Environ. Scl. Technol. 1982, 16, 247-250
A Novel Integrative Technique for Locating and Monitoring Polynuclear Aromatic Hydrocarbon Discharges to the Aquatic Environment John J. Black,” Thomas F. Hart, Jr., and Penelope J. Black
Roswell Park Memorial Institute, Buffalo, New York 14263
w A simple integrative technique for locating and monitoring polynuclear aromatic hydrocarbon discharges to aquatic environments is described. The technique involves anchoring artificial substrates cut from a commercial oil-adsorbant cloth (3M Co.) near suspected sources of contamination. Analytical methodology involves mild ethanolic extraction and liquid-liquid partitioning to isolate a polynuclear aromatic hydrocarbon containing fraction that is amenable to analysis by high-pressure liquid chromatography.
Introduction A screening method with the capability to detect and locate point sources of hydrocarbon pollutants prior to the implementation of an intensive point-source monitoring program would have considerable utility. We describe here a simple, relatively inexpensive analytical technique that uses polypropylene substrates (PST) for integrative monitoring of polynuclear aromatic hydrocarbons (PAH) in aquatic environments. Experimental Procedures Substrates. Polypropylene substrates (PST)measuring 5.0 cm X 10.0 cm were cut from sheets of a commercially available oil-adsorbant cloth (3M Brand Oil Sorbant Type 151). Substrates were wrapped in foil to avoid contamination prior to use. The bouyant PST’s were attached to an anchored wooden float by a short nylon line and allowed to trail freely in the current. Use of spring-loaded metal clips facilitated rapid substrate installation and removal under field conditions. After exposure periods of 24-96 h, substrates were removed, immediately wrapped in foil, and returned to the laboratory where they were stored at . -10 OC prior to extraction and analysis. Solvents. All hydrocarbon solvents and water were redistilled in glass. Dimethyl sulfoxide (Burdick and Jackson) and acetonitrile (J. T. Baker, HPLC grade) were used without further purification. Sample Extraction and PAH Isolation. PST’s were placed in 43 mm X 123 mm cellulose thimbles (Whatman) and extracted with 95% ethanol in a Soxhlet apparatus for 4 h (approximately16 cycles). The ethanol extract (250 mL) was transferred to a separatory funnel containing 230 mL of water and 250 mL of cyclohexane, and a nonpolar fraction was isolated by liquid-liquid partitioning. After solvent exchanging of the cyclohexane for dimethyl sulfoxide (Me2SO),a PAH-containing fraction was isolated by Me2S0partitioning and back-extraction procedures similar to those utilized by Dunn for isolation of PAH from marige organisms (1). Details are provided here for completeness. As per Dunn, the solvent exchanged Me2S0 was transferred along with a 5-mL Me2S0 rinse of the evaporation flask to a separatory funnel containing 10 mL of hexane. After vigorous shaking, the MezSO hypophase was transferred to a second separatory funnel. The hexane phase in the first funnel was extracted a second time with a fresh 10-mL volume of Me2S0, and this was also added to the 0013-936X/82/0916-0247$01.25/0
second Beparatory funnel. Following addition of 40 mL of water and 20 mL of cyclohexane to the second funnel, the PAH was back-extracted into the cyclohexane. The aqueous Me2S0 phase was drained into a third funnel and extracted again with 20 mL of cyclohexane. The two cyclohexane phases were combined and washed with distilled water. This fraction, containing the bulk of the PAH, was reduced in volume to approximately 3 mL and transferred to a conical tube where the hydrocarbons contained in this fraction were concentrated into an accurately measured volume of Me2S0 (0.5-1 mL) by evaporating off the cyclohexane. The resulting analytical fractions were transferred to septum-covered injection vials (Varian Associates) and stored in the dark prior to analysis. High-pressure Liquid Chromatography (HPLC). Analyses were performed on a Perkin-Elmer Series 3 liquid chromatograph. With a flow rate of 0.6 mL/min, a brief isocratic segment (2.5 min; 55% acetonitri1e:water) was followed by a linear gradient to 100% acetonitrile (25 min). A 20-min hold at 100% acetonitrile was required to elute residuals and return the chromatographic .signals to baseline. The column was equilibrated at the initial solvent composition for 20 min prior to another analytical run. A fixed-wavelength 254-nm detector (Perkin-Elmer LC-15) and an in-house repacked column (2.1 mm i.d. X 250 mm, Vydac TP 201, 10-pm ODS packing material) were used for routine screening analyses. For detailed analysis of some samples, a Perkin-Elmer PAH,, column was connected in series with an absorbance detector (Perkin-Elmer LC-75) and a fluorescence detector (Perkin-Elmer 650-10s). An exciting wavelength of 300 nm and an emission wavelength of 420 nm were utilized for fluorescence detection. Peaks were identified by their retentions relative to an internal reference compound (chrysene) and by trace enrichment. A bypass valve on the fluorescence detector enabled the flow cell to be taken “off-line”for spectral scanning. Fluorescence spectra obtained via this modification are used to confirm compound identifications.
Results and Discussion So that the origin of PAH in aquatic environmentscould be determined, a number of investigations have measured concentrations in sediments, mollusks, crustacea, and fish (2-4). While these approaches, especially measurement of sediment PAH (4-8),have some value for localizing sources of these compounds, they may have limited usefulness in some aquatic situations. For example, sediment analyses may largely reflect historical data and/or sediments may be difficult to obtain at a given site. Measurement of PAH in biota may be complicated by the fact that most biota, especially fish, metabolize and eliminate many compounds of interest such as carcinogenic PAH (4, 9-13). Measurements of PAH in water have employed materials such as XAD resins and urethane foams to concentrate these pollutants prior to analysis (14-1 7). These methods have been largely used as a substitute for liquid-liquid partitioning methods applied to a single water
0 1982 American Chemical Society
Environ. Sci. Technoi., Voi. 16, No. 5, 1982 247
sample, although large volumes of water have been sampled over extended periods in some drinking-water studies (18). These techniques, though highly useful, may be expensive or difficult to apply in some situations. The use of inert adsorbtive substrates to directly monitor waterborne PAH would overcome many of these disadvantages. To be valid for use as a hydrocarbon-monitoring substrate, the material chosen should produce an artifact-free reagent blank, readily adsorb hydrocarbons, provide consistent recoveries of the adsorbed compounds, and be inexpensive and easy to use. In our search for a suitable substrate we initially tested samples of urethane foam. Organic solvent extracts of the foam always produced artifactual peaks that interfered with the HPLC analyses. Alternatively the PST, when extracted by using the analytical methodology described in this report, did not produce significant interference to the analysis of PAH compounds. To develop an extraction method for the isolation of a PAH/PST adsorbate, three solvents of differing polarity were tried. Solvents utilized in these experiments were hexane, methylene chloride, and ethanol. Prolonged extractions (24 h) with any of these solvents resulted in a fraction that formed a gel during subsequent concentration steps. Gel formation was particularly evident with hexane and methylene chloride. The nature of this gel is unknown but may be due to extraction of lubricants used in the manufacture of the polypropylene cloth or perhaps extraction of small amounts of the polymer. By trial and error it was determined that a 4-h ethanolic extraction (14-16 Soxhlet cycles) yielded consistently gel-free procedural blanks that did not contain significant amounts of materials that interfered with the chromatographic analysis. While the rate of adsorption of PAH from water could not be easily measured, as a test of the efficiency of the solvent extraction and partitioning scheme a l-mL hexane solution containing 50 pg each of four PAH compounds was distributed onto a PST. Recoveries of the PAH “spiked” onto the PST varied with ring number and the degree of alkyl substitution. The following extraction efficiencies were measured by using the previously described ethanolic extraction and MezSO partitioning scheme: phenanthrene, 79% ; methylanthracene, 64% ; chrysene, 69%; benzo[ghi]perylene,44%. These efficiencies were relatively low and apparently reflected partitioning behavior between the polar extracting solvent and the nonpolar PST, i.e., recoveries of aJkyl-substituted PAH < parent structure and 3 ring > 4 ring > 5 ring. An environmental simulation involving brief exposure of a substrate to a dilute creosote/water mixture (50 pL of creosote in 3.5 L of water) indicated that PST readily adsorbed PAH compounds from the water (19). To test the PST in a field situation, ten western New York aquatic sites on Lake Erie and the Niagara River were selected as potential sources of PAH. The sites were selected on the basis of information contained in the New York State Industrial Chemical Survey (20) and on the basis of high levels of PAH detected in previous HPLC screening analyses of sediments (19) collected at some of the sites. In general, PST anchored near potential sources of PAH yielded analytical fractions that, when subjected to HPLC analysis, produced chromatogramsindicative of significant amounts of UV-absorbing materials, whereas unexposed PST, i.e., procedural blanks and/or PST exposed in areas of presumably cleaner water, did not exhibit significant levels of UV-absorbing materials. Additionally, PST exposed at a given site for 96 h yielded chromatograms in248
Environ. Sci. Technol., Vol. 16, No. 5, 1982
P
Figure 1. Chromatograms produced by HPLC analysis of PST exposed for 24 and 96 h of UV absorbance at 254 nm. Abbreviations: P, phenanthrene; A, anthracene; FN, fluoranthrene; Py, pyrene: BA, benzanthracene: BaP, benzo[a Ipyrene.
L
uL
BLANK
BLANK
Figure 2. HPLC elution patterns produced by PST exposed for 96 h at sites near suspected sources of hydrocarbon pollution. Reagent blanks are shown for comparison (UV absorbance at 254 nm).
dicative of larger amounts of adsorbed materials than PST exposed for 24 h (Figure 1). However, the increase noted did not appear to be strictly proportional. This either may have indicated variable rates of hydrocarbon output at a given site or may reflect inherent kinetics of the PST methodology. Reproducibility of the technique in field testing appeared to be excellent. Paired duplicates of field-exposed PST produced results that displayed less than 2% variation between any pair of chromatographic peaks. Typical results obtained from PST monitoring of two putative PAH contributing sites are shown in Figure 2. General differences between the character of the chromatographicpatterns observed in analysis of PST fractions from different sites were observed, suggesting that different types of hydrocarbon sources may have been located (see also Figure 3). For routine screening purposes a low-resolution HPLC column and detection& UV absorption at 254 nm appeared adequate. A trace enrichment of a PST adsorbate with use of this mode of detection is shown in comparison with a PST procedural blank (Figure 3). The PST adsorbate shown in Figure 3 was also analyzed on a high-
EMISSION
UNKNOWN
jk, \
BLANK BaP
Flgure 3. Trace enrichment of a PST absorbate with PAH (UV absorbance at 254 nm). Abbreviations: F, fluorene: P, phenanthrene: A, anthracene, Fn, fluoranthene; Py, pyrene 2-methyiphenanthrene: MeA, methylanthracene; BA, benzanthracene: BaP, benzo[a ] pyrene.
+
\ 220 WAVELENGTH
320
420
WAVELENGTH
Flgure 5. Emission and excitation spectra obtained from the putative benzo[a]pyrene peak in the ship canal: PST adsorbate of Figure 4 compared with spectra obtained from authentic benzo[a Ipyrene.
I\
CLEAN WATER CONTROL
BLANK
Flgure 4. Fluorescence HPLC chromatograms, highresolution column (Perkln-Elmer PAH,,), showing results of analysis of PST exposed for 96 h cian-water control vs. suspected site of hydrocarbon pollution (excitation, 300 nm, emission 420 nm). Reagent blank shown for comparison.
resolution column (Perkin-Elmer PAHlo) by using the dual mode of compound detection (see Experimental Procedures). Studies have indicated that this column has a high degree of selectivity for PAH compounds and is capable of resolving the 16-priority pollutant PAH, including isomeric PAH pairs that are not resolved by other columns (21,22). The results of this analysis by fluorescence detection are shown in Figure 4. A PST procedural blank and a clean water control are also shown for comparison. These data also indicated the presence of a series of compounds with retention times consistent with identifications as PAH. Fluorescence spectroscopy was used to confirm the identity of the suspected benzo[a]pyrene peak detected
in this sample. Close matches of emission and excitation spectra recorded from authentic benzo[a]pyrene and the putative benzo[a]pyrene peak indicated that the sample component was in fact benzo[a]pyrene (Figure 5). This particular PST was anchored near a major steel manufacturing industry and clearly recorded the presence of a source of PAH. Observation at this location at the time of PST retrieval indicated the presence of a typical rainbow-colored hydrocarbon slick. In conclusion, these field and laboratory experiments clearly indicated the ability of PST to adsorb PAH from water, and although efficiencies of the extraction method for some PAH compounds were in the low range, the reproducibility of the PST technique was excellent. The low extraction efficiencies would not appear to be a serious handicap providing a standard time interval is applied in sampling. We chose to use a 96-h exposure based on our experience in a particular field situation. Other time intervals may be appropriate in a different environmental setting. Although point-source discharges may be episodic, because the PST technique provides an integrated record of PAH accumulation over the time of field exposure, the use of the technique should facilitate rapid "tracking" of these hydrocarbons to their putative sources. Specific effluents under suspicion could then be subjected to detailed scrutiny to determine actual discharge rates. The technique may be of value to regulatory agencies for locating and/or monitoring point sources of PAH. By modification of the PST analytical procedure it may be possible to use PST to monitor other hydrophobic pollutants. However, it should be noted that the methods of detection employed in the present study are fairly selective for PAH and Environ. Scl. Technoi., Vol. 16, No. 5, 1982 249
Environ. Sci. Technol. 1982, 16, 250-254
therefore other modes of detection may be subject to interference. Literature Cited Dunn, B. P. In “Polynuclear Aromatic Hydrocarbons: Chemistry and Biological Effects”; Bjorseth, Alf, Dennis, A. J., Eds.; Battelle Press: Columbus,OH, 1979; p 367-377. Brown, R. A,; Weiss, F. T. “Fate and Effects of Polynuclear Aromatic Hydrocarbons in the Aquatic Environment”, Publication No. 4297; American Petroleum Institute: Washington, D.C., 1978. Neff, J. M. “Polycyclic Aromatic Hydrocarbons in the Aquatic Environment: Sources, Fates and Biological Effects”;Applied Sciences: London, 1979. Black, J. J.; Hart, T. F.; Evans, E. In “ChemicalAnalysis and Biological Fats: Polynuclear Aromatic Hydrocarbons“; Cooke, M., Dennis, A. J., Eds.; Battelle Press: Columbus, OH, 1980; p 343-355. Black, J. J. Arch. Environ. Contam. Toxicol., in press. Lopez-Avila,V.;Hites, R. A. Environ. Sci. Technol. 1980, 14, 1382-1390.
Hites, R. A.; LaFlamme, R. E.; Windsor,J. G. Environ. Sci. Res. 1980,16, 397-403.
Grimmer, G.; Bohnke, H. Cancer Lett. (Shannon, Irel.)
Neoplasia”; Kraybill, H. F.; Dawe, C. J., Harshbarger, J. C., Tardiff, R. G., Eds.; N.Y. Academy of Sciences: New York, 1977; p 505-521. Pedersen, M. G.; Hershberger, W. K.; Juchau, M. R. Bull. Environ. Contam. Toxicol. 1974,12, 481-486. Walton, D. G.;Penrose, W. R.; Greene, J. M. J . Fish. Res. Board Can. 1972, 35, 1547-1552. Dressler, M. J . Chromatogr. 1979, 165, 167-206. Benoit, F. M.; LeBel, G. L.; Williams, D. T. Bull. Environ. Contam. Toxicol. 1979,23, 774-778. Alben, K. Environ. Sci. Technol. 1980, 14, 468-470. Saxena, J.; Kozuchowski, J.; Basu, D. K. Environ. Sci. Technol. 1977,11,682-685. Basu, D. K.; Saxena, J. Environ. Sci. Technol. 1978, 12, 795-798. Black, J. J. Roswell Park Memorial Institute, 1980, un-
published data. Department of Environmental Conservation Report on Industrial Chemical Survey,Bureau of Industrial Programs; New York State, 1979. Ogan, K.; Katz, E. J . Chromatogr. 1980, 188, 115-127. Glazer, J.; Riggen, R.; Cole, T. In “Chemical Analysis and Biological Fate: Polynuclear Aromatic Hydrocarbons”; Cooke, M., Dennis, A. J., Eds.; Battelle Press: Columbus, OH, 1980; p 439-454.
1976,1,75-84.
Leversee, G. J.; Giesy, J. P.; Landum, P. F.; Gerould, S.; Bowling,J. W.; Fannin, T.; Haddock, J.; Bartell, S. Arch. Environ. Contam. Toxicol., in press. Malins, D. C. In “AquaticPollutants and Biological Effects with Emphasis on Neoplasia”;Kraybill, H. F., Dawe, C. J., Harshbarger, J. C., Tardiff, R. G., Eds.; N.Y. Academy of Sciences: New York, 1977; p 482-496. Bend, J. R.; James, M. 0.; Dansette, P. M. In “Aquatic Pollutants and Biological Effects with Emphasis on
Received for review June 30,1981. Revised manuscript received November 2,1981. Accepted January 19,1982. This study was supported by a grant from the National Science Foundation and an EPA contract ((2164851 to J.J.B.). Although the research described in this article has been funded wholly or in part by the U.S. Environmental Protection Agency it has not been subjected to the Agency’s required peer and policy review and therefore does not necessarily reflect the views of the Agency, and no official endorsement should be inferred.
Trace Enrichment of Fluorinated Organic Acids Used as Ground-Water Tracers by Liquid Chromatography Klaus J. Stetzenbach,* Stephen L. Jensen, and Glenn M. Thompson Department of Hydrology and Water Resources, University of Arizona, Tucson, Arizona 8572 1, and Laramie Energy Technology Center, PO Box 3395 University Station, Laramie, Wyoming 82071
rn A method was developed for trace enrichment of fluorinated organic acids used as ground-water tracers to allow detection at the low ppb level. By replacement of the injection-valve sample loop with a 3-cm column packed with an octadecylsilane bonded phase, tracer can be extracted from a large water sample as it is pumped through the injection valve. The method increases sensitivity by 3 orders of magnitude while retaining the ease and precision typical of HPLC analysis. Introduction The need to trace ground-water flow and monitor waste-disposal sites has prompted the investigation of several classes of compounds to be used for this purpose (I). This research has resulted in the adoption of a group of fluorinated aromatic organic acids for use as tracer compounds. These compounds are particularly well suited for this purpose because they are exotic in the environment, extremely stable, and not sorbed by the aquifer To whom correspondence should be addressed at Laramie Energy Technology Center. 250
Envlron. Scl. Technol., Vol. 16, No. 5, 1982
materials, and they can be separated from other groundwater constituents by bonded-phase high-performance liquid chromatography (BP HPLC). As anions they are readily soluble in water and nonvolatile, which greatly increases the reliability of the quantitative analysis during the monitoring of breakthrough curves. In order to be usable as tracers it is also necessary that these compounds be routinely and accurately measured in the low ppb level or below. The fluorinated aromatic organic acids to be used as tracers are most readily analyzed by means of BP HPLC using a mobile phase of phosphate buffer and methanol, which allows UV detection at wavelengths as low as 200 nm. For this work the absorption bands between 200 and 230 nm provide the best sensitivity. At these wavelengths, standard HPLC methods that permit the direct injection of only 20-200-pL samples typically provide good measurement capability only in the 0.1-1.0 ppm range. Therefore, so that the desired detection limits can be achieved, an on-line precolumn concentrating technique is needed to increase the sensitivity while maintaining the ease and precision typical of HPLC analysis. The removal of organic compounds from large volumes of water by adsorption onto resins followed by elution with
0013-936X/82/0916-0250$01.25/0
0 1982 American Chemical Society
