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### RESULTS AND DISCUSSION

### Aromatic Compounds in Petroleum and Petroleum Products

Fig. 16A
Table 1
Figure 17
Figure 16B
(Table 1)

For discussion purposes, it is convenient to divide the ACs listed in Table 1 into the low-molecular-weight ACs (LACs) and the high-molecular-weight ACs (HACs). This has the convenience of dividing the ACs approximately into: a) the more water-soluble and acutely toxic compounds, LACs; and b) the less water-soluble and more chronically toxic compounds, HACs (National Research Council 1985). The LACs are more prone to dissolution, evaporation, and bacterial degradation and, hence, their levels in the environment generally would decline as the spilled oil weathered. The HACs, on the other hand, are more resistant to dissolution, evaporation, and bacterial degradation and tend to persist in the environment. It is also evident from Table 3 that the LACs comprised about 97% of the ACs in PBCO (generally those components to the left of fluoranthene in Fig. 18).

The pattern of selected components in petroleum, petroleum products, combustion products (from an urban marine sediment), and other materials such as creosote (from coal tar) that may have been associated with the spill sites were compared (Fig. 18, Fig. 19, Fig. 20, Fig. 21). The selected ACs are graphed with the peak areas normalized to one selected alkyl-substituted phenanthrene (C2PHN) component. Comparing the different graphs, one can see that the relative concentrations of the more volatile components, such as naphthalene, and the less volatile components, such as benzo[a]pyrene, varied in the different products. For example, the PBCO collected from the sea surface 11 days following the spill had lost some of the more volatile (and water-soluble) components (e.g., naphthalenes) compared to the C2PHN component in fresh PBCO (Fig. 18). The composition of ACs in a sample collected from a patch of oil from the beach at Snug Harbor, PWS in July 1990, about 15 months after the Exxon Valdez spill (Fig. 19), was quite similar to the 11-day weathered PBCO except for lower levels of naphthalene, C1- and C2- naphthalenes. The lack of continued weathering could have been due to several factors; for example, the formation of an outer layer or crust could have sealed the material from further weathering, or the tar mat may have been in the high intertidal zone and thus less exposed to wave action and other degradation processes.

The pattern of ACs in two other crude oils, Cook Inlet crude oil (CICO) and Kuwaiti crude oil was evaluated and found to be similar to that in the PBCO except for the dibenzothiophenes (DBTs). Prudhoe Bay crude oil contained a considerably larger number and amount of DBTs relative to the C2PHN component; the DBTs were mostly absent in the CICO, but higher in Kuwaiti crude than in PBCO (Fig. 19). The No. 2 fuel-oil pattern (Fig. 20) appeared somewhat similar to PBCO even though No. 2 fuel-oil is basically comprised of a distillation fraction of the crude oil. Bunker fuel, a heavy distillate fraction of petroleum (that generally is diluted with No. 2 fuel-oil to obtain the desired viscosity) contains more of the less volatile components of crude oil (Fig. 20). A pattern of creosote is included (Fig. 21) because molluscs were collected from one station near creosoted pilings and the question was raised if the ACs in those samples were from creosote. Three major components of creosote (phenanthrene, fluoranthene, pyrene), a coal tar distillate, are also major components in combustion products as shown in an extract of marine sediment from an urban area (Duwamish III sediment, Fig. 21), but are not major components in PBCO. These patterns will be important in subsequent sections for comparisons to patterns from analyses of shellfish. It is important to note that when comparing patterns, only similarities or differences can be pointed out. Generally, a pattern should not be linked to a particular source unless the study was appropriately designed and the ambiguities (such as those above) considered.

### Fluroescent Aromatic Compounds in Bile of Fish

Tables 4
5

As discussed earlier, the main utility of bile analyses was to quickly assess exposure of fish to ACs and then to use those results to set priorities for corresponding samples of edible tissue for more comprehensive analysis for individual ACs by GC/MS. Prioritization was done using bile concentrations reported on the basis of bile weight, allowing the selection of edible tissues samples for GC/MS analysis from fish that had elevated concentrations of FACs in their bile. Conversely, fish which exhibited low concentrations of biliary FACs were probably less exposed to ACs and thus were of lower immediate interest.

In this report, the concentrations of biliary FACs in fish will be compared using results based on bile protein. Excretion of metabolites of ACs into bile of fish can be affected by many factors, such as species, age, and feeding status. Previous laboratory studies have shown marked increases in protein concentrations in bile of nonfeeding fish compared to feeding fish (Collier and Varanasi 1987). Moreover, it was shown in these studies that the variation between concentrations of biliary FACs in some fish from the same area were reduced when differences in bile protein were taken into account. In the subsistence study, pink salmon were sampled from a number of areas (e.g., Chenega Bay, Chiniak, Old Harbor, Port Lions, Tatitlek) that included fish being caught upstream in rivers and from the open sea. Results of bile analyses showed significant differences in concentrations of biliary FACs and protein between pink salmon caught in open water compared to pink salmon caught in streams from the same area. For example, at Chenega Bay in 1989, significantly higher mean concentrations of biliary FACs and protein were found in pink salmon caught upstream (220,000 ng PHN equiv./g bile and 36 mg bile protein/g bile, respectively) compared to concentrations in pink salmon from the open sea (7,700 ng PHN equiv./g bile and 1.5 mg bile protein/g bile, respectively, Vol. II, Section A). However, when FACs were normalized for protein, similar concentrations of biliary FACs were observed in pink salmon sampled from the stream (6,400 ± 3,600 ng PHN equiv./mg bile protein) and open sea (4,600 ± 1,900 ng PHN equiv./mg bile protein) and were not significantly different (p = 0.40). 

#### Biliary FACsPHN in Salmon

Table 4
Fig. 22

In 1989, coho salmon were sampled from a number of sites to allow statistical evaluation of the concentrations of FACs in their bile. The results of concentrations of biliary FACsPHN in coho salmon are summarized in Figure 23. Levels of biliary FACs based on bile protein were significantly higher in coho salmon from Ouzinkie, Akhiok, and Tatitlek than concentrations in fish from Angoon, the reference site. Salmon, a pelagic species, can migrate over large distances, thus the finding of elevated concentrations of FACs in Tatitlek fish may be due to these salmon passing through contaminated waters near the site of the oil spill before capture. Insufficient numbers of bile samples from other species of salmon (e.g., sockeye, chum) were sampled to be able to compare FACs between 1989 and 1990.

In 1990, sufficient numbers of chum and sockeye salmon were collected from some areas to allow site comparisons of the concentrations of FACs found in bile of these fish. Mean concentrations of FACs in bile of sockeye salmon from Karluk and Larsen Bay in 1990 were not significantly different from concentrations in sockeye salmon from Yakutat, the designated reference site (Fig. 24). In addition, concentrations of FACs in bile of chum salmon from Old Harbor and Port Graham in 1990 were similar (760 and 750 ng PHN equiv./mg bile protein, respectively), but were significantly lower than concentrations in chum salmon from Ouzinkie (Fig. 25).

#### Biliary FACsPHN in Bottomfish

Table 5

In 1989, concentrations of FACsPHN in bile of halibut from Chenega Bay, Ouzinkie, and Port Lions were significantly higher according to GT2 comparison intervals than concentrations in bile of halibut from Angoon, the reference site (Fig. 26). Statistical comparisons of bile analyses of halibut collected in 1990 were not possible because of insufficient sample size. Moreover, statistical comparisons of bile analyses of other species of bottomfish shown in Table 5 were not possible because bottomfish were rarely sampled from more than three sites, making even qualitative comparisons difficult.

In 1989, concentrations of FACs in bile of Pacific cod from Chenega Bay were significantly higher according to GT2 comparison intervals than concentrations in bile of Pacific cod from Larsen Bay (Fig. 27). Although statistical comparisons could not be made with Pacific cod from Angoon, the reference site, because only one fish was collected, the level of FACs in bile of this fish (480 ng PHN equiv./mg bile protein) was much lower than concentrations in bile of Pacific cod from Chenega Bay. In 1990, no significant differences were found among concentrations of FACs in bile of Pacific cod from all the subsistence sites.

#### Temporal Changes in Biliary FACs in Salmon

Fig. 22

#### Aromatic Compounds in Edible Flesh of Fish

Table 1
Table 6
Table 6
Table 6

On the basis of individual fish samples, excluding the smoked salmon samples, only one sample of fish muscle was moderately contaminated with ACs (Table 7) and only 3 samples had concentrations between 50 and 99 ng/g (category 2). The concentrations exceeded 10 ng/g (category 1) in only 16 samples of fish, the majority of which were collected in 1989. The concentrations of ACs in the edible flesh of the other fish samples were less than 10 ng/g and were generally comparable to the concentrations detected in the same or related species from the reference sites, Angoon and Yakutat. Pink salmon was the only species sampled at several sites in both years, but for the most part, only a small number of samples was collected. The small number of samples and low concentrations of ACs in most samples makes it difficult to evaluate temporal trends of ACs in fish muscle.

In contrast to the samples of fresh fish, samples of smoked salmon from Old Harbor and Tatitlek contained 8,100 and 23,000 ng/g of ACs, respectively (Table 7). The concentrations of benzo[a]pyrene (a carcinogenic HAC) in these samples were 6 ng/g (Old Harbor) and 18 ng/g (Tatitlek), which are low values but are still higher than in any of the other fish collected in this study. It is well known that the smoking process results in the accumulation of ACs in the tissue, hence more samples of smoked fish should be analyzed so that better comparisons can be drawn among the different smoking procedures. Such information should be valuable to subsistence hunters. 

#### Fluorescent Aromatic Compounds in Bile of Marine Mammals

Table 8
Table 9

In contrast to fish, we have not determined whether biliary protein can be used as a normalizing factor to account for differences in the composition of bile of marine mammals. Thus, the concentrations of FACs in these pinnipeds are expressed as ng PHN equivalents per gram bile. The levels of biliary protein in bile of these marine mammals, however, were measured and are given in Tables 8 and 9. As with fish, the range of concentrations of FACs was wide (170-210,000) ng PHN equiv./g of bile. Even though oil was observed to be present on 13 harbor seals at the time samples were collected, the concentrations of FACs varied widely in those animals indicating that many factors, including the degree of absorption/ingestion of oil, metabolism, and excretion determine the levels of metabolites in bile. Hence, considerable caution should be used in comparing the presence of oil on the exterior of the animals with FACs in bile. For example, it would not be known when or how the animal came in contact with the oil. However, 9 of the 12 bile samples with the highest concentrations of FACsPHN (>8,000 ng/g) were from animals that were visibly oiled. Only 4 of the 14 bile samples from harbor seals with concentrations of FACsPHN less than 8,000 ng/g were from animals that were visibly oiled. 

Samples from nine Steller sea lions were collected in 1989 and one in 1990; five were collected from PWS and five from the Gulf of Alaska. The concentrations of FACs in bile from Steller sea lions were generally much lower than the concentrations of FACs in bile from harbor seals (except for one sample, concentrations of FACs in bile from sea lions were <5,000 ng/g bile). None of the sea lions sampled were visibly oiled. Sea lions did not seem to avoid oil, as some were observed swimming in oiled areas. The results for FACs in marine mammals are detailed in Vol. II, Section C.

Temporal trends in concentrations of FACs in bile of mammals were not statistically evaluated due to an insufficient number of samples.

#### Aromatic Compounds in Edible Flesh of Marine Mammals

Table 10

On an individual tissue sample basis, the concentrations of ACs were greater than 10 ng/g in 18 tissue samples from 16 harbor seals, 16 of which were blubber samples (Table 11). Of these 16 blubber samples, four were moderately contaminated with ACs (category 3) and all four were from visibly oiled animals collected from the heavily oiled areas of Herring Bay and Bay of Isles, PWS in 1989. Blubber samples from 10 of the 13 visibly oiled harbor seals (Vol. II, Section D) had concentrations of ACs in categories 2 and 3. The only sample of muscle tissue that exceeded 10 ng/g (16 ng/g) was collected from a visibly heavily-oiled harbor seal pup in Seal Bay in 1989. The only sample of liver tissue that exceeded 10 ng/g (15 ng/g) was collected from a harbor seal in Herring Bay in 1990. The kidney samples that were analyzed had category 1 concentrations of ACs (concentrations were <10 ng/g). Tissue samples collected from seven harbor seals harvested by subsistence hunters in PWS in 1990 had AC concentrations less than 10 ng/g. The concentrations of ACs in edible tissue (blubber, liver, kidney, and muscle) from the marine mammals are detailed in Vol. II, Section D by species, site, and year sampled. 

The limiting factors for statistical comparison of FACs in bile samples from animals with ACs in tissues include the small number of samples and the lack of control studies. As the data base for ACs in marine mammals expands, we will be able to better establish the correlation of FACs in bile with ACs in tissue.

In the blubber samples with elevated concentrations of ACs, the concentrations of alkyl-substituted ACs were similar to or greater than the concentrations of the corresponding nonsubstituted AC (e.g., the ratios of the concentrations of C1PHN to PHN in sample numbers 60-1325 and 60-1412 (duplicate with 60-1424) were 1.7 and 1.8, respectively, Vol. II, Section D). A similar ratio was found in PBCO (e.g., C1PHN to PHN = 2.1) and generally these ratios were opposite of what was observed in the very few fish in this study where ACs were detected in their muscle tissue. Laboratory studies have shown that the alkyl-substituted ACs are more readily metabolized by fish than the nonalkyl substituted components (Schnell et al. 1980). Also, the ratios of LACs/HACs in harbor seals that had greater than 100 ng/g of ACs in their blubber were very high (e.g., 1,000 for sample 60-1325, and 270 for duplicate samples 60-1412/1424, compared to ca. 400 for PBCO). These patterns of ACs in blubber may correspond to the oil these two animals were exposed to because both were collected in 1989, and both were visibly oiled. 

As noted earlier, FACs in bile of fish are indicative of relatively recent exposure to ACs, because ACs that are taken up by the animal are readily metabolized and excreted in their bile. Further, metabolites of ACs in bile of fish have been correlated to exposure to ACs in laboratory and field studies (Stein et al. 1987); however, much less is known about metabolism and excretion of ACs in marine mammals. The limited data available on biotransformation enzyme systems and metabolite profiles of ACs in marine mammals suggest that the extent of metabolism of xenobiotic compounds in marine mammals is intermediate between terrestrial mammals and fish (Boon et al. 1992, Norstrom et al. 1992). Their results suggest that the metabolic capability of cetaceans (e.g., whales) is similar to fish, while the metabolic capability of pinnipeds (e.g., harbor seals, Steller sea lions) is greater than fish or cetaceans, but less than terrestrial mammals. Thus, the marked differences in ACs profiles between pinniped blubber and fish muscle would not appear to be due solely to differences in biotransformation capacities. Differences in kinetics of uptake of ACs, route of exposure, and lipid content of the tissues may all contribute to the differences in ACs profiles observed; however, additional studies are needed to determine the physiological basis for the observed differences.

#### Aromatic Compounds in Invertebrates

Table 12
Table 12
Table 13
Table 13

In this section, those samples that were moderately or highly contaminated (category 3 or 4) will be discussed. The 14 stations that had one or more invertebrate samples with concentrations of ACs in categories 3 or 4 are included in Table 14, and this table shows that stations CHE7, WNB1, WNB3, and KOD3 had the most samples in categories 3 and 4.

#### Comparison of ACs in Molluscs by Area

Tables 12
Table 14

The Chenega Bay sampling area was the first subsistence collection area in the path of the spilled oil and included a large geographic area in southwestern PWS (Fig. 3). Due to the concerns of the community and the potential for shoreline oiling, 12 sampling stations for invertebrates were established. Oil was observed in the area or on the beach at 3 of the sampling stations (CHE1, CHE 10, CHE24), and 4 of the 12 Chenega sampling stations (CHE1, CHE7, CHE10, CHE24) had samples (41 out of 194) with concentrations of ACs that exceeded 100 ng/g. Thirty-one of the 41 samples that were in category 3 or 4 were from CHE7, although the spilled oil may not have been the main source of ACs at this site. This issue will be discussed in a later section. Oil was observed in the area of station CHE1 in 1989 following the spill (this station was located adjacent to the village of Chenega Bay). Booms were put in place to prevent oiling of the beach which received little or no direct oiling. Six of 38 invertebrate samples from CHE1 had concentrations of ACs greater than 100 ng/g (category 3 or 4). At CHE10, there was a tar mat about 1 m wide that extended most of the length of the beach. Oil was also observed on the beach at station CHE24, where one mussel sample of 10 collected had a concentration of ACs in category 3. The high, low, and median concentrations of ACs in the target mollusc samples from CHE1, CHE7, and CHE10 (Fig. 28) demonstrate the wide range of concentrations found in these samples. 

One of the next subsistence seafood sampling areas in the path of the spilled oil was Windy Bay on the southwestern part of the Kenai Peninsula. All of the Windy Bay sampling stations were oiled, but two stations (stations 1 and 3, two small islands in the mouth of Windy Bay) were moderately to heavily oiled. During the three sampling years, 106 invertebrate samples were collected from these 2 stations. Of the 106 samples, only 34 contained concentrations of ACs that exceeded 100 ng/g (Table 12 and Table 14). Only 3 of the 81 invertebrate samples collected from the other 3 Windy Bay stations located at the head of Windy Bay had concentrations of ACs that were greater than 100 ng/g (Table 13 and Table 14). As stated above, the high, low, and median concentrations (Fig. 29) show that the concentrations varied considerably for the target mollusc samples at the most contaminated Windy Bay sampling stations. The mean concentrations of ACs in molluscs from the other sampling stations on the Kenai Peninsula (Lower Cook Inlet, Table 13) were mostly in the not contaminated category.

The spilled oil was carried along Kodiak Island and southward along the Alaska Peninsula. Several areas in this region were selected for invertebrate sampling, however, invertebrate samples from only five stations (K7, KOD3, OHA4, CHG1, CHG3) had concentrations of ACs greater than 100 ng/g (Table 14). At K7, only 1 sample of 4 had concentrations of ACs greater than 100 ng/g; at OHA4, 4 of 26; at CHG1, 2 of 9; and at CHG3, 3 of 10 had concentrations of ACs greater than 100 ng/g. However, at station KOD3, 26 of 30 samples had concentrations of ACs greater than 100 ng/g, 9 of which were greater than 1,000 ng/g (Fig. 7, Fig. 8, Fig. 14). The high, low, and median concentrations of ACs in mussel and clam samples from station KOD3 (Fig. 29) were similarly high in 1989 and 1990, and all were moderately or highly contaminated. Station KOD3 is located on Near Island about a quarter of a mile from Kodiak's boat harbor; therefore, the potential source of the ACs in the samples from KOD3 may have been from boating activity (also see page 32). Station OHA4 is near the village of Old Harbor and within 1/4 mile of a boat harbor. Five samples from the Chignik sampling area (stations CHG1 and CHG3, Fig. 14) were moderately contaminated; however, the concentrations of ACs in these samples were just over 100 ng/g wet weight. The mean concentrations of ACs in molluscs from the Kodiak Island and Alaska Peninsula sampling stations, other than the five discussed here, were mostly in the not contaminated category (<10 ng/g, Table 13) and were comparable to reference area values.

#### Comparison of ACs in Molluscs by Species

#### Aromatic Compounds in Mussels 

Table 13
Table 13

The concentrations of total ACs that exceeded 100 ng/g in mussel samples are listed in order of decreasing concentration by station in Table 15. The highest concentrations of ACs in individual mussel samples were from Windy Bay stations WNB1 (1989) and WNB3 (1990), 18,000 and 5,500 ng/g, respectively. Only 15 mussel samples of 367 were highly contaminated (those with >1,000 ng/g of ACs) and they were from the Chenega Bay, Windy Bay, and Kodiak sampling areas. Forty-seven mussel samples were moderately contaminated (100 to 1,000 ng/g) and were from the three areas above plus the Old Harbor and Chignik sampling areas. It is notable that of these 47 mussel samples, many were in the low end of the moderately contaminated category (i.e., the concentrations of ACs were <200 ng/g in 28 of the 47 samples). Even at the more contaminated Windy Bay stations WNB1 and WNB3, only 15 of the 32 moderately or highly contaminated mussel samples had concentrations greater than 200 ng/g. Furthermore, four samples from each of two sample collections from WNB3 (March 1990 and July 1990) were from the upper intertidal or splash zone and the presence of oil was noted during the March 1990 sample collection. These eight samples were highly contaminated, whereas none of the other mussel samples from WNB3 had concentrations of ACs in concentration category 4. 

#### Aromatic Compounds in Butter Clams

Table 13

Based on the concentrations of ACs in individual samples (208 total), nine samples of butter clams (eight from the Kodiak station KOD3 sampling site collected in 1990 and one from Chenega Bay station CHE7) were highly contaminated with ACs (Table 16). Only 18 samples of butter clams were moderately contaminated with ACs, all of which were from Tatitlek, Chenega Bay, Kodiak, and Old Harbor sampling areas. 

#### Aromatic Compounds in Littleneck Clams

Table 17

#### Aromatic Compounds in Chitons

Table 18
Table 13

#### Aromatic Compounds in Other Invertebrate Species and Miscellaneous Samples

In summary, the sampling design for this study did not include sample collections to explain species differences in concentrations of ACs in mussels, clams, or chitons. The differences in concentrations observed may be relative to exposure but could be due to other factors as well (e.g., uptake and depuration). Only 25 of the more than 1,000 invertebrate samples analyzed were considered highly contaminated (>1,000 ng/g). Of these 25 samples, all were mussels or clams except for one sample of chitons, and all 25 samples were from three sampling areas (Chenega Bay, Windy Bay, Kodiak Island). Invertebrate samples that were either moderately or highly contaminated were primarily from some stations at Chenega Bay (CHE1, CHE7, CHE10, CHE24), Windy Bay (WNB1, WNB3), Kodiak (KOD3), and Old Harbor (OHA4). Even so, the concentrations of ACs in many of the invertebrate samples from the above stations were in the not-contaminated category, and most of the mean concentrations of ACs in these samples fell in either the not-contaminated or minimally-contaminated categories. 

#### Temporal Changes in the Concentrations of Aromatic Compounds in Molluscs

Fig. 30

The GT2 plots show that mussel samples from WNB1 collected in April 1991 had significantly lower concentrations of ACs than those collected in 1989 (Fig. 31). The eight mussel samples from the higher part of the tidal zone of the WNB3 station (discussed on page 25) were the most contaminated with ACs. The concentrations of ACs in these eight samples from the two collections were not significantly different, however, the concentrations of ACs in all of the other mussel samples from WNB3 were significantly lower than those eight samples, and the remaining samples from WNB3 were only minimally or moderately contaminated with ACs. The concentrations of ACs were significantly lower in mussels from WNB3 collected in 1991 compared to those collected in July 1990, but not significantly lower than those collected in March or April 1990. Thus, it is difficult to see a declining trend in concentrations of ACs at WNB3. At WNB2, the concentrations of ACs were not significantly lower in mussels collected in 1991 compared to those collected in 1990. As at Chenega Bay, evaluation of temporal trends in the concentrations of ACs was limited by the sampling scheme (e.g., a limited number of samples).

Most butter clam samples from Chenega Bay station CHE1 were minimally contaminated or not contaminated with ACs (Fig. 32). The concentrations of ACs in all of these samples were not significantly different. The concentrations of ACs in butter clams from CHE7 were moderately or highly contaminated with ACs and were significantly lower in samples collected in April 1990 and June 1991 than in those collected in February 1990. The concentrations of ACs in the samples collected in June 1990 were not significantly different from those collected in February 1990. Also, the concentrations of ACs were significantly lower in butter clams from CHE10 collected in August 1991 than those collected in 1990, with the concentrations being low in all of the samples. 

The concentrations of ACs in butter clams from Kodiak station KOD3 did not decline significantly over the four sampling events in 1990 and the majority of the mean concentrations of ACs were in the highly contaminated category (Fig. 33). Moreover, the concentrations of ACs were consistent for three of the four sampling events.

Most butter clams from Old Harbor station OHA3 were not contaminated with ACs (Fig. 34). In fact, this is a good example of where the concentrations of ACs in samples are so low that they should not be compared statistically. As noted earlier, only two butter clams collected from Old Harbor station OHA4 in 1989 were moderately contaminated with ACs (Table 16). However, all of the other samples collected from this station during 1989 and 1990 had concentrations of ACs that were in category 1 or the lower part of category 2. 

Overall, most of the littleneck clam samples were not contaminated or minimally contaminated and the concentrations of ACs did not differ significantly (Fig. 35). The concentrations of ACs in littleneck clams from station CHE1 were consistently near the low end of the minimally contaminated category and there was no significant decline in the concentrations of ACs in these samples over time. The concentrations of ACs in littleneck clams from station CHE7 were near the 100-ng/g level and were consistent over the 2 years of sampling. Possible reasons for this will be addressed in a later section. The concentrations of ACs in littleneck clams from CHE10 collected in August 1991 were significantly lower than for those collected in April 1990 and June 1991. Littleneck clams from the two sample collections from Kodiak station KOD3 were moderately contaminated and the concentrations of ACs were not significantly different. Again, as for butter clams from KOD3, additional investigation is needed to determine the source of the ACs. The concentrations of ACs were significantly lower in littleneck clams from Windy Bay station WNB4 in April 1991 than for those collected in July 1990.

#### Patterns of Aromatic Compounds in Invertebrates

The samples that were used for pattern comparison generally were randomly selected from those with the higher concentrations of ACs from the stations of interest. For example, examination of the patterns of ACs in selected mussel samples from Chenega Bay station CHE1 and Windy Bay station WNB1 (Fig. 36) show that they are almost identical, suggesting a common source of contamination. These two chromatographic patterns contain many of the alkyl-substituted components typical of petroleum, and, as would be expected, they show considerable losses of the more volatile and more water-soluble components of the LACs (the components to the left of phenanthrene, Fig. 18) when compared to the patterns obtained with PBCO. 

As noted earlier, Windy Bay stations WNB1 and WNB3 could be considered to be one station. Temporal comparison of the patterns of ACs in mussels from Windy Bay stations WNB1 and WNB3 from 1989, 1990, and 1991 shows a continuing increase in the relative concentrations of some of the HACs and dibenzothiophenes (Figs. 36 and 37), otherwise, these three patterns for ACs in mussels from stations WNB1 and WNB3 are very similar to each other. Petroleum from the Exxon Valdez spill was known to impact stations WNB1 and WNB3, and Chenega Bay station CHE1, thus the spilled petroleum was probably the source of the ACs in the molluscs, even though the patterns in the fresh PBCO and the molluscs are not identical. 

The pattern of ACs in mollusc samples from Chenega Bay station CHE10 was somewhat different than the pattern for molluscs from CHE1. For example, a mussel sample from CHE10 (Fig. 38) contained many of the same alkyl-substituted components as those in the mussels from stations CHE1, WNB1, and WNB3. However, the pattern also contained ACs suggestive of combustion products (e.g. phenanthrene, fluoranthene, pyrene, and benz[a]anthracene) when compared to the patterns of ACs in an urban sediment from Puget Sound, Washington (Fig. 21). There was a tar mat on the beach at CHE10 assumed to be from the spilled oil, which could account for the petroleum pattern in the mollusc sample. However, the source of the combustion products pattern in this sample is not clear.

The concentrations of ACs were high in some invertebrates from sampling stations that were not observed to have been exposed to the spilled PBCO. In some of these samples (e.g., Chenega Bay station CHE7) the ratio of LACs/HACs was considerably lower than in samples that probably were exposed to the spilled oil (e.g., WNB3 samples). The pattern of ACs in a butter clam from Chenega station CHE7 (Fig. 38) was similar in some ways to that for the mussel sample from CHE10, but it contained less of the C3PHN, C1DBT, C2DBT, and C3DBT components. The pattern for ACs in the butter clam sample from CHE7 also had similarities to the patterns for combustion products in the urban sediment extract described above and to creosote (Fig. 21). Alkyl-substituted ACs suggestive of petroleum were also present in the butter clam sample, but appear to be a minor component in this example. Creosoted pilings were noted to be in the CHE7 area where the butter clam samples were collected; however, in the absence of a carefully designed sampling and analysis study, it would be difficult to say that the ACs in the clams were from the creosoted pilings. The comparison of patterns shows that caution is important when correlating patterns to potential sources. At the same time, these comparisons show that, if used judiciously, pattern comparison may provide an in-depth look into the potential sources of exposure.

Tatitlek station T1 also was not suspected to have been exposed to the spilled PBCO, even though samples from these stations contained ACs. The patterns of ACs in butter clams from Tatitlek station T1 (Fig. 39) contain the ACs indicative of combustion products, but they were relatively free of the alkyl-substituted phenanthrenes and dibenzothiophenes found in PBCO and in the Windy Bay mussels. 

As noted earlier, molluscs from two Kodiak Island stations (KOD3 and OHA4) were moderately or highly contaminated with ACs. The patterns of ACs in a mussel sample from each station (Fig. 40) were very similar to the patterns of ACs in mussel samples from WNB3 (Fig. 37), except for the higher relative amounts of the more volatile and water-soluble components of the LACs in the KOD3 and OHA4 samples. Higher concentrations of ACs typical of combustion products were present in the OHA4 sample, including fluoranthene, pyrene, and benz[a]anthracene. The pattern of ACs in a sample of butter clams collected from station KOD3 in March 1990 (Fig. 41) was nearly identical to that for mussels collected at the same station in April 1990 (Fig. 40). Also, for butter clams collected at three different times in 1990 at KOD3 (March, May, and September), the pattern of the ACs typical of petroleum in these samples remained consistent over the relatively short period of time, even though the ACs typical of combustion products relative to the C2-phenanthrene peaks appeared to decrease (Fig. 41 and Fig. 42). The consistent pattern of petroleum-related ACs in these samples implies that the clams were being continually exposed to the same source of these ACs over time (probably a local source of ACs, not the Exxon Valdez oil spill). Furthermore, the pattern of ACs in littleneck clams collected from KOD3 in April 1990, (Fig. 41) was nearly the same as that in butter clams from the same station collected in March 1990 (Fig. 41). These patterns suggest that the mussels and the two clam species collected from station KOD3 were all exposed to a similar source(s) of ACs.

The preceding examples of pattern comparison illustrate the need to utilize a thorough and ongoing sampling scheme and to consider the many ambiguities involved when comparing the patterns of ACs in biological samples to the patterns in sources. Also, these pattern comparisons show that the proper use of pattern comparison can be a powerful asset for identifying sources of contamination by ACs.

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