Anal. Chem. 1985, 57, 1133-1138
1133
Isomer-Specific Determination of Hexachlorodibenzo-p -dioxins by Oxygen Negative Chemical Ionization Mass Spectrometry, Gas Chromatography, and High-pressure Liquid Chromatography Walter F. Miles,* Narine P. Gurprasad, and Greg P. Malis Laboratory Services Division, Food Production & Inspection Branch, Agriculture Canada, Ottawa, Ontario K1A OC6, Canada
All ten hexachlorodlbenrcbpdloxln (HCDD) Isomers, prepared by mlcropyrolysls of chlorophenates, were studled by using mass spectrometry, gas chromatography, and hlgh-pressure liquid chromatography. Of several lonlratlon modes studled, oxygen negatlve chemical lonlratlon was the most sultable for Isomer ldentiflcatlon. Thls technique enabllng Isomer spectflc Identlflcatlon to be carrled out was applled to hexachlorodloxln lmpurltles In pentachlorophenol samples and detected small amounts of 4:2 substituted Isomer not prevlously known.
Polychlorinated dibenzo-p-dioxins (PCDDs) are a group of toxic compounds found in a variety of substrates including chlorinated phenols which are widely used as wood preservatives and fungicides. The PCDDs are formed as a byproduct of the manufacture of chlorinated phenols under alkaline conditions at high temperature. Hexachlorodibenzo-p-dioxins (HCDDs) have been found in pentachlorophenol and tetrachlorophenol (1-9) along with hepta- and octachlorodioxins. There are a total of ten HCDD isomers and the toxicity of the isomers depends on the position of the chlorine substituents, with the most toxic isomers apparently belonging to the 2,3,7,8 substituted group (10). In the case of the HCDDs these isomers include 1,2,3,4,7,8-, 1,2,3,6,7,8-, and 172,3,7,8,9-HCDDs. Because of the toxicity of some PCDDs (11) and their presence as contaminants in industrial chemicals such as chlorinated phenols, there is a need for a highly sensitive and isomer specific analytical method for these compounds, Many methods for the determination of PCDDs have appeared in the literature (12, 13) but all of these methods tend to use chromatography, either gas or liquid, as the means of distinguishing between dioxin isomers. Reviews of the mass spectrometry of PCDDs have appeared (14,15) and a variety of ionization methods have been used including electron impact, positive ion chemical ionization, negative ion chemical ionization, and atmospheric pressure chemical ionization. Of these methods, only oxygen negative chemical ionization (ONCI) using a Townsend discharge source and methane negative chemical ionization (MNCI) show significant differences between dioxin isomers. The discharge source is necessary when using oxygen as a chemical ionization reagent gas because a hot filament would rapidly burn out in such an environment. The two major fragmentation pathways reported for PCDDs using ONCI are shown in Figure 1. Pathway I gives rise to an (M - 19) ion and pathway 11, first reported for 2,3,7,8tetrachlorodibenzo-p-dioxinby Hunt and Harvey (16),leads to the formation of either one or two product ions depending upon the chlorine distribution on the two aromatic rings and
Flgure 1. Fragmentation pathways of PCDDs uslng ONCI. Reprinted with permission from ref 14. Copyright 1979 John Wiley and Sons, Ltd.
yields information defining the number of chlorine atoms on each ring. The present work was undertaken to examine the utility of ONCI using Townsend discharge as a selective mode of ionization for an isomer specific method of analysis for HCDDs. Analytical conditions were found that allow an isomer specific separation and detection of all the ten HCDDs using HRGC, HPLC, and ONCI MS. The procecure is applied to samples of pentachlorophenol.
EXPERIMENTAL SECTION Synthesis of HCDDs. The ten isomers of HCDD were prepared by the pyrolysis of various potassium chlorophenate combinations as described in the literature (5,17). The chlorophenols used were 2,3,4-, 2,3,6-, 2,4,5-, and 2,4,6-tri and 2,3,4,5- and 2,3,5,6-tetra, all from Fluka (Buchs, Switzerland) and all were purified by recrystallization from ethanol/ water or hexane. Pentachlorophenol was 99+ % pure from Reichold Chemicals (New York) and was used without further purification. Additional Reference Samples. 1,2,3,4,7,8-HCDDwas obtained from KOR Isotopes (Cambridge, MA) and 1,2,3,6,7,8-and 1,2,3,6,7,9-HCDDswere gifts from J. J. Ryan (Health and Welfare Canada). Extraction and Cleanup of Pentachlorophenol Samples. A 0.5-g portion of pentachlorophenol was dissolved in 5 mL of benzene (if needed, a small amount of methanol is added dropwise till solution is achieved) and this solution was quantitatively transferred with 3 X 5 mL of benzene to a 35 cm X 22 mm i.d. glass column packed with 50 g of neutral alumina (Fisher A-540 activated at 110 "C overnight). A 135-mL portion of benzene was passed through the column and the eluant collected and concentrated to 1-2 mL on a rotavapor. This solution was transferred quantitatively with 3 X 2 mL of 2% CH2Clz/hexaneto a 25 cm X 1.45 cm i.d. glass column packed with basic alumina (Woelm basic alumina, activity 2) and the column was eluted with 100 mL of 2% CH,Cl,/hexane followed by 100 mL of 30% CH,Cl,/hexane. The 30% CHPCl/hexaneeluant was collected and concentrated to 1-2 mL on a rotavapor, followed by complete removal of solvent under a gentle stream of nitrogen. HPLC. A Waters Model 6000 chromatographic pump was used along with a Schoeffelvariable absorbance detector, Model SF-770, at 235 nm for both reverse-phase and normal-phase HPLC.
0003-2700/85/0357-1133$01.50/00 1985 American Chemical Society
1134
ANALYTICAL CHEMISTRY, VOL. 57, NO. 6, MAY 1985 1118.0,
I
i 1?4679 + 124685 HCOOS
lB8.0,
I
123683 HCUO
i
5E,81
ii
21E
It
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1 I
390
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fi E 156
123469 HCUU
1
il
c
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:46
250
380
358
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I .ii
Flgure 2. ONCI mass spectra of all HCDD isomers.
Jl!,I
c k
288
258
388
358
480
I
ANALYTICAL CHEMISTRY, VOL. 57, NO. 6, MAY 1985
1135
Table I. Relative Sensitivities of ONCI, EI, and MNCI for different PCDDs
PCDD 1,3,7,8-tetra 1,2,4,7,8-penta 1,2,3,4,7,8-hexa 1,2,3,4,6,7,8-hepta octa
ionization method ONCI E1 MNCI 0.38 1.9 3.5 3.9 3.8
0.08 1.8
1.0 1.0 1.0 1.0 1.0
7.7
6.2 6.7
Figure 3. Mass chromatogram of mlz 210 and 246 of all HCDD isomers: 50 m CPS-1, 140-200 at 20 "Clmin, 200-240 "C at 10
"Clmin.
Table 11. Relative Retention Times of All 10 HCDD Isomers on 50 m CPS-1, 140-200 at 20 OC/min, 200-240 at 10 OC/min 37 I
100.0,
HCDD isomer
I
r;0
re1 retention time 1.000
1.006 1.064 1.069 1.117
1.132 1.174
1.232 1.276 was used as a reagent gas in the Townsend discharge mode, source pressure was 0.5 torr, temperature was 140 "C, discharge current was 65 FA, and discharge voltage was 1.2-1.5 kV. E.I. In the electron impact mode source conditions were as follows: temperature, 140 "C; 50 eV; emission current, 0.35 mA. MNCI. Methane negative chemical ionization was carried out at source pressure 0.5 torr, temperature 140 "c,emission current 0.35 mA, and 55 eV. All source temperatures reported in this paper are those set on the instrument. The manufacturer's operating manual indicates the true temperature is 40-50 O C higher.
I00.B
1
SOUHCE lB0C
RESULTS AND DISCUSSION
R.E
150
200
258
300
350
400
Flgure 4. ONCI spectra of 1,2,3,6,7,8-HCDDat different source temperatures.
Sample injection was carried out with a Rheodyne Model 7105 injector with either a 2-mL loop (RP-HPLC) or a 50-pL loop (NP-HPLC). Reverse phase utilized a Du Pont Zorbax ODS preparative column, 250 nm X 9.4 mm i.d., 10 wm particle size, using 100% methanol at a 2 mL/min flow rate. Extracts of pentachlorophenol samples were partially dissolved (some of the octachlorodioxin did not go into solution) in a MeOH:CHC13: toluene (2:7:1) mixture, with up to 1 mL injected. The fraction eluting from 25 to 41 min was collected and analzyed for HCDDs by GC/MS. Normal-phase HPLC was used for the separation of 1,2,4,6,8,9and 1,2,4,6,7,9-HCDDsaccording to the method of Lamparski and Nestrick (17). GC/MS. A Finnigan 4500 mass spectrometer equipped with pulsed positive/negative ion chemical ionization and Townsend discharge ionization was used for this work. Samples were introduced via a Finnigan Model 9610 gas chromatograph equipped with a J and W Model I1 capillary on-column injector. GC columns used in this study included 14 m DB-1,50 m CPS-1, 50 m CPS-2,30 m DB-225, and 30 m DB-5. GC operating conditions varied and are given with the chromatograms. Data acquisition and mass spectrometer control were carried out with a Finnigan 2300 INCOS data system. ONCI. When oxygen (Canadian Liquid Air, U.H.P., 99.99%)
All ten HCDDs were synthesized by micropyrolysis of various potassium chlorophenates according to the procedure of Buser (5). The products were cleaned up on an alumina microcolumn (5) and contained some lower and higher chlorinated dioxins in the case of the PCP pyrolizates as well as the expected HCDD isomers. Oxygen Negative Chemical Ionzation. The ONCI spectra of all ten HCDDs, including the 124689/124679 pair which were not resolved on any of the GC columns used, are given in Figure 2. In addition to the (M - 19) ion at m/z 369 and the ether cleavage ions at m/z 210 and 244, aJl the isomers show clusters at m/z 388 (M-), m/z 353 (M - Cl)-, and weaker peaks a t m / z 334 (M - 19 + 35)- and m/z 306 (M - 19 63)-. It is easy to distinguish between the 3:3 and 4:2 chlorine substitution in the HCDDs by the ether cleavage ions at m / z 210 for 3:3 and m/z 246 for 4:2 substitution (Figure 3). In contrast to the findings of Hass et al. (14) it was found that in most cases the base peak of the mass spectra of the HCDDs was due to (M - C1)- rather than (M - 19)- although this may have resulted from the higher temperature at which our spectra were obtained. The essential fact that emerges from a study of the ONCI mass spectra of the HCDDs (Figure 2) is that each fragmentation pattern is unique and that we now have another criteria in addition to chromatography to aid in isomer identification. A study of response vs. amount injected for 1,2,3,6,7,8HCDD showed response to be linear over a wide range of concentrations (10 pg to 10 ng) and the relative abundances
+
1136
,
ANALYTICAL CHEMISTRY, VOL. 57, NO. 6, MAY 1985
35:
1
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Flgure 5. MNCI mass spectra of all HCDD isomers.
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i
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ANALYTICAL CHEMISTRY, VOL. 57, NO. 6, MAY 1985 100.0
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124679 OR 124689 HCDD
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Figure 7. Mass chromatogram of m l z 210 and 246 of a pentachlorophenol extract on CPS-1.
1
WE
150
124679 OR 124585 HCDD
200
250
300
350
40e
Flgure 6. ONCI spectra of 1,2,4,6,8,9- and 1,2,4,6,7,9-HCDD.
of the fragment ions were also quite reproducible. A detection limit of approximately 10 pg of 1,2,3,6,7,8-HCDDwas found when monitoring three ions in the multiple ion detection mode. Figure 4 shows the ONCI spectra of 1,2,3,6,7,8-HCDDa t two different source temperature. This shows the relative abundances of fragment ions to be quite sensitive to temperature and that this parameter must be carefully controlled to obtain reproducible spectra. In the case of the Finnigan 4500 GC/MS it is necessary to use the same GC temperature program and interface temperature as well as source temperature to obtain reproducible spectra. The relative abundance of the M- ion increased with increasing temperature but the total ionization yield decreased. When the temperature was lowered to 100 "C to maximize total ion yield, the chromatographic resolution was decreased due to peak broadening. Methane Negative Chemical Ionization. The MNCI spectra of all ten HCDDs are given in Figure 5. The most intense clusters in all cases are due to M-, (M - Cl)-, and (M - 2C1)-, and although each spectra is unique, there is no information about the number of chlorine atoms on each ring. Table I gives the relative sensitivities obtained for all ions from m/z 100 to 500 for 10 ng each of a variety of chlorinated dioxins using ONCI, EI, and MNCI. The electron impact sensitivity for each isomer is given as unity to demonstrate the relative sensitivities of the other ionization techniques. ONCI is more sensitive than E1 for PCDDs containing five or more chlorine atoms and gives the most structural information. MNCI gives the greatest sensitivity of any technique for the higher chlorinated dioxins, but less information on isomers than ONCI. The use of oxygen as a reagent gas was also found to be very effective in keeping the source clean and thus contributed to a higher sample throughout. Thus it was concluded that ONCI was the best method to use as it combined maximum structural information with good sensitivity. Chromatography. A variety of capillary GC columns were tried in this study and the best separation was obtained on a 50-m CPS-1 column as shown in Figure 3. Table I1 shows the relative retention times of the 10 HCDD isomers on the CPS-1 column. However, this did not achieve base line sep-
aration for all ten isomers and indeed there was no separation for 1,2,4,6,7,9- and 1,2,4,6,8,9-HcDDson any GC column used. 1,2,3,6,7,9and 1,2,3,6,8,9isomers were slightly separated on CPS-1 and to a lesaer extent on CPS-2, but this pair appeared as one peak on all the other columns that were tried. In order to separate the 1,2,4,6,8,9/1,2,4,6,7,9 isomers, it was necessary to use normal-phase absorption (silica) HPLC as reported by Lamparski and Nestrick (17). Figure 6 shows the ONCI spectra obtained from the two HPLC fractions. The spectra are similar but not identical, and comparison with the 1,2,4,6,8,9/1,2,4,6,7,9composite spectra in Figure 2 shows it to be intermediate between the two pure isomers; especially the ratio of the mlz 355 and 390 clusters. Figure 7 is a chromatogram of a pentachlorophenol extract cleaned up by HPLC. In this case it is apparent that all ten hexa isomers are present in the sample including small amounts (see maximum intensities Figure 7) of the 4 2 substituted isomers even though they are only slightly resolved on GC from the 3 3 substituted isomers. If any other ionization mode including E.I. were used, then the 4:2 substituted isomers would probably not be detected. A data system library search on the first HCDD &C peak of another PCP extract confirmed the presence of both 1,2,4,6,7,9- and 1,2,4,6,8,9-hexachlorodioxins in the sample. These results demonstrate that the combination of ONCI, HRGC, and HPLC permits the isomer specific separation of all 10 HCDDs, even those in small quantities. A study of the isomer distribution of these compounds in products containing PCDDs may help in reducing the amounts of the more toxic components and could be expanded to dioxin cogeners in other products.
Registry NO. 1,2,4,6,7,9-HCDD,39227-62-8; 1,2,4,6,8,9-HCDD, 58802-09-8; 1,2,3,4,6,8-HCDD,58200-67-2; 1,2,3,6,7,9-HCDD, 64461-98-9; 1,2,3,6,8,9-HCDD,58200-69-4; 1,2,3,4,7,8-HCDD, 39227-28-6; 1,2,3,6,7,8-HCDD,57653-85-7; 1,2,3,4,6,9-HCDD, 58200-68-3; l,2,3,7,8,9-HCDD919408-74-3; 1,2,3,4,6,7-HCDD, 58200-66-1; pentachlorophenol, 87-86-5.
LITERATURE CITED (1) Firestone, D. EHf, Envkon. Health Perspect. 1973, 5 , 59. (2) Crummett, W. 6.; Stehl, R. H. EHP, Environ. Health Perspect. 1973, 5 , 15. (3) Vlllanueva, E. C.; Jennlngs, R. W.; Burse, V. W.; Kimbrough, R. D. J. Agrrc. FoodChem. 1975, 23, 1089. (4) Buser, H. J . Chromatogr. 1975, 707, 295. (5) Buser, H. J . Chromatogr. 1975, 774, 95. (6) Blaser, W. W.; Bredeweg, R. A.; Shadoff, L. A.; Stehl, R. H. Anal. Chem. 1976, 48, 984. (7) Buser, H. Anal. Chem. 1976, 48, 1553. (8) Pfeiffer, C. D. J. Chromatogr. Scl. 1978, 74, 3813. Kaley, R. G.; Michael, P. R. J . Chromatogr. (9) Mieare, J. P.; Hlcks, 0.; Scl. 1977. 15. 275. (10) ituser, H.;'Rappe, C. Anal. Chem. 1984, 5 6 , 442. (1 1) Hlgglnbotham; et al. Nature (London) 1968, 220, 702. (12) Crummet, W. B. Chemosphere 1963, 72, 429. f13) Polychlorinated Dlberno-pdloxins: Limitations to the current analytical techniques, 1981, Publication NRCC No. 18576, Ottawa, Canada.
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Anal. Chem. 1985, 57, 1138-1144
(14) Hass, J. R.; Friesen, M. D.; Hoffman, M. K. Ofg. Mass Spectrom. 1979, 14, 9. (15) Mahle, N. H.; Shadoff, L. A. Blomed. Mass Spectrom. 1982, 9 , 45. (le) Hunt, D. F.; Harvey, T. M.; Russell, J. w. J . them. sot., Chem. Commun. 1975, 151.
(17) Lamparskl, L. L.; Nestrlck, T. J. Chemosphere 1981, 10, 3 . RECEIVEDfor review September 19,1984. Accepted December
17, 1984.
Assessment of Adsorption/Solvent Extraction with Polyurethane Foam and Adsorption/Thermal Desorption with Tenax-GC for the Collection and Analysis of Ambient Organic Vapors Mary P. Ligocki and James F. Paekow* Department of Chemical, Biological, and Environmental Sciences, Oregon Graduate Center, 19600 N. W. Von Neumann Dr., Beaverton, Oregon 97006
Two methods for the collection of ambient organic vapors at the ng/m3 to pg/m3 level were utllired In field sampling at a residential site in Portland, OR, during the winter and spring of 1984. The methods were adsorptlon/solvent extraction with polyurethane foam plugs (ASE/PUFP) and adsorption/ thermal desorption wlth Tenax-GC cartrldges (ATD/TenaxGC). ASE/PUFP was used with a single sample flow rate In a single channel of the sampler. ATD/Tenax-GC was used with two different sample flow rates In two separate channels. Each method was found to be well sulted to the analysis of compounds In a speclflc range of volatility. Some Intermediate-volatility compounds were determined with all three sampling channels. The coefficients of varlation for the three channels pooled over seven events were 9-38% for compounds In the range of volatility between acenaphthene and pyrene. The low sample volumes used wlth ATD/Tenax-GC for determinations at the ng/m3 level make It an attractive method for many applications.
Over the past 2 decades, increasing numbers of trace organic compounds have been identified in the atmosphere. These include the polycyclic aromatic hydrocarbons (PAHs) and their derivatives, pesticides, and polychlorinated biphenyls (PCBs), many Qf which are of concern due to their toxic and carcinogenic properties. In most cases, the distribution and fate of these compounds are only poorly understood. Since knowledge of the physical state of an atmospheric contaminant is vital for the understanding of its operative physical and chemical removal processes, and since the vapor pressures of many trace organics, including the PAHs, pesticides, and PCBs, fall in the range where substantial amounts of a given compound are expected in both the vapor and aerosol phases, a comprehensive organic air sampling system must include both aerosol and vapor measurement capabilities. A popular sampler configuration for the determination of atmospheric organic compounds includes a glass fiber filter followed by an adsorbent such as polyurethane foam (PUF) or Tenax-GC (1-3). PUF has the advantages of being convenient to handle and inexpensive but exhibits breakthrough of volatile compounds ( 4 ) . Although Tenax-GC exhibits less breakthrough, small cartridges have a high flow resistance and large adsorbent beds may be prohibitively expensive for many
Air Intake 1 0 2 mm g l a s s holder
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H o l d e r f o r PUFPs (atalnless s t e e l )
Polyurethan
foam plugs (PUFPs)
a n d pump ( 6 0 0 mL1mln)
Figure 1. Air sampler designed for the collection of ambient particulate and vapor phase organic compounds.
applications. In addition, Tenax-GC may degrade during sampling in the presence of reactive gases such as O3 and NOz (5). Billings and Bidleman (4) compared PUF and Tenax-GC in high volume field sampling for PCBs. Both adsorbents retained the less volatile PCBs adequately, but some breakthrough was observed on PUF for the more volatile PCBs. Solvent extraction was used to recover the analytRs from both types of adsorbents in that study. In conjunction with a sampling program to determine the scavenging of atmospheric organics by rain (6),an air sampler was developed which has the capability to collect organic compounds ranging in volatility from trichloroethene to coronene, in both the vapor and aerosol phases. It utilizes adsorption/solvent extraction (ASE) with PUF for the determination of low volatility organics, with analysis by capillary gas chromatography/mass spectrometry (GC/MS). Adsorption/thermal desorption (ATD) with Tenax-GC is used with
0003-2700/85/0357-1138$01.50/00 1985 American Chemical Society
