobichoke, ON, Canada, 1993. (17) Plomley, J. B.; Koester, C. J.; March, R. E. Org. Mass Spectrom. 1994, 29, 372-381. (18) Plomley, J. B.; Koester, C. J.; March, R. E. Proceedings of the 42nd ASMS Conference on Mass Spectrometry and Allied Topics, Chicago, IL, May 29June 3, 1994; p 718. (19) Plomley, J. B.; Mercer, R. S; March, R. E. Proceedings of the 43rd ASMS Conference on Mass Spectrometry and Allied Topics, Atlanta, GA, May 2226, 1995; p 230. (20) Hamelin, G.; Brochu, C.; Moore, S. Organohalogen Compd. 1995, 23, 125130.

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an applied single-frequency waveform is matched empirically to the fundamental axial secular frequency, ωz,0, of the ion. While ωz,0 for a given ion species can be calculated readily,21 the actual fundamental axial frequency of ions focused near the center of the ion trap is subject to frequency shifts due to space charge effects,23 nonideal trap geometry, concentration ranges, the radio frequency (rf) trapping potential, and the inadequacies associated with automated frequency calibration. Let us examine each mode of resonant excitation in turn. SFI is carried out at a fixed value of qz by the application of a supplementary ac signal across the end-cap electrodes in dipolar fashion. This usual mode of resonant excitation has been well documented and characterized using thermometer molecules such as n-butylbenzene.22 The optimization of SFI requires an examination of qz, a matching of the applied frequency with ωz,0, and variation of both the duration of irradiation and the amplitude of the ac potential. SFM involves changing, modestly, the amplitude of the rf drive potential applied to the ring electrode such that ions move in and out of resonance with the applied singlefrequency waveform.23 Modulation of the rf amplitude changes qz, which in turn affects βz (a complex function of the trapping parameters) and thus ωz,0 so as to produce a frequency sweep over a narrow range (e.g., 1 or 2 kHz). The CAD efficiency of SFM depends upon the ion dwell time per voltage step, the modulation range (i.e., the ratio of off- and on-resonance durations), the amplitude of the irradiating waveform, and the total duration of the CAD episode. MFI involves the application of a waveform consisting of several frequency components while the value of the trapping parameter qz is held constant. The frequency components or bandwidth of the irradiating waveform bracket the axial secular frequency of the ion so as to compensate for frequency shift due to space charge effects, the stretched geometry of the ion trap, the imprecision of the ion trap frequency calibration procedure, and ion axial excursion from the ion trap center. The number of frequency components within a given bandwidth is determined by the frequency interval of 500 Hz: here, bandwidths of 1 and 2 kHz were used. While other types of multifrequency irradiation have been reported, such as random noise,24 swept-frequency,25 and broad-band excitation,26-29 smallbandwidth irradiation waveforms such as those used in this investigation have received little attention. When random noise and broad-band excitation waveforms are used, nascent product ions can be ejected by a frequency component in the waveform, resulting in reduced ion trap sensitivity. (21) March, R. E.; Hughes, R. J. Quadrupole Storage Mass Spectrometry; Chemical Analysis Series 102; John Wiley and Sons: New York, 1989. (22) Louris, J. N.; Cooks, R. G.; Syka, J. E. P.; Kelley, P. E.; Stafford, G.; Todd, J. F. J. Anal. Chem. 1987, 59, 1677-1685. (23) Schachterle, S. Proceedings of the 42nd ASMS Conference on Mass Spectrometry and Allied Topics, Chicago, IL, May 29-June 3, 1994; p 714. (24) McLuckey, S. A.; Goeringer, D. E.; Glish, G. L. Anal. Chem. 1992, 64, 14551460. (25) McLuckey, S. A.; Goeringer, D. E.; Glish, G. L. J. Am. Soc. Mass Spectrom. 1991, 2, 11. (26) Julian, R. K.; Cooks, R. G. Anal. Chem. 1993, 65, 1827-1833. (27) Vedel, F.; Vedel, M.; March, R. E. Int. J. Mass Spectrom. Ion Processes 1991, 108, R11. (28) McLuckey, S. A.; Goeringer, D. E.; Glish, G. L. Proceedings of the 40th ASMS Conference on Mass Spectrometry and Allied Topics, Washington, DC, May 31-June 5, 1992; p 1013. (29) Vedel, F.; Vedel, M.; March, R. E. Int. J. Mass Spectrom. Ion Processes 1990, 99, 125.

EXPERIMENTAL SECTION Chemicals. A solution containing 200 pg/µL T4CDD/F, 1000 pg/µL each of P5CDD/F, H6CDD/F, H7CDD/F, and 2000 pg/µL of OCDD/F was supplied by the MOEE for the optimization of the CAD process. Note that T4CDD/F is an accepted abbreviation for tetrachlorodibenzo-p-dioxin/tetrachlorodibenzofuran, where the numerical subscript indicates the number of chlorine substituents on the dioxin or furan skeleton; thus P5 indicates a pentachlorinated species, H6 a hexachlorinated species, etc. Instrumentation. All experiments were performed on a Varian (Walnut Creek, CA) Saturn 4D GC/MS equipped with a Varian waveform generator, autosampler (Model 8200), and septum programmable injector with a high-performance insert. Saturn version 5.0 software was used for data acquisitions, while prototype software (Toolkit) was utilized for the application of multiple scan functions during a chromatographic run. The ion trap was operated in mode II21 (i.e., with the end-cap electrodes grounded) with dipolar resonance ejection at a fixed frequency of ∼485 kHz (1-4 V0-p) to give mass-selective ejection at qz ≈ 0.89. The trap manifold temperature was held constant at 260 °C. Helium buffer gas was present in the ion trap cavity at ∼1 mTorr. Optimal sensitivity of the ion trap was achieved using a filament emission current of 50 µA and an electron multiplier voltage of 1800 V (the latter tuned to give a gain of 105). The tunable target ion current for the AGC algorithm was 35 000, thus ensuring optimal ionization times for all analytes under examination. Mass calibration (V0-p/amu) was performed in EI mode using perfluorotributylamine (PFTBA, or FC-43). Automated frequency calibrations were achieved using m/z 69 and 131 from PFTBA; this calibration procedure requires some explanation. The calculation of ωz,0 (and thus the single frequency required for resonant excitation) is based upon the determination of a trap calibration factor (TCF). The TCF is calculated automatically using a calibrant ion from PFTBA, the theoretical frequency required to excite that ion and the empirically determined rf drive amplitude required to bring the applied waveform into resonance with the axial frequncy of ion motion. For example, let us consider m/z 69 from PFTBA. A default low-mass cutoff of 300 V0-p is specified that corresponds to qz ) 0.6147 for m/z 69; the corresponding theoretical ωz,0 for this value of qz is 250.0 kHz. Thus, a single-frequency waveform of 250.0 kHz, of predetermined optimal amplitude, is applied so as to reduce the ion signal due to m/z 69 to a preset minimum value (which is close to zero and corresponds to complete dissociation of m/z 69). When the preset minimum current due to m/z 69 is not reached, the rf amplitude is changed in defined increments until m/z 69 falls into resonance with the applied waveform. In this example, let us define 302 V0-p as the rf amplitude at which the preset minimum current is observed (and corresponds to the disappearence of m/z 69). At 302 V0-p, m/z 69 has a ωz,0 of 252.3 kHz. This frequency differs by +2.3 kHz from the theoretical frequency. A TCF may now be determined such that TCF ) (252.3-250)/250 ) 0.0092. The algorithm for calculating waveform frequencies utilizes the lowmass cutoff value, ion mass/charge ratio, and the most recent TCF value. For example, when a low-mass cutoff of 300 V0-p is specified for m/z 69, the waveform algorithm calculates the applied frequency from the relationship

applied frequency ) theoretical ωz,0(1 + TCF)

(1)

Gas chromatography and injector conditions necessary for the optimal separation of all dioxins and furans have been described elsewhere.17,18 A 30 m (rather than 60 m) × 0.25 mm i.d. × 25 µm DB-5 (diphenyldimethylpolysiloxane) fused-silica capillary column (J&W Scientific, Folsom, CA) was used in order to expedite MS/MS optimization. The transfer line temperature was held at 260 °C. Parent Ion Isolation. A custom-made scan function was used for ionization, parent ion isolation, CAD, and daughter ion analysis. Parent ion isolation was performed in two stages, corresponding to coarse isolation and fine isolation. The isolation of m/z 320 from T4CDD will serve as an example. EI (∼70 eV) was performed at an rf drive amplitude of 1000 V0-p (low-mass cutoff of 160 amu), such that qz ≈ 0.44 for m/z 320. The corresponding Dehmelt trapping well depth (vide infra) of 55 eV ensures that nascent m/z 320 ions are trapped efficiently. During the ionization period, a multifrequency waveform (MFW-A) was applied at 30 V0-p to eject resonantly ions of m/z < 320, thereby ejecting matrix ions and reducing space charge. After ionization, a second multifrequency waveform, MFW-B, was used to eject ions of m/z > 320. The applications of MFW-A and MFW-B represent coarse isolation, since a 10 kHz notch was created between the two waveforms. This frequency notch corresponded to the secular frequencies of m/z 312-329; thus, ions in a mass range of 17 amu remained stored. Unit ion isolation of m/z 320 was achieved by ramping the rf amplitude, in two stages, so as to eject m/z 319 at βz ≈ 1. During this low-mass ejection phase, axial modulation was engaged. The rf amplitude was then decreased slightly with simultaneous application of MFW-C at 20 V0-p, a broad-band waveform capable of ejecting high-mass ions of m/z>320. Since the time over which the rf amplitude is decreased is less than that necessary for highmass ion ejection, MFW-C was applied for a further 5 ms at 20 V0-p while the rf amplitude was held constant. The strategy for high-mass ejection is rationalized on the basis that, as qz increases, the difference in ωz,0 between successive masses increases. Therefore, the risk of parent ion ejection by MFW-C is minimized at large qz values. In general, parent ions from the most intense peak in the mass spectral isotopic clusters of T4CDD/F, P5CDD/F, H6CDD/F, H7CDD/F, and OCDD/F were isolated for CAD optimization. After nominal parent ion isolation, the rf amplitude was adjusted to give an appropriate qz value for the ion prior to CAD. A cooling time of 1 ms or more can be inserted into the scan function at this juncture. Daughter ions formed by CAD were scanned out from the ion trap, with axial modulation active, at a rate of 5555 amu/ s. The narrowest practical mass scanning range was specified in order to reduce the duration of this portion of the scan function and, thereby, increase the number of scan functions across a chromatographic peak. Resonant Excitation Studies. All resonant excitation studies were performed with the parent ion at a working point of qz ≈ 0.4. A constant working point permits ready comparison among the resonant excitation modes since ions of different mass/charge ratios will have the same secular frequency. The selection of this working point was based upon previous experience with dioxins.18,19 (i) Single-Frequency Irradiation. Parent ions were irradiated for 30 ms at qz ≈ 0.4 with a single-frequency sinusoidal waveform of variable amplitude, applied across the end-cap Analytical Chemistry, Vol. 68, No. 14, July 15, 1996

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Table 1. Parent Ion Mass, Resonant Excitation Cutoff Mass, and Optimal Irradiation Frequency Offsets for the Tetra- to Octachlorinated Dioxins and Furans

congener group

parent ion mass

T4CDF P5CDF H6CDF H7CDF OCDF

306 340 374 408 444

T4CDD P5CDD H6CDD H7CDD OCDD

320 356 390 424 458

resonant excitation cutoff mass for qz ) 0.4

optimal irradiation frequency offset (Hz)

Furans 134.8 149.7 164.8 179.7 195.6

+400 +400 +400 +300 +300

140.9 156.8 171.8 186.8 201.8

+200 +300 +300 +400 +300

Dioxins

electrodes in dipolar fashion. To match the applied waveform frequency with ωz,0, it was necessary to offset the applied frequency from its calibration-based value by as much as +400 Hz. Compiled in Table 1 are the mass/charge ratios of the parent ions isolated from each PCDD/PCDF congener group, along with the low-mass cutoffs required to maintain a constant trapping parameter of qz ≈ 0.4 for each parent ion and the frequency irradiation offsets. (ii) Secular Frequency Modulation. SFM was implemented by varying the rf voltage amplitude. To optimize the conversion efficiency associated with the loss of COCl•, SFM experiments were carried out under two sets of conditions: (1) the parent ion was modulated over a range of ∆V ) 6 V0-p, with an ion dwell time of 5 ms/V0-p, giving five CAD segments lasting 150 ms, and (2) the parent ion was modulated over a range of ∆V ) 4 V0-p with an ion dwell time of 5 ms/V0-p, for a total of five CAD segments lasting 100 ms. Each rf voltage amplitude increment corresponds to a change in ωz,0 of ∼200 Hz. Under both sets of conditions, a sine waveform of variable amplitude was applied at ωz,0 for an ion with qz ≈ 0.4; no irradiation frequency offset was applied to the calculated waveform. (iii) Multifrequency Irradiation. MFI involved the construction of waveforms consisting of frequency components at 500 Hz intervals. Conversion efficiencies corresponding to the loss of COCl• were examined as a function of waveform bandwidth and amplitude. Both 1 kHz (three frequency components) and 2 kHz (five frequency components) bandwidth waveforms, centered at the theoretical ωz,0 for an ion with qz ≈ 0.4, were applied in separate experiments for periods of 30 ms. Waveforms were generated by random phasing and were clocked out at a frequency of 2.5 MHz. A time domain signal with 5000 data points corresponds to a sample rate of 400 ns/data point, producing a signal with a 2 ms pulse duration. Thus, for a CAD episode lasting 30 ms, 15 pulses were applied. RESULTS AND DISCUSSION Single-Frequency Irradiation. To examine the accuracy of the frequency calibration and to ensure that the applied waveform was resonant with ion motion, the commercially available Toolkit software was modified to include an irradiation frequency offset. This offset changes the irradiation frequency in increments of 100 Hz from that determined by the calibration procedure. The irradiation frequency calculated by the automated trap calibration 2348

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Figure 1. Ion abundance curves obtained by plotting molecular and fragment ion abundances (expressed as percentages of total ion current) as a function of the amplitude V0-p of a single-frequency waveform: (a) T4CDD and (b) T4CDF. (a) 9, M; 2, [M - COCl]; [, [M - Cl]; b, [M - 2COCl]. (b) 9, M; 2, [M - COCl]; [, [M - COCl - Cl]; b, [M - COCl - 2Cl].

had to be adjusted by as much as +400 Hz (see Table 1) in order to minimize both the amplitude and the duration of the applied waveform required for optimal conversion efficiency (CE), where CE is defined as

CE )

∑D P0

i

(2)

Di represents the signal intensities of the daughter ions and P0 is the pre-CAD signal intensity of the isolated parent ion. The criteria for determining resonance between the applied waveform and ωz,0 are disappearance of the molecular ion signal, maximum total daughter ion signal, and minimum values of both the amplitude and the duration of irradiation. When the optimal irradiation frequency offset was used, CEs for the loss of COCl• were higher by 24% for P5CDF, 29% for P5CDD, and 16% for H6CDD when compared to the CEs without the offset. While the mismatch of the applied waveform and ωz,0 could be rectified by increasing the amplitude of the applied waveform, this corrective procedure tended to affect adversely the efficiency of the CAD process. Ion abundance curves, obtained by plotting molecular and fragment ion abundances (expressed as percentages of total ion current) as a function of the amplitude V0-p of a singlefrequency waveform, were generated for each of the congener groups, as exemplified in Figure 1 for 2,3,7,8-T4CDD (a) and 2,3,7,8-T4CDF (b) (hereafter referred to as TCDD and TCDF, respectively). For comparison, the breakdown curves used for the optimization of the TSQMS by the MOEE are illustrated in Figure 2 for TCDD (a) and TCDF (b) analysis in MS/MS mode; argon was used as the collision gas at a pressure of ∼3.3 mTorr.

b

Figure 2. Ion breakdown curves obtained with a triple-stage quadrupole mass spectrometer, where argon was used as the collision gas at a pressure of ∼3.3 mTorr: (a) T4CDD and (b) T4CDF. (a) 9, M; [, [M - COCl]; 2, [M - 2COCl]; f, [M - Cl]; b, [M - COCl-Cl]. (b) 9, M; [, [M - COCl]; 2, [M - COCl - Cl]; f, [M COCl - 2Cl].

While for all PCDDs and PCDFs, the loss of COCl• was the dominant dissociation channel in both the ion trap and the TSQMS, comparison between the CAD processes in the ion trap and the TSQMS is far from simple. The time scale of the CAD process in the TSQMS is in the microsecond range, while that for the ion trap is in the millisecond range; the target mass in the TSQMS is normally greater than that of helium used in the ion trap; ion kinetic energies in the TSQMS are greater than those attained in the ion trap; and, in addition, the ion activation processes differ. In the TSQMS, the projectile ion suffers some 1-4 collisions with target species, while resonantly excited ions in the ion trap each suffer some 400-600 collisions (in 30 ms) with helium, in which ions undergo a complex process of activation and deactivation. The transfer of ion kinetic energy to internal energy in a single collision is greater in the TSQMS than in the ion trap. In Figure 1a, the precipitous decrease in the TCDD molecular ion abundance as the waveform amplitude is increased from 200 to 300 mV is accompanied by the appearance of product ions [[M - COCl•]+, [M - Cl•]+ and [M - 2COCl•]•+. As the waveform amplitude is increased further, the abundance of the [M-2COCl•]•+ fragment ion increased at the expense of the [M - Cl•]+ fragment ion. While all three fragmentation channels for TCDD shown in Figure 1a were observed also by TSQMS, the [M - (CO)2Cl•]+ fragmentation channel observed by TSQMS was not accessed in the ion trap. In Figure 1b, fragmentation of the TCDF molecular ion in the ion trap occurs over a waveform amplitude range of 300-600 mV0-p, clearly a more gradual breakdown than for TCDD. The onset of fragmentation for TCDF, which occurs at a waveform amplitude some 100 mV0-p greater than that required for TCDD fragmentation, is accompanied by the appearance of the fragments

corresponding to the loss of COCl•, COCl2, and COCl•3. The fragment ion abundances are [M - COCl•]+ > [M - COCl2]•+ ≈ [M - COCl•3]+. In both parts a and b of Figure 1, the maximum abundances of the [M - COCl•]+ product ion are attained at a waveform amplitude of 600 V0-p; thus, a measure of reaction channel selectivity is achieved readily. The ion breakdown curves for TCDD and TCDF obtained with the TSQMS and shown in Figure 2 indicate that the optimal fragmentations of molecular ions to [M - COCl•]+ occur at a collision energy (Elab) of ∼25 eV, resulting in CEs of ∼58% for TCDD and ∼80% for TCDF. In the ion trap, the same reaction channel (i.e., [M - COCl•]+) is accessed for TCDD and TCDF, with CEs of 72% and 89%, respectively. A significant difference between Figures 1 and 2 is that appearance energies for the observation of fragment ions in the TSQMS reflect clearly a hierarchy of activation energies, while all of the fragmentation channels accessed in the ion trap are observed once a critical waveform amplitude is applied. For TCDF in the TSQMS, for example, [M - COCl•]+ and [M - Cl•]+ have appearance energies of 5 and 10 eV, respectively, while those for [M - COCl2]•+ and [M - COCl3•]+ are indistinguishable at ∼12.5 eV. The loss of COCl• from PCDD/F is a rearrangement process (low activation energy, low frequency factor, tight transition state), whereas the loss of Cl• is due to single-bond cleavage (high activation energy, high frequency factor, loose transition state), and yet the production of [M - COCl•]+ increases as a function of waveform amplitude (up to 600 mV0-p only for T4CDF). Such a trend is opposite to that expected if the activation energy for single-bond cleavage was higher than that for rearrangement. Yet, as the TSQMS data indicate that the activation energy for [M COCl•]+ is less than that for [M - Cl•]+, the frequency factor for the loss of COCl• must be higher than that for Cl• loss in order to explain the results observed in the ion trap for TCDD. In Figure 3 are represented in stacked bar graph format the daughter ion average abundances for each of five PCDD congener groups obtained under optimal conditions for each of SFI, SFM, and MFI; the corresponding data for five PCDF congener groups are shown in Figure 4. The SFI waveform amplitudes (in mV0-p) used were T4CDD, 600; P5CDD, 600; H6CDD, 600; H7CDD, 700; OCDD, 600; T4CDF, 600; P5CDF, 700; H6CDF, 700; H7CDF, 900; and OCDF, 1000. For T4CDD, an optimal SFI waveform amplitude of 600 mV0-p was required in order to obtain the highest CE for the COCl• loss reaction channel, and the ratio of [M - COCl•]+/[M - Cl•]+/[M - 2COCl•]•+ was 72:16:4; this ratio can be compared with that obtained from the TSQMS breakdown curve where, at an optimal collision energy of ∼25 eV, the corresponding ratio was 58:2:32. For T4CDF, the ion trap optimal amplitude of 600 mV0-p gave a ratio of 89:4:4:3 for [M - COCl•]+/[M - Cl•]+/[M - COCl2]•+/[M - COCl•3]+, whereas at an optimal collision energy of ∼25 eV on the TSQMS, the corresponding ratio was 80:0:10:7. Such different distributions of daughter ion current for the ion trap and the TSQMS arise from the different activation modes. Clearly, more internal energy is deposited into the parent ion with the TSQMS at optimal collision energies for the loss of COCl•, as shown by the dominance of the higher reaction channel products [M - 2COCl•]+ for T4CDD and [M - COCl2]+ and [M - COCl3•]+ for T4CDF in Figure 2. In Figures 3 and 4, it is indicated that, except for OCDD and OCDF, SFI yields CEs corresponding to the loss of COCl• greater than 70%. A comparison between like congeners of PCDD and Analytical Chemistry, Vol. 68, No. 14, July 15, 1996

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Figure 3. Daughter ion average abundances (expressed as a percentage of the initial molecular ion signal intensity) obtained for each of five PCDD congener groups obtained under optimal conditions for each of SFI, SFM, and MFI.

Figure 4. Daughter ion average abundances (expressed as a percentage of the initial molecular ion signal intensity) obtained for each of five PCDF congener groups obtained under optimal conditions for each of SFI, SFM, and MFI.

PCDF reveals that the optimal amplitude required to effect the loss of COCl• is generally larger for the PCDFs. For example, while a SFI waveform amplitude of 700 mV0-p is optimal for the loss of COCl• from H7CDD, 900 mV0-p is required to effect the same loss from H7CDF. The cumulative evidence from Figures 3 and 4 suggests that a lower activation energy is required to effect the loss of COCl• from PCDD relative to PCDF. The loss of Cl• with SFI is common to both the PCDDs and PCDFs but is more predominant for the PCDDs (as high as 17%) than for the PCDFs (