Anal. Chem. 2001, 73, 1374-1376
Technical Notes
Methylene Retention Indexes for Isolated Toxaphene Congeners Jeffrey G. McDonald,† Walter Vetter,‡ and Ronald A. Hites*,†
Environmental Science Research Center, School of Public and Environmental Affairs and Department of Chemistry, Indiana University, Bloomington, Indiana, 47405, and Department of Food Chemistry, University of Jena, D-07743 Jena, Germany
Toxaphene was a heavily used, broad-spectrum insecticide, which was banned in most countries in the 1980s. Early data suggested that a limited number of congeners in the technical mixture were responsible for its toxicity to insects. However, toxaphene research has historically focused on analyzing total toxaphene, largely due to insufficient analytical methodology to measure the individual congeners. In recent years, congener-specific toxaphene research has flourished due to analytical advances leading to the identification of several congeners, about 25 of which are commercially available. However, the high price of these standards may inhibit toxaphene research in some laboratories. We report here the methylene retention indexes for 28 isolated toxaphene congeners. When used in conjunction with mass spectrometry, methylene retention indexes provide an alternative method for identifying these compounds when direct comparison with standard compounds is not practical. Toxaphene was a broad-spectrum insecticide consisting of a complex mixture of chlorinated bornanes and camphenes. The Hercules Co. first produced it in 1947 and then patented the process in 1951.1 Toxaphene was produced by the photochlorination of camphene, an isomerization product of R-pinene, which was extracted from softwood tree stumps.2 Toxaphene has 6-10 chlorines/molecule and can contain over 32 000 possible congeners,3 although the technical mixture contains only about 700 compounds.4 Of these, only a limited number of congeners have been individually isolated and characterized. Throughout the 1960s and 1970s, the use of toxaphene increased dramatically as it replaced DDT, which was under heavy scrutiny due to increasing concern about its toxic effects on wildlife.5 In 1986, the U.S. Environmental Protection Agency * Corresponding author:
[email protected]. † Indiana University. ‡ University of Jena. (1) Buntin, G. A. U.S. Patent 2 565 471, 1951. (2) Saleh, M. A. In Toxaphene: Chemistry, Biochemistry, Toxicity and Environmental Fate; Ware, G., Ed.; Reviews of Environmental Contamination and Toxicology; Springer-Verlag: New York, 1991; pp 1-85. (3) Vetter, W.; Luckas, B. Sci. Total Environ. 1995, 160-161, 505-510. (4) Jansson, B.; Wideqvist, U. Int. J. Environ. Anal. Chem. 1983, 13, 309321. (5) Howdeshell, M. J.; Hites, R. A. Environ. Sci. Technol 1996, 30, 220-224.
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banned toxaphene’s use in the U. S., citing similar concerns over its toxicity and persistence in the environment.6 Many other countries followed suit, and today, virtually all developed countries have banned toxaphene’s use; however, it still may be produced and used in underdeveloped countries such as China, Pakistan, and Mexico.7 Because of its toxicity, persistence, and heavy use, toxaphene is one of 12 chlorinated compounds designated for “international action” by the United Nations Environmental Program.8 Until recently, toxaphene measurements were based on “total toxaphene”. This simplification was due to the complexity of the technical mixture and the resulting difficulties in making the analytical measurements. These difficulties stifled attempts to conduct congener-specific research on toxaphene.9 However, as early as 1978, research showed that several pure congeners in technical toxaphene were 4 times more toxic to insects than the technical mixture. This research demonstrated the need to pursue toxaphene research on a congener-specific basis.10 Such research has now become possible through advanced analytical methodology, which has allowed the isolation and structural elucidation of specific congeners. It is now possible to study the unique behavior and accumulation of specific toxaphene congeners in sediments and biota and in other environmental media and to measure the enantiomeric ratios of biologically relevant congeners.9 The commercial availability of individual toxaphene congeners has increased in recent years. In the early 1990s, Parlar and coworkers isolated and identified 23 congeners.11,12 Other researchers have isolated and identified other congeners of technical toxaphene as well.13-16 However, most of these congeners are (6) (7) (8) (9)
(10) (11) (12) (13) (14)
Fed. Regist. 1982, 47, 53784. Voldner, E. C.; Li, Y. F. Chemosphere 1993, 27, 2073-2078. http://worldwildlife.org/toxics/progareas/pop/priority.htm. Vetter, W.; Oehme, M. In New Types of Persistent Halogenated Compounds; Paasivirta, J., Ed.; Toxaphene. Analysis and Environmental Fate of Congeners; Springer: New York, 2000; pp 237-287. Chandurkar, P. S.; Matsumura, F.; Ikeda, T. Chemosphere 1978, 7, 123130. Burhenne, J.; Hainzl, D.; Xu, L.; Vieth, B.; Alder, L.; Parlar, H. Fresenius J. Anal. Chem. 1993, 346, 779-785. Hainzl, D.; Burhenne, J.; Barlas, H.; Parlar, H. Fresenius J. Anal. Chem. 1995, 351, 271-285. Vetter, W.; Klobes, U.; Krock, B.; Luckas, B.; Glotz, D.; Scherer, G. Environ. Sci. Technol. 1997, 31, 3023-3028. Krock, B.; Vetter, W.; Luckas, B.; Scherer, G. Chemosphere 1996, 33, 10051019. 10.1021/ac001119t CCC: $20.00
© 2001 American Chemical Society Published on Web 02/03/2001
Table 1. Measured Methylene Retention Indexes (on a DB-5MS Column) for the Parlar 22 Standard and Other Isolates of Technical Toxaphenea IUPAC name 2-endo,3-exo,5-endo,6-exo,8,8,10-heptachlorobornane 2-exo,3-endo,6-exo,8b,9c,10a-hexachlorobornane 5,5,6-exo-8,9,10-hexachlorocamphene 5-exo,6-endo,8,9,9,10-hexachlorocamphene 2-exo,3-endo,5-exo,9,9,10,10-heptachlorobornane 5-exo,6-endo,7-anti,8,9,10-hexachlorocamphene 2-endo,3-exo,5-endo,6-exo,8b,9c,10a-heptachlorobornane 2,2,5,5,9c,10a,10b-heptachlorobornane 5,5,6-exo,8,9,9,10-heptachlorocamphene 2-endo,3-exo,5-endo,6-exo,8b,8c,10a,10c-octachlorobornane 2-endo,3-exo,5-endo,6-exo,8,8,9,10-octachlorobornane 5,5,6-exo,8,8,9,9,10-octachlorocamphene 2,2,5-endo,6-exo,8c,9b,10a-heptachlorobornane 2,2,5,5,9b,9c,10a,10b-octachlorobornane 2,2,3-exo,5-endo,6-exo,8c,9b,10a-octachlorobornane 2-endo,3-exo,5-endo,6-exo,8b,9c,10a,10c-octachlorobornane 2-exo,3-endo,5-exo,8c,9b,9c,10a,10b-octachlorobornane 2,2,5-endo,6-exo,8b,8c,9c,10a-octachlorobornane 2,2,5-endo,6-exo,8c,9b,9c,10a-octachlorobornane 2-exo,5,5,8c,9b,9c,10a,10b-octachlorobornane 2-endo,3-exo,5-endo,6-exo,8b,8c,9c,10a,10c-nonachlorobornane 2,2,5,5,8c,9b,10a,10b-octachlorobornane 2,2,5-endo,6-exo,8b,8c,9c,10a,10c-nonachlorobornane 2,2,3-exo,5,5,8c,9b,10a,10b-nonachlorobornane 2,2,5-endo,6-exo,8c,9b,9c,10a,10b-nonachlorobornane 2,2,5,5,8c,9b,9c,10a,10b-nonachlorobornane 2-exo,3-endo,5-exo,6-exo,8b,8c,9c,10a,10c-nonachlorobornane 2,2,5,5,6-exo,8c,9b,9c,10a,10b-decachlorobornane
Parlar no.
A/V no.
Cl no.
measured RI
B7-1000 B6-923
7 6 6 6 7 6 7 7 7 8 8 8 7 8 8 8 8 8 8 8 9 8 9 9 9 9 9 10
2072.6 2088.3 2105.8 2110.0 2147.5 2162.7 2190.2 2200.4 2225.4 2238.8 2258.4 2280.3 2297.2 2319.9 2352.1 2357.6 2357.6 2373.2 2373.2 2382.2 2415.1 2423.8 2483.9 2498.2 2505.8 2539.4 2543.8 2685.0
11 12 B7-1453 15 21 25 26 31 32 38 39 40 41 42a 42b 44 50 51 56 58 59 62 63 69
B7-1001 B7-499 B8-1413 B8-1412 B7-515 B8-789 B8-531 B8-1414 B8-1945 B8-806 B8-809 B8-2229 B9-1679 B8-786 B9-1046 B9-715 B9-1049 B9-1025 B9-2206 B10-1110
a Congeners are listed by their IUPAC name and then by Parlar number and A/V number where applicable. The errors for the retention indexes were all between 0.02 and 0.03 units, less than the precision of the reported retention indexes.
available only in limited quantities or are not commercially available at all. The complex separations involved in the isolation and identification of these congeners makes their price and availability prohibitive to many laboratories, potentially hindering the advance of congener-specific toxaphene analyses. As an alternative to identifying specific congeners by direct comparison with standards, we present here the methylene retention indexes for 28 congeners found in technical toxaphene. EXPERIMENTAL SECTION Toxaphene Nomenclature. The nomenclature of toxaphene congeners has not been uniform, and attempts have been made to reduce the complex IUPAC nomenclature; this has led to the Parlar numbering system, the Andrews and Vetter (A/V) system, and others. We will primarily use the Parlar and A/V nomenclatures because they are the most widely used and accepted. Several of the compounds we discuss here are chlorocamphenes, for which A/V nomenclature does not apply. Vetter and Oehme have published a complete review of toxaphene nomenclature.9 Origin of Isolates. We obtained four mixtures, comprising a total of 28 isolated congeners of technical toxaphene. The first was the commercially available Parlar 22 standard (Dr. Ehrenstorfer GmbH, Augsburg, Germany), consisting of 23 congeners (a pair of congeners coelute). The second mixture contained two congeners, B7-1453 and B8-1412. The third contained B7-1000. The fourth mixture contained two compounds, B6-923 and B7(15) Miskimmin, B. M.; Muir, D. C. G.; Schindler, D. W.; Stern, G. A.; Grift, N. P. Environ. Sci. Technol. 1995, 29, 2490-2495. (16) Coelhan, M.; Parlar, H. Chemosphere 1996, 32, 217-228.
1001, commonly referred to as Hx-Sed and Hp-Sed, respectively.15 The second, third, and fourth standards are not commercially available at this time. For assigning retention indexes, we used a standard mixture containing 17 n-alkanes, ranging from n-octane (C8H10) to n-tetracontane (C40H82) with only the even carbon numbers (AccuStandard, New Haven, CT). Instrumental Analysis. Toxaphene isolates and n-alkenes were analyzed using a Hewlett-Packard gas chromatographic (6890) mass spectrometer (5973) operated in electron capture, negative ionization (ECNI) and positive chemical ionization (PCI) modes. All toxaphene congeners reported here were ionized using ECNI conditions, while the n-alkenes were ionized using PCI conditions. The n-alkanes were ionized using PCI conditions so the individual peaks could be assigned by using the (M - H)+ ions in the respective mass spectra. Using PCI and ECNI modes allowed us to analyze both the toxaphene congeners and the n-alkanes without having to make modifications to the instrument, which would have increased the uncertainty in the measurements. To be sure that there was no systematic offset in retention times between the PCI and ECNI ionization modes, we checked the retention times of methoxychlor olefin and p,p′-DDT, which ionize well in both modes. These retention times agreed within (0.005 min, which gives errors in the retention indexes that are less than the errors suggested below. The GC/MS instrument was equipped with a 60-m DB-5MS column (250-µm i.d.; 0.25-µm film thickness; J&W Scientific, Folsom, CA). The injection port was maintained at 285 °C; 2 µL was manually injected in the pulsed injection, splitless mode at Analytical Chemistry, Vol. 73, No. 6, March 15, 2001
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25 psi with 1.9-min vent time. Helium was the carrier gas at a head pressure of 17.2 psi and average velocity of 25 cm/s. The temperature program began at 100 °C for 1 min and was then ramped at 20 °C/min to 210 °C, 0.8 °C/min to 270 °C, and 25 °C/min to 325 °C, and held for 10 min at 325 °C for a total run time of 93.7 min. The transfer line between the GC and the MS was heated to 280 °C. The ion source was operated at 150 (ECNI) and 250 °C (PCI), and the quadrupoles were operated at 106 °C in both modes. Methane was used as the reagent gas at a manifold pressure of 2 × 10-4 (ECNI) and 4 × 10-4 Torr (PCI). Retention times for all relative peaks were assigned using Hewlett-Packard ChemStation software. Because of the stepped temperature program, the retention time spacing of the n-alkanes was not linear. We determined that the best-fit functional relationship between the retention index scale and the GC retention times of the n-alkanes was logarithmic. Thus, the retention indexes were calculated from a logarithmic interpolation between the two n-alkanes that bracketed the retention time of each respective congener. For the congeners measured here, the n-alkanes used were C20H42, C22H46, C24H50, C26H54, and C28H58. RESULTS AND DISCUSSION The retention indexes for the 28 toxaphene congeners we measured are given in Table 1. Congeners are listed by their IUPAC names and then by Parlar or A/V numbers where applicable. Figure 1 shows the bornane and camphene structures with numbered carbons for use as an aid in interpreting the IUPAC nomenclature. The standard errors for the measured retention indexes were calculated based on triplicate measurements. Errors were calculated using basic principles of error propagation, including quadratic sums of relative and absolute errors and log transformations where necessary. Due to the exceptional reproducibility of modern gas chromatographs, these errors ranged between (0.02 and (0.03 retention index units, less than the precision with which we report the retention indexes themselves. We suggest an error
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Figure 1. Structure of bornane (A) and camphene (B) with carbons numbering based on IUPAC nomenclature. Potential chlorine positions on the primary carbons of structure A are also labeled.
of (0.1 retention index units as a more liberal value to account for potential differences in gas chromatographs and column manufacturers. When these retention indexes are used to identify potential congeners of technical toxaphene, it is important to employ mass spectrometry for positive identification of the congeners. Isotopic ratios resulting from the two isotopes of chlorine can provide conformation that the potential congener has the proper degree of chlorination. Purity of the peak can also be confirmed by mass spectrometry, allowing for the determination of potential coelution or other interferences. Other detectors such as electron capture GC detectors do not provide this capability.
Received for review September 19, 2000. Accepted December 13, 2000. AC001119T
