Extractable Organic Matter in Municipal Wastewaters. 2. Hydrocarbons: Molecular Characterizationt Robert P. Eganhouse* and Isaac R. Kaplan
Department of Earth and Space Sciences and Institute of Geophysics and Planetary Physics, University of California, Los Angeles, California 90024 Hydrocarbons isolated from southern California municipal wastewaters were analyzed by high-resolution gas chromatography and computer-assisted gas chromatography-mass spectrometry. Most of the hydrocarbons (56-77%) are not chromatographically resolvable and probably derive from petroleum products such as lubricating oils. Normal, iso-, and acyclic isoprenoid alkanes along with alkylcyclohexanes and numerous series of substituted benzenes and polycyclic aromatic hydrocarbons compose the major fraction of the resolvable hydrocarbons. Polycyclic terpenoids occur as trace constituents and appear also to be of ancient, not recent biosynthetic, origin. However, the virtual absence of 17a(H),lsa(H),2lp(H)28,30-dinorhopane, a specific marker of California oils, indicates that locally produced petroleum is, at most, a minor contributor to these wastewaters. A homologous series of long-chain alkylbenzenes presumably derived from the LAS-type detergents was identified. These compounds seem to be abundant and ubiquitous domestic wastewater constituents that might be exploited as anthropogenic waste tracers in the marine environment.
Introduction Municipal wastes are widely recognized as an important source of petroleum hydrocarbons to the ocean (1-4). More specifically, a number of recent studies have implicated local sewage discharges as the cause of hydrocarbon pollution in both river and marine waters and sediments (4-10). Often the connection between source and sedimentary sink has been established by gas chromatography (GC) alone or by GC in conjunction with ancillary fingerprinting techniques such as infrared and UV-visible absorption and fluorescence spectroscopy. Even though these techniques can provide a means of correlation for a complex organic mixture such as petroleum, they are generally incapable of elucidating detailed structural information without considerable effort. Computer-assisted gas chromatography-mass spectrometry (GC-MS), on the other hand, has been successfully used to verify precise structures of individual sewage tracers such as coprostanol (11, 12). Furthermore, computer manipulation of spectral data (e.g., mass fragmentography), allows rapid examination and tentative identification of important series of marker compounds, typically present only at trace levels (13, 14). +Publication No. 2218, Institute of Geophysics and Planetary Physics, University of California at Los Angeles. 0013-936X/82/0916-0541$01.25/0
Several studies in recent years have, in fact, used GCMS in the examination of municipal wastewaters (10, 15-19). However, most of them have focused either on the major components of the total extractable organic matter, most of which are polar nonhydrocarbons (20,21), or on specific environmentally significant compounds such as the chlorinated hydrocarbons. A notable exception is the recent study made by Barrick (10) in the Seattle-Puget Sound area. This report differs from those previous in that it presents an extensive and detailed molecular inventory of the hydrocarbon compositions of five effluents from southern California. Because of the multiple inputs of petroleum to the ocean in this region (5, 22), we were mainly interested in establishing a comprehensive description of wastewater hydrocarbons for later use in organic source differentiation in marine sediments. Therefore, a large number of samples were analyzed to monitor both temporal and plant-to-plant variability. One highly important aspect of this study was to identify compounds that originate from sewage. These will be essential in future studies of the environmental distribution and fate of wastewater organics.
Experimental Section Sampling and Extraction. Samples of final effluent were collected from the four major treatment plants in southern California according to methods outlined earlier (2,20); locations of these plants and general characteristics of their effluents are described therein. For a brief summarization, duplicate 3-L 24-h composite samples of final effluent were obtained at various intervals during 1979. One of the two samples was immediately preserved at pH 1 (HC1) with hexane and refrigerated at 10 OC; the other was filtered with a Whatman GF/A filter, and 2 L of the filtrate was preserved and stored in the same fashion. All samples were extracted with chloroform (CHCl,). The combined hexane/CHCl, extracts were concentrated, dehydrated, treated for sulfur removal, and esterified (BF,-MeOH). Separation of the esterified extracts into general compound classes was achieved by chromatography on silica gel coated thin-layer plates (CH2C12).Five bands including the total hydrocarbons (THC) were removed and eluted with appropriate solvents. Following high-resolution GC analysis, selected THC fractions were rechromatographed by pentane elution and separated into two or more subfractions. In most cases only two subfractions were isolated: (1)total aliphatics (Al, R, 0.92-1.0); (2) total aro-
0 1982 Amerlcan Chemical Society
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matics (Ar, Rf 0-0.92). One sample (Hyp-7 mi, October 15,1979) was separated into three subfractions: (1)Al (Rf 0.92-1.0); (2) Arl (R, 0.86-0.92); (3) Ar2 (Ef 0.05-0.86). The A1 subfraction contained normal, branched, and cyclic hydrocarbons, whereas the Arl and Ar2 subfractions were isolated on the basis of their UV fluorescence/absorption and chromatographic properties. For example, the Arl band absorbs UV strongly and contains many of the alkyl-substituted benzenes in addition to a portion of the >C20 cyclics. The Ar2 subfraction includes a broad fluorescent band (Rf0.05-0.35) corresponding to the polycyclic aromatic hydrocarbons (PAH). Gas Chromatography. Following gravimetric analysis (cf. ref 2,2O), the hydrocarbons were chromatographed on high-resolution glass capillary columns (SE-54,15 m X 0.25 mm) with Carlo Erba FTV 2150 and 2350 instruments equipped with flame ionization detectors and "Grab"-type split/splitless injectors (23). The analyses were performed by splitless injection of hexane solutions according to the following protocol: 40-260 "Cis,, at 4 "C/min, injector/ detector temperature 275 "C, linear carrier (He) velocity 30 cm/s. Hydrocarbon concentrations reported here were cbmputed by using response factors determined from an n-alkane standard that was run the same day (20). In order to test the efficiency of hydrocarbon recovery, known amounts of two standards, triisopropylbenzene (RS,)and deuterated tetracosane (RS,), were added to each effluent sample prior to preservation. On the basis of the gas chromatographic analyses, recoveries of these compounds were 59 f 7% and 64 f 8%,respectively. No attempt has been made to correct the hydrocarbon concentrations reported here for recovery. Selected THC fractions submitted to GC-MS analysis and subsequently separated into A1 and Ar subfractions were also analyzed by GC using a Hewlett-Packard 5830A instrument equipped with a split/splitless injector and flame ionization detector. In these cases, a fused silica capillary column (SE-54,30 m X 0.25 mm) was employed under conditions similar to those described above except that the column upper temperature limit was 275 "C. Gas Chromatography-Mass Spectrometry. GC-MS analysis of selected representative hydrocarbon fractions was performed on a Finnigan 4000 quadrupole mass spectrometer interfaced with a Finnigan 9610 gas chromatograph. Chromatographic separation was achieved with a fused silica capillary column (30 m X 0.25 mm, J & W; wall coated with SE-54) under the following conditions: injector temperature, 275 "C; column temperature, 35-280 OC at 4 "C/min; linear carrier (He) velocity, 30 cm/s. Mass spectrometric conditions were as follows: ion-source temperature, 240 OC; electron-beam energy, 70 eV. Electron impact mass spectral data were acquired and processed with a Finnigan INCOS 2300 data system. The identifications presented here are based upon one or combinations of the following: (1)comparison of sample spectra with those of authentic reference compounds, computer library spectra and/or published spectra; (2) coinjection with authentic standards; (3) gas chromatographic retention times (SE-54). As used here, the term "mass fragmentography" refers to the computer-assisted extraction and display of a specific ion from the total ion current.
Results and Discussion General Characteristics. General features of the wastewater hydrocarbons are given in Table I, and an inventory of prominent constituents identified by GC/MS is provided in Table 11. Representative gas chromatograms of the total hydrocarbon fractions for each effluent 542
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can be found in Figure 1; for reference, Figure 2 provides greater detail on individual aliphatic and aromatic components of the Hyperion 7-mile (Hyp-7 mi) effluent. Based upon our analyses, the outstanding characteristic of the hydrocarbons in southern California's wastewaters is the consistency in their composition. Although a few specific compounds (e.g., benzothiophenes) were found in only one or several of the effluents, the vast majority of the identifiable constituents were common to all. Variations between effluents and with time for any one effluent appear only in the relative abundances of the major compound groups. Even in the case of specific trace constituents such as the pentacyclic triterpanes, mass fragmentography reveals nearly identical distributions for all of the effluents (Figure 3). A most striking example of this compositional uniformity is the JWPCP effluent (Joint Water Pollution Control Plant, Los Angeles County; cf. ref 2), which was sampled 24 times during 1979. Gas chromatograms of the total hydrocarbons from these samples are virtually superimposable. In agreement with recent work (9,10,15) the predominant resolvable hydrocarbons are the normal alkanes. Total n-alkanes recovered from unfiltered samples range from 0.066 to 6.29 mg/L, and their concentrations are linearly correlated with that of the total hydrocarbons (r = 0.97, Table I). By comparison, Seattle's Metro waste effluent contains approximately 0.05-0.15 mg of n-alkanes/L (particulates only; ref IO). The variation in total n-alkane contents we observed for unfiltered samples probably results from both the influent nature and, more importantly, the waste treatments involved. For example, the high values found for the Hyp-7 mi samples reflect the fact that this effluent is a slightly diluted sludge. The other wastewaters sampled are either primary effluents or a combination of primary plus secondary effluents and, with respect to total solids and extractable organics, are much more dilute (2). Whether filtered or not, all effluents we examined had roughly equivalent proportions of total aliphatic and aromatic hydrocarbons (2); however, filtered samples typically showed an enhancement in the relative concentrations of the lower and medium molecular weight aromatics (viz., alkylbenzenes, naphthalene, and biphenyl plus alkyl homologues). This may reflect some fractionation of the aromatics on the basis of water solubility (24). Planimetric measurements of the gas chromatograms showed the majority (5677%) of wastewater hydrocarbons to exist as nonresolvable branched, cyclic, and aromatic constituents (cf. Table I and ref 3,9,1O) which appear as broad unimodal and bimodal humps (i.e., unresolved complex mixtures (UCM); cf. Figures 1 and 2). Such humps are frequently encountered in petroleum-polluted sediments (4,7,9, 25, 26) and are often attributed to weathered and/or microbially degraded petroleum residues. However, they are also characteristic of refined products such as lubricating and fuel oils (3,6, 7,27) and may occur as the microbial decomposition and resynthesis products of biogenic materials (28). For most samples examined here, the UCM was unimodal, centered in the range 1800-2300 (Kovats Index) and extended from 1000 to 3200; bimodal distributions centered at 2100-2200 and 2600-2700 were also observed. Qualitatively, the appearance of the UCM is quite similar to that reported in the Seattle study (IO). Whereas the presence of the UCM in these samples is strongly suggestive of a petroleum origin, the variable nature of the distributions is most likely to changing input of assorted degraded and refined petroleum residues and wastes. Van
Table I. General Characteristics of Total Hydrocarbon Fractions in Southern California Municipal Wastewaters, 1979 c n-alkanes, max mean %res /.lg/L UCM maxb n-alkane OEPC HC's, % Pr/n-C,, effluenta sampling date THC, mg/L Unfiltered Samples 0.51 39.5 1.01 405 1930 10.6 1/15/79 JWPCP 0.54 1.05 573 2100/2660 14.2 2/15/79 0.48 1.08 2120/2680 7 04 14.3 3/14/79 0.51 45.5 1.05 174 2130 16.6 4/4/79 0.48 1.04 592 2100/2700 13.0 5/15/79 0.38 1.05 679 220512700 15.6 6/15/79 0.46 32.5 1.02 580 213512730 15.9 7/16/79 0.43 1.08 1180 2285 20.9 8/15/79 0.46 1.05 719 214012625 18.2 9/13/79 0.43 31.1 1.05 49 1 2200 16.1 10115179 11115 179d 0.48 1.08 532 2175 14.0 12/13/79 0.62 34.1 1.10 113 1820 5.1 1/15/79 Hyp-5 mi 0.55 27.9 1.07 96 1890 6.9 4/18/79 0.53 24.5 1.00 66 1995 5.7 7/16/79 0.59 30.7 1.10 105 1980 6.5 10/16/79 0.53 36.8 1.14 6290 1890 397 1/15/79 Hyp-7 mi 0.52 30.9 1.17 5090 1870 314 4/18/79 0.53 26.2 1.16 3500 1960 7/16/79 297 0.51 29.3 1.17 4490 1950 297 10/15/79 0.55 1.14 43.0 97 1865 5.7 1/15/79 OCSD 0.71 30.4 1.08 125 1860 10.9 4/12/79 0.55 38.6 1.12 98 1945 7.0 7/16/79 0.15 31.1 1.11 108 1910 7.8 10/16/79 0.57 39.6 1.11 781 1 77 512660 11.8 1/16/79 CSD 0.49 37.5 1.08 271 2285 4/17/79 8.2 0.49 51.9 1.07 10.4 1/11/79 305 1675 0.47 36.9 1.07 1710 1640 18.7 10/16/79 Filtered Samples 0.51 30.2 0.99 98.3 2160 1/15/79 2.18 0 37 1.04 50.5 2215 2/15/79 1.78 0.43 1.02 108 2255 2.36 3/14/79 0.44 1.01 35.4 60.1 2170 1.53 4/4/79 1.00 0.37 48.7 2255 1.29 5/15/79 0.32 1.00 78.9 2200 2.13 6/15/79 0 44 23.3 1.04 66.6 2280 2.18 7/16/79 0.40 155 2180 1.08 3.29 8115/79 0.38 1.02 52.1 2175 1.61 9/13/79 0.46 1.10 55.2 1870 31.6 2.19 10/15/79 111 15/79d 12113179d 0.56 1.02 22.5 34.6 1850 Hyp 5 mi 1.92 1/15/79 0.54 1.04 32.1 47.2 1790 3.04 4/18/19 0.62 1.00 51.3 2035 22.9 2.90 7/16/79 0.60 1.01 26.7 43.9 2020 10116/79 3.10 0.58 263 1860 1.16 36.0 Hyp-I mi 17.6 1/15/79 0.53 1.17 33.7 203 1840 13.2 4/18/79 0.53 1.14 27.9 128 1930 9.42 1/16/79 0.50 1.14 35.9 10115/79 97.4 1900 7.08 0.47 32.2 2230 1.09 30.8 OCSD 1/15/79 1.23 0.53 1.04 25.4 25.0 1985 4/12/19 1.63 0.57 0.96 44.3 33.0 1895 7/16/79 1.73 10/16/19d 1/16/79 CSD 1.22 85.7 1670 1.02 0.59 44.2 99.5 2130 0.52 4/17/79 1.78 1.07 43.0 7/17/79 108 2310/1530 2.21 1.01 0.45 31.7 10/16/79 253 1690 0.53 44.2 2.66 0.87 a JWPCP (Joint Water Pollution Control Plant, Los Angeles County), Hyp-5 mi (City of Los Angeles, 5-mile outfall), Hyp7 mi (City of Los Angeles, 7-mile outfall), OCSD (Orange County Sanitation District), CSD (City of San Diego). Maxima of the unresolved complex mixture given in Kovats indices; if bimodal, numerator refers to index of the dominant mode. Mean OEP is the average of all running OEP values calculated for n-C,, t o n-C,. Sample lost, JWPCP
Vleet and Quinn (9) found that 85% of the hydrocarbons in Providence, RI, effluents were contained in the UCM. However, their analyses were performed on (packed) columns of lower resolution than those used in this study. Thus, comparison of their results with ours is difficult. In the past, the pristane/n-C1, ratio has been used to indicate the degree of microbial degradation because nalkanes are metabolized more readily than branched species (15). The consistent ratios we observed, particularly in the case of the Hyp-5 mi and Hyp-7 mi effluents
(Table I), which represent the resultant products of different treatments to essentially the same influent, suggest but do not prove that waste treatment (e.g., anaerobic digestion) probably has little effect on the catabolism of hydrocarbons. This same conclusion was reached by Giger et al. (15),who examined primary and secondary wastewater effluents. In contrast, undisturbed surface sedimenb (0-1 cm) in the immediate vicinity of the JWPCP outfalls (29) as well as surface sediments in deeper portions of the adjacent San Pedro Basin (5,29)contain almost no normal Envlron. Scl. Technol., Vol. 16, No. 9, 1982 543
Table 11. Selected Compounds Identified in Wastewater Effluents from Southern Californiau concn peak no. compound range,b pg/L means of identificationC Alkanes n-alkanes, C,,-C,, 70-6300 SM, C, RT, STD d isoal kanes 4-450 SM, RT 2-methylundecane 3 2-me thyldodecane 8 2-methyltridecane 11 2-methyl te tradecane 14 2-methylpentadecane 16 2-methylhexadecane 19 7-650 SM, RT acyclic isoprenoids: 2,6-dimethylundecane 5 2,6,1O-trimethylundecane 9 2,6,10-trimethyldodecane 12 2,6,10-trimethyltridecane 14 2,6,10-trimethylpentadecane 17 2,6,10,14-tetramethylheptadecane 22 2,6,10,14-tetramethylnonadecane 25 unknown branched C,,:, 30 Cyclics n-alkylcyclohexanes 4-450 SM, RT pentylcyclohexane 2 hexylcyclohexane 6 hep tylcyclohexane 10 oc tylcycl ohexane 13 nonylcy clohexane 15 decylcyclohexane 18 undecylcyclohexane 20 dodecylcy clohexane 21 tridecylcyclohexane 23 tetradecylcyclohexane 24 pentadecylcyclohexane 28 hexadecylcy clohexane 31 hep tadecylcy cldhexane 32 octadecylcyclohexane 33 alkyldecahydronaphthalenes 0.5-50 SM C, Decalins 1 C, Decalins 4 C, Decalins 7 steranes
