Environ. Sci. Technol. 1993, 27, 1826-1831
Chlorinated Polycyclic Aromatic Hydrocarbons: Method of Analysis and Their Occurrence in Urban Air Ulrlka L. Nilsson' and Conny E. Ostman Analytical Chemistry Division, National Institute of Occupational Health, S-17 1 84 Solna, Sweden
A method for sensitive and selective analysis of chlorinated polycyclic aromatic hydrocarbons (Cl-PAHs) is described. Two-dimensional liquid chromatography was applied for the isolation of a fraction containing C1-PAHs and polycyclic aromatic hydrocarbons (PAHs). Separation, selective detection, and identification of C1-PAHs were performed by the use of GC-MS utilizing chemical ionization, selected-ion monitoring, and negative-ion detection. Samples from an urban area were investigated with respect to the occurrence, identity, and concentration of C1-PAHs. Among the identified compounds were 1and 4-chloropyrene and 6-chlorochrysene. The former two exhibit strong mutagenic effects, and the later binds to the TCDD receptor and is an AHH-inducing agent. The level of 1-chloropyrene was found to be about 10 pg/m3in the street and about 40 pg/m3 in the road tunnel. The C1-PAHsfound in urban air were in agreement with those predicted from laboratory studies of reactivity and formation of chloro-substituted PAHs using gaseous chlorine. The presence of some specific isomers of chloro-substituted PAHs indicated degradation products of chloro-added PAHs. Introduction Chlorinated polycyclic aromatic hydrocarbons (C1PAHs) are a group of compounds comprising polycyclic aromatic hydrocarbons with three or more rings and one or more chlorine atoms attached to the ring system. The C1-PAHscan be divided in two groups: chloro-substituted PAHs, which have one or more hydrogen atoms substituted by chlorine, and chloro-added Cl-PAHs, which have two or more chlorine atoms added to the molecule and, thus, loosing the aromaticity in the ring to which the chlorine atoms are attached. During recent years, 61-PAHs have been detected in environmental samples of various origin. Shiraishi et al. demonstrated their presence in chlorinated tap waters (1). Tausch and Stehlik detected chlorosubstituted PAHs, ranging from four to seven aromatic rings, in fly ash from an incineration plant for radioactive waste (2). In this case, the concentrations were reported to be in the same range as those of the parent PAHs. Polychlorinated PAHs were detected by Eklund and Stromberg (3) and by Oehme et al. ( 4 ) in emissions from coal combustion and municipal waste incineration. Also chlorobromo derivatives of PAHs were shown to occur in fly ash from a municipal waste incinerator, according to Sovocool et al. (5). Formation of polychlorinated pyrenes during PCB fires has been reported by Rappe et ala (6). Moreover, Haglund et al. (7) reported the occurrence of a large number of chlorinated PAHs in automobile exhausts, snow, and urban air. Regarding automobile exhaust, the major source of chlorine was thought to be dichloroethane, which is added to leaded fuels. Interest has been focused on C1-PAHs since a number of these compounds have shown a response in test systems 1826
Envircn Sci. Technol , Vol. 27, No 9, 1993
for biological activity. Several chlorinated derivatives of PAHs exhibit mutagenic activity toward Salmonella typhimurium in the Ames assay. It was found by Colmsjij et al. (8,9) and Rannug et al. (10) that some chloro-added PAH derivatives, such as chloro-added pyrenes and chloroadded benzo[elpyrenes, had different mutagenic properties compared to the corresponding chloro-substituted PAHs. The chloro-added derivatives exhibited direct mutagenic effects, while the chloro-substituted derivatives showed significant mutagenic effects only in the presence of a metabolizing system. It was also demonstrated that some chloro-added derivatives of perylene exhibit extremely strong direct mutagenic effects (10). Further, Lofroth et al. found that 7-chlorobenz[alanthracene, 9-chloroanthracene, and 6-chlorochrysene showed strong direct mutagenic effects (11). The latter compound has also been shown to have a high affinity to the TCDD receptor (12) and to be a potent AHH inducer (13). The aim of this work was to identify and determine the levels of possible biologically active chlorinated PAHs in urban air, taking both the particulate and the semivolatile phases into consideration. Isolation of a C1-PAH/PAH fraction was performed with a two-dimensional HPLC system and using selective analysis with gas chromatographylmass spectrometry utilizing chemical ionization in negative-ion detection mode (GC/MS-NICl). Experimental Section
Chemicals and Solvents. All of the Cl-PAH reference substances, except 1-chloropyrene(9596 purity, Cambridge Chemicals), were synthesized by the authors (14-1 7). The identity of the compounds were established by utilizing nuclear magnetic resonance spectroscopy (270 MHa, JMMEX270, Jeol Japan). Capillary gas chromatography (GC) was used for control of the purity. Standard solutions of C1-PAHs used for quantification were prepared using cyclohexane as the solvent. The silica gel used for preseparation of the samples was prepared from 0.063 to 0.2 mm particle-size Kieselgel 60 (Merck, Darmstadt, Germany), which was heated to 450 "C for 24 h. Subsequent deactivation was done by adding 10% (w/w) distilled water. The deactivated silica was stored in cyclohexane until used. Analytical-grade solvents obtained from Merck were used after distillation in an all-glass apparatus. The HPLC mobile phase was HPLC-grade hexane (Rathburn Inc., Walkerburn, Scotland), which was used as received. Sample Collection. An anodized aluminum sampler equipped with a glass fiber filter (127-mn1, type AIE, Gelman Sciences Inc., Ann Arbor, MI) and two polyurethane foam plugs (PUF) (density = 25 kg/m3,70 rnm i.d. X 70 mm, Specialplast, Gillinge, Sweden) coupled in series were used for the sampling. Particles were collected on the glass fiber filter, and the semivolatile phase was adsorbed on the first PUF. The second PlJF was used as a control of break-through in the sampler. A high-volume sampler pump gave a flow of 13.6 m3/h. Samples were 0013-936X/93/0927-1826$04 00/5
0 1993 American Chemical Soclety
collected during 24 h, yielding sampled air volumes of 325 m3. Prior to sampling, the glass fiber filter was washed with methanol and acetone, and the PUFs were treated with boiling water for 2 h, washed with acetone, and Soxhlet-extracted with 300 cycles of benzene. When investigating the sampling system with regard to the formation of artifacts during sampling, the glass fiber filter was replaced by a Teflon filter (Pallflex T60A20, Pallflex Inc.). Six air samples from an urban street environment with heavy traffic (29 000vehicleslday) and two air samples from a road tunnel (1.5 km long, 19 000 vehicleslday) were collected. After Soxhlet extraction, the street air samples were pooled into two samples representing two subsequent weeks. Sample Cleanup. Soxhlet extraction was used to desorb the organic compounds collected on the filter and the adsorbed organic material on the PUFs. The internal standard (50 pg of monochloro-2,2'-binaphthylper m3 of air) was adsorbed on the filter and injected into the PUFs. Extraction was then performed by using 200 cycles of benzene during 24 h. Benzene was used instead of dichloromethane in order to avoid chlorine during the evaluation of the sample treatment. The two solvents exhibit comparable recoveries of the investigated C1-PAHs, and as is subsequently shown, the use of dichloromethane did not induce any artifact formation during Soxhlet extraction. It is thus possible to substitute benzene for dichloromethane. This is to be preferred due to the high toxicity of benzene. A total of 20 pL of dodecane was added to the extracts before the solvent was removed. The extracts were dissolved in 200 pL of cyclohexane and were then preseparated on an all-glass column (i.d. = 6 mm) with deactivated silica to a height of 60 mm. A single fraction containing aliphatics, mono- and dicyclic aromatics, PAHs, and C1-PAHs was eluted with 18 mL of cyclohexane and concentrated to 100 pL. After silica cleanup, an HPLC back-flush method with a cyanopropyldimethylsilica column in normal-phase mode was used, in order to isolate possible chloro-added derivatives of PAHs (17). Chloro-added PAHs could be collected in a back-flush fraction, while chloro-substituted PAHs, parent PAHs, aliphatics, and mono- and dicyclic aromatics eluted before the flow reversal, Le., in the straight-flush fraction. This fraction was collected and further cleaned up with respect to chloro-substituted PAHs. The C1-PAHs were then isolated together with the PAHs by using a two-dimensional HPLC coupled column system described elsewhere (18, 19). Aliphatic and mono- and dicyclic aromatic compounds were eluted from the column system. Aromatic compounds with three or more rings were subsequently back-flushed out of the column system as one peak. By coupling a 2-(l-pyrenyl)ethyldimethylsilica column in series with a nitrophenylpropyldimethylsilica column, the C1-PAHs with three or more aromatic rings were obtained within the PAH fraction. The overall absolute recoveries of individual chloro-substituted C1-PAH components with three to five aromatic rings were found to be in the range of 90-95%. By the use of an internal standard technique, a relative recovery of 100% was obtained with a coefficient of variation of 10%. Instrumental. A Waters 590 solvent delivery module (Waters Associates, Milford, MA) was used to deliver a flow of 1.0 mLlmin of hexane (HPLC grade, Rathburn Inc., Walkerburn, Scotland) through the coupled column
system. The columns utilized in the cleanup procedure were as follows: a cyanopropyldimethylsilyl column (250 X 4.6 mm, Hibar, 5 pm, Merck, Darmstadt, Germany) and a nitrophenylpropyldimethylsilyl column (70 X 4.0 mm, Nucleosil-NOz, 5 pm, Macherey-Nagel, Duren, Germany) coupled in series with a 2-(l-pyrenyl)ethyl-modifiedsilica column (150 X 4.6 mm, Cosmosil PYE, 5 pm, Nakarai Chemicals, Ltd., Kyoto, Japan). A Rheodyne injector (Model 7125, Rheodyne, Cotati, CA) equipped with a 100pL loop was used as the injection system. Detection was performed using a UV detector (L-4000, Merck-Hitachi, Germany) tuned at 254 nm. Back-flushing the column system and collecting the back-flush peak was performed by utilizing a 4- and a 3-port switching valve, respectively (Valco Instruments Co. Inc., Houston, TX). Each valve was equipped with a two-position helical-drive air actuator operated by a digitalvalve interface. A PC microcomputer, using a relay actuator card, controlled the valve switching. Operation of the valves and registering the UV-detector response was performed by an ELDS 900 PC computerbased laboratory data system (Chromatography Data Systems AB, Stenhamra, Sweden). The GC-MS system consisted of a Varian 3400 gas chromatograph (Varian, Walnut Creek, CA) coupled to an Incos 50 quadrupole mass spectrometer (Finnigan MAT, San Jose, CAI. The GC was equipped with a split1 splitless injector operating at a temperature of 310 "C. The capillary column was a DB-5 (5% phenylmethylsiloxane gum, 30 m X 0.25 mm i.d., df = 0.10 pm, J&W Scientific),and helium was used a carrier gas. During the splitless injection, with a duration of 2 min, the column temperature was kept at 70 "C. It was then increased linearly a t a rate of l0"Imin up to 300 OC which was kept for 10 min. A temperature of 310 "C was maintained in the GC-MStransfer line. The MS was running in negativeion chemical ionization (NIC1) mode, using methane of 99.995% purity as a reactant gas at an ion source pressure of 1.3 mbar. The chloro-substituted PAHs are stable, and the mass spectra exhibit dominating molecular ions when using electron impact ionization as well as chemical ionization. Two sequence scan descriptors for selectedion monitoring (SIM) were used for selective detection of monochloro- and dichlorosubstituted PAHs with parent molecular weights of 178, 202, 228, 252, and 276. Monochloro derivatives of the different parent PAHs were monitored using the molecular ions 2121214,2361238,262/ 264, and 2861288. The isotopic ratio between the two molecular ions was used as an identification of a monochloro derivative. Dichloro derivatives were monitored in a similar maner using the molecular ions 246/248,270/272, 296/298, and 3201322 with the isotopic ratio as an identification of a dichloro derivative. The time invervals for the different masses were calibrated using the retention of standard components. Mass 289 was used for monitoring the internal standard monochloro-2,2'-binaphthyl.
Results and Discussion Response and Identification. The response of the detection system with respect to C1-PAHswas investigated using 1-chloropyrene,7-chlorobenz[alanthracene,and 1,6dichloropyrene. All three compounds showed a linear response in the range of 0.5 to 1000 pg with coefficients of correlation better than 0.9990. For this reason, linear responses in the investigated concentration range were assumed for all the investigated C1-PAHs. The mass Enviton. Sci. Technol., Vol. 27, No. 9,1993
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Table I. Relative Retention and Relative Response Factors for 20 Cl-PAH Reference Compounds Calculated against 1-Chloropyrene.
compound
retention
response
9-chlorophenanthrene 9-chloroanthracene 9,lO-dichlorophenanthrene 9,lO-dichloroanthracene 8-chlorofluoranthene 3-chlorofluoroantheen 4-chloropyrene 1-chloropyrene x,3-dichlorofluoranthene 1,3-dichloropyrene 1,8-dichloropyrene 1,6-dichloropyrene 6-chlorochrysene 7-~hlorobenz[a]anthracene 6,12-dichlorochrysene 7,12-dichlorobenz[al anthracene z-chlorobenzo [e]pyrene 6-chlorobenzo[a]pyrene z-chlorobenzo~hilperylene y-chlorobenzo[ghil perylene
0.846 0.838 0.940 0.940 0.967 0.974 0.999 1.000 1.067 1.082 1.093 1.094 1.140 1.145 1.218 1.219 1.288 1.298 1.442 1.459
13 1900 980 1100 4400 2100 3600 10000 8200 9200 12000 6700 48 4400 2700 980 1200 5100 4000 3300
a
Temperature program, see Experimental Section. Coefficients
of variation, relative retention
