Environ. Sci. Technol. 1993, 27, 699-708

Comparison of High-Performance Liquid Chromatography/F luorescence Screening and Gas Chromatography/Mass Spectrometry Analysis for Aromatic Compounds in Sediments Sampled after the Exxon Valdez Oil Spill

Margaret M. Krahn,' Gina M. Ylltalo, Jon Buzltls, Sln-Lam Chan, and Usha Varanasl

Environmental Conservation Division, Northwest Fisheries Science Center, National Marine Fisheries Service, National Oceanic and Atmospheric Administration, 2725 Montlake Boulevard East, Seattle, Washington 98 112

Terry L. Wade, Thomas J. Jackson, and James M. Brooks

Geochemical and Environmental Research Group, Texas A&M University, 833 Graham Road, College Station, Texas 77845

Douglas A. Wolfe and Carol-Ann Manen

National Ocean Service, Coastal Monitoring and Bioeffects Assessment Division, Office of Ocean Resources Conservation and Assessment, National Oceanic and Atmospheric Administratlon, Rockville, Maryland 20852

After the grounding of the Exxon Valdez, sediment samples were collected to determine the degree and distribution of the oiling. Sixty sediments from 10 sites in Prince William Sound, AK, were analyzed for Prudhoe Bay crude oil (PBCO) using a rapid HPLC screening method that measured fluorescence at wavelength pairs specific for two- and three-ring petroleum-related aromatic compounds (ACs). Concentrations of individual ACs in the sediments were also determined by GUMS to compare the results of the two methods. Concentrations of ACs measured by HPLC screening were highly correlated with the sums of ACs determined by GUMS, thus validating the screening method as an effective tool for estimating concentrations of petroleum-related ACs in sediments. Moreover, differences in HPLC chromatographic patterns among sediments suggested different sources of contam- ination, e.g., crude oil or diesel fuel. Finally, GUMS analyses confirmed that PBCO was a primary source of contamination in many sediments.

Introduction

The grounding of the Exxon Valdez on March 24,1989, spilled 11 million gallons of Prudhoe Bay crude oil (PBCO) into the largely uncontaminated waters of Prince William Sound, AK. As part of the Natural Resource Damage Assessment effort, thousands of sediment samples were collected during the next 3 years to determine the distribution of the spilled crude oil. Because analysis of all these samples by gas chromatography/mass spectrom- etry (GC/MS) would be prohibitively expensive and excessively time-consuming, an alternative was sought. We had recently developed a high-performance liquid chromatography (HPLC) method with fluorescence de- tection to rapidly screen for aromatic compounds (ACs) in sediments from urban sites (1). Accordingly, we used this semiquantitative method to estimate PBCO concen- trations in sediment samples from the oil spill area. Thus, a relative ranking of the contamination levels in these sediments was obtained and was used to identify samples for priority analysis by GC/MS.

Previously, many of the methods employed to screen for petroleum-related ACs in sediments were nonchro- matographic procedures, e.g., synchronous scan or normal fluorescence spectrometry (2-5). However, these methods are of limited value because they do not separate interfering

biogenic compounds from the analytes (6, 7). Alterna- tively, Killops (8) used an HPLC procedure to assess qualitative differences in levels of petrogenic contami- nation in sediment extracts, but no quantitations were made. Another chromatographic procedure-flash evap- oration/pyrolysis GC/MS-screened for ACs in sediments, but the detection limit for ACs was high (5 ppm) and elemental sulfur interfered with the analyses (9).

The spillage of PBCO into Prince William Sound offered an opportunity to test the HPLC method on sediment samples contaminated by a distinct, identified source. These PBCO-contaminated sediments differed from the previously analyzed urban sediments in the variety and types of ACs present. For example, sediments from urban estuaries contained high concentrations of many anthro- pogenic compounds, including high proportions of the four- to six-ring ACs characteristic of the combustion of fossil fuels (1, 10). In contrast, the predominant ACs in fresh PBCO are one- to three-ring alkylated ACs (11-15). As crude oil is degraded in time by physical, chemical, and microbial processes, the aromatic fraction of the weathered oil soon becomes dominated by those ACs that are most resistant to weathering, i.e., highly alkylated naphthalenes, phenanthrenes, and dibenzothiophenes (16,17). Accord- ingly, to detect and quantitate the ACs in sediment samples collected following the spill of PBCO, we selected the two sets of fluorescence wavelengths, Le., for naphthalenes/ dibenzothiophenes and for phenanthrenes, that had been used previously to detect metabolites of these ACs in fish bile (11, 18).

More than 400 sediment samples from the Alaskan oil spill have been screened to date. In this paper, the HPLC results are presented for 10 sites (six depths at each site) that represent a range in degree of oiling and in types of cleanup treatments used to remove the oil from the environment. In addition, detailed analyses of these samples were performed by GUMS, so that the HPLC screening and the GUMS methods of measuring PBCO contamination could be compared for sediments contam- inated by an oil spill. The concentrations of ACs measured by HPLC screening were found to be highly correlated with the sums of ACs determined by GC/MS, thus validating screening as a tool for estimating concentrations of petroleum-related ACs in sediments. Interestingly, HPLC chromatographic patterns were not consistent among all the extracted sediments. The differences were not due solely to the degree of weathering of the crude oil,

0013-936X/93/0927-0699$04.00/0 0 1993 Amerlcan Chemical Society Environ. Sci. Technol., Vol. 27, No. 4, 1993 699

4 Block Island 9 Sleepy-Bay 5 Herring Bay 10 MacLeod Harbor

Extent of Exxon V8ldezOli Coverage: April 20, 1989 (non-continuour)

Figure 1. Chart of Prince William Sound showing the sites from which sediment samples were collected. The Exxon VaMer was grounded near Bligh Island on March 24, 1989, and the path of the spill is shown on the chart.

but reflected different sources of ACs, e.g., crude oil or diesel fuel. Consequently, the source of contamination needed to be confirmed from GC/MS results by comparing relative concentrations of petroleum-related ACs in the sediments to those from possible sources.

Experimental Section

Standards, Marine lubricating oil, No. 2 fuel oil, Kuwaiti crude oil, and South Louisiana crude oil were American Petroleum Institute reference oils. Weathered Kuwaiti crude oil was collected from a near-shore trench, near Abu Ali, Saudi Arabia, in May 1991. Diesel fuel was obtained from a supplier in Seattle, WA. A sample of PBCO was obtained from the oil remaining in the hold of the Exxon Vuldez, and a weathered PBCO sample was collected from the upper tidal zone of Snug Harbor (Knight Island) about 16 months after the spill occurred. A stock solution was prepared by dissolving PBCO in methylene chloride to give a final concentration of 5.40 pg/pL. A series of dilutions, each containing polystyrene internal standard (2500 ng/pL), wasprepared from the stockPBCO to give multilevel calibration standards (at concentrations of 1340, 269, 67.3, 13.5, and 2.7 ng/pL) for quantitating PBCO in sediments.

Collection of Sediment Samples. The sediment samples were collected from selected sites (Figure 1) between June 27 and July 22,1990, during cruise DA90-01 of the NOAA RIVDuuidson. Except a t MacLeod Harbor, surficial (top 2 cm), intertidal (0 m) sediments were collected by a beach party within 1 h of low tide. Subtidal sediments (including the 0-m sample from MacLeod Harbor) were collected by scuba divers a t depths of 3,6, and 20 m. From each of these depths (depth refers to

height of water column above sediment), three replicate samples, each a composite of eight randomly located subsamples along a 30-m transect, were placed into previously cleaned glass jars. In addition, samples from 40 and 100 m were collected using a Shipek, Van Veen, or Smith-MacIntyre grab sampler. Surficial sediments were composited from at least three separate grabs at each site to make up three replicate samples. For each site/depth combination, approximately equal portions from the three replicates were subsequently composited to make up the samples analyzed and reported in this paper. Samples were stored on ice for up to 4 h prior to final compositing on board ship and were frozen at -80 OC immediately thereafter and kept frozen until analyzed.

Degree of Intertidal Oilingand Treatment History of the Sites. The sites (Figure 1) reported here represent a range in the degree of intertidal oiling and in the treatments used to remove the oil (Table I). Houghton, Lees, and Ebert (19) reported the degree of oiling based on scientists’ observations at each site. Three of the sites (Olsen Bay, Port Fidalgo, MacLeod Harbor) were not oiled by the spill and were not treated (category 1). Seven of the oiled sites (Table I) represented the types of treatment undertaken in attempts to remove the oil from the intertidal areas: category 2, (a) untreated in “set aside” areas or (b) low-pressure washes or bioremediation; and category 3, high-pressure, hot-water washes.

Sonic Extraction of Sediments for Screening Anal- yses. Sonic extraction and HPLC/fluorescence analysis were used to screen the sediment samples from the oil spill area for fluorescent ACs. Equipment and procedures were identical to those reported earlier (1). Briefly, sediment (1.0 g), sodium sulfate (10 g), activated copper

700 Environ. Sci. Technoi., Voi. 27, No. 4, 1993

Table I. Degree of Intertidal Oiling and Treatment History for Sites in Prince William Sound

site degree of intertidal oiling treatment history'

Bay of Isles light/moderateb 2b

Chenega Island oiled 2a Herring Bay light/heavyb 2a and 2b MacLeod Harbor none 1 Northwest Bay moderate/heavyb 3 Olsen Bay none 1 Port Fidalgo none 1 Sleepy Bay moderatel heavyb 3 Snug Harbor light/moderate/ heavyb 2a and 2b

a Categories of treatment to remove oil from the intertidal areas were defined as follows: 1, sites with no known oiling and no record of treatment; 2, oiled sites (a) untreated in "set aside" areas or (b) moderately treated (e.g., by manual pickup, low-pressure washes, or bioremediation); 3, oiled sites subjected to hot-water, high-pressure wash. As reported by Houghton et al. (19).

Block Island heavyb 3

(1 mL), and methylene chloride (20 mL) were mixed together in a centrifuge tube. The tubes were placed into a sonic bath for 15 min and then centrifuged at 1500 rpm for 5 min. Each extract was decanted into a 50-mL concentrator tube. To the sediment remaining in each tube was added 10 mL of methylene chloride. The resulting mixture was stirred and sonicated for 5 min and then centrifuged and decanted as before. This step was repeated, and the three extracts were combined. The polystyrene HPLC internal standard (100 pL; 50 pg/pL) was added, and each solution was concentrated by evaporation to -4 mL.

HPLC/Fluorescence Screening of Sediment Ex- tracts. A portion (15 pL) of the extract was injected onto a size-exclusion HPLC column and was eluted isocratically with methylene chloride (flow of 2.5 mL/min); the fluo- rescence was monitored at 260/380 nm (phenanthrene wavelengths; where the phenanthrenes and the polystyrene internal standard fluoresce) and at 290/335 nm (naph- thalene wavelengths; where the naphthalenes and the dibenzothiophenes fluoresce). The HPLC system had been calibrated with dimethylnaphthalene to determine the beginning of the elution of the fraction containing the ACs (retention times of >8.2 min for our HPLC system) (I). Then, the chromatographic areas of this AC fraction were integrated at each set of wavelengths (I), and the PBCO equivalents (pg/g, wet wt) were calculated by comparison to equivalent areas from the multilevel series of PBCO standards. The retention times for the size- exclusion column are remarkably stable over time, seldom changing more than 0.05 min during the analysis of more than 200 samples over a period of several months.

Fresh, rather than weathered, PBCO was used as a standard because the degree of weathering of the oil in each sediment was unknown. Under our analytical conditions, -1 ppm (pg/g) PBCO can be quantitated at phenanthrene (PHN) or naphthalene (NPH) wavelengths. Limits of detection can be lowered by concentrating the extract or by injecting more extract onto the HPLC column. However, if sediments contain substantial proportions of ACs from sources other than PBCO, e.g., fuel oils, lubricating oils, or pyrogenic ACs, HPLC screening concentrations might be more appropriately calculated in terms of phenanthrene or naphthalene equivalents as described in our earlier study (I).

Sediment Extraction for GC/MS Analysis. The method for sediment extraction, cleanup, and analysis by

GC/MS has been described previously (20) and is sum- marized here briefly. A surrogate standard (1,l-dichlo- robenzene-d*, naphthalene-de, acenaphthene-dlo, phenan- threne-dlo, chrysene-dlz, and perylene-d12 was added to the sediment samples after freeze-drying (approximately 10-50 g of sediment based on the original wet weight), and the mixture was then Soxhlet-extracted for 4 h with methylene chloride. The extract was concentrated by evaporation, and the solvent was replaced with hexane.

Cleanup of Sediment Extracts for GC/MS Analysis. Separation of aromatic hydrocarbons from lipids was accomplished by alumina/silicagel chromatography. Silica gel was slurry-packed in methylene chloride over alumina, and copper was added to the top of the column to remove elemental sulfur. The extract, in hexane, was transferred to the column. The aliphatic compounds were first eluted with pentane and then the ACs were eluted with 1:l methylene chloride/pentane. The fraction containing ACs was concentrated for analysis by GC/MS in the selective ion mode (SIM).

GC/MS Analysis for ACs in Sediments. The mass spectrometer was tuned daily to the standard Hewlett- Packard autotune parameters using perfluorotributyl- amine (PFTBA). The GC/MS was initially calibrated and detector linearity was determined by injection of standards (including all surrogate standards) a t four concentrations (0.2-2.5 ng/pL). A linear relationship between concen- tration and response was demonstrated by a correlation of a t least 0.99 before analysis of samples was initiated. The samples were injected in the splitless mode onto a 0.25 mm X 30 m (0.032-pm film thickness) DB-5 fused- silica capillary column at an initial temperature of 60 "C, temperature programmed at 10 "C/min to 300 "C, and held at the final temperature for 6 min. Peaks were identified from their molecular ions, the ratio of the quantitation ion to the secondary ion, and retention time.

Quality Assurance for Sediment Screening Anal- yses. Each set (13) of sediment samples for HPLC screening was accompanied by a method blank, a reference sediment material, and a replicate of one of the samples. In addition, the HPLC system was calibrated daily and duplicate HPLC analyses were performed on -10% of the samples.

Quality Assurance for GC/MS Analyses of Sedi- ments. Each set (12-15) of sediment samples for GC/MS was accompanied by a combusted sand system blank (including freeze-drying, reagents, solvents, and surrogate standards) and National Institute of Standards and Technology reference material which were carried through the entire analytical scheme in a manner identical to samples.

Statistical Methods. The relationship between con- centrations of ACs in sediments determined by GC/MS and concentrations measured by HPLC screening of the same samples were evaluated by correlation (21). The concentrations obtained by each method were first log- transformed to improve homogeneity in the variances.

Results

Statistical Comparisons of HPLC Screening and GC/MS Methods. The concentrations of fluorescent ACs in extracts in sediments from Prince William Sound were

Environ. Sci. Technol., Vol. 27, No. 4, lg93 701

Table 11. Aromatic Compounds in Sediments As Analyzed by HPLC Screening and by GC/MS

depth (m)

0 3 6

0 3 6

0 3 6

0 3 6

0 3 6

0 3 6

0 3 6

0 3 6

0 3 6

0 3 6

PBCO equiv (screening) ( d g )

NPHX PHNX

6 4 2

35 27 34

13 2 1

690 9 2

3 7 6

230 64 98

1 1 2

1 1 1

11 12 13

56 2 1

26 11 5

87 88

110

41 3 1

970 26 5

5 25 23

410 180 310

4 3 5

0 2 2

33 43 47

100 3 4

AC sums (GCiMS) (ngig)" CNPHs CDBTs CPHNs

depth (m)

200 <18* <15

18 25 30

12 <19

11

294 58 26

47 91 71

576 46 96

50 11 16

<27 <6 <5

11 11 57

118 5

39

168 33 31

33 56 62

61 31 58

43 1 154 84

<21 <33 <21

571 167 437

<18 <12 <19

<7 <13 <13

54 62 28

47 <14 <16

Bay of Isles 370 20 105 40 97 100

Block Island 100 20 160 40 173 100

Chenega Island 136 20 77 40

150 100 Herring Bay

998 20 388 40 210 100

MacLeod Harbor 25 20 87 40 68 100

Northwest Bay 809 20 417 40

1234 100 Olsen Bay

27 20 29 40 32 100

Port Fidalgo <14 20 <26 40 <26 100

Sleepy Bay 162 20 190 40 131 100

Snug Harbor 99 20 9 40 6 100

PBCO equiv (screening) ( d g )

NPHX PHNX

11 8

11

7 63 16

1 9

10

5 8 5

15 18 6

31 15 17

4 10 19

3 2 7

17 4

12

5 31 20

29 22 30

22 210 48

3 25 24

12 23 15

58 67 26

130 58 46

15 31 54

8 5

20

77 11 32

12 75 54

AC sums (GUMS) (ngig)" CNPHs CDBTs CPHNs

307 105 78

78 59

154

<9 27 71

55 80 71

101 119 83

97 91

183

68 125 111

<8 <22 168

116 100 155

41 86

206

477 122

<22

95 129 93

<32 <33

82

167 59 48

<18 <19 <18

357 116 191

<5 <26 <22

<12 <6 38

65 46 50

<18 28 15

1169 311 99

290 380 309

68 119 251

447 191 161

115 164 119

1097 305 506

41 53 99

20 <16 110

149 101 177

19 97

152

Summed concentrations: CO-C4 naphthalenes, CNPHs; Co-Cd phenanthrenes, CPHNs; cO-c3 dibenzothiophenes, CDBTs. NPHs and DBTs fluoresce at NPH wavelengths and PHNs at PHN wavelengths. <, concentrations less than the summed limits of quantitation.

estimated by HPLC/fluorescence (Table 11) at wavelengths chosen to detect the most important classes of compounds present in weathered PBCO, i.e., the alkylated naphtha- lenes and dibenzothiophenes that fluoresce at naphthalene wavelengths and the alkylated phenanthrenes that fluo- resce at phenanthrene wavelengths. Although the Alaskan sediments were collected more than 1 year after the spill and the PBCO they contained had weathered, concen- trations are reported in terms of equivalents of freshPBCO. No single weathered PBCO standard could be represen- tative of the many different degrees of weathering dis- played by the Alaskan sediments, because weathering is environmentally dependent and may be influenced by length of exposure to sun, water, microbial activity, and other factors in the environment.

To compare the semiquantitative results obtained by HPLC screening with the quantitative results from the detailed GUMS method, concentrations of individual ACs determined by GC/MS were summed by parent structure (Table 11): CO-C4 naphthalenes (CNPHs), Co-Cs di- benzothiophenes (CDBTs), and CO-C4 phenanthrenes (E-

PHNs). The summed ACs were then divided into two groups by their fluorescence characteristics-the CPHNs that fluoresce at phenanthrene wavelengths and the CNPHs + CDBTs that fluoresce at naphthalene wave- lengths. When the summed concentrations were below the limit of quantitation (preceded by <), an amount equal to half the limit of quantitation was included in the statistical analysis. The PBCO equivalents a t phenan- threne wavelengths from HPLC screening (Table 11) were highly correlated (r = 0.726, p I 0.0001) with the CPHNs from GUMS (Figure 2A). A similar correlation ( r = 0.731, p I O.OOO1) was found for the PBCO equivalents calculated at naphthalene wavelengths and the CNPHs + CDBTs (Figure 2B).

HPLC Chromatographic Patterns of Petroleum Products. Analytes eluted from the size-exclusion HPLC column according to molecular weight and size, with compounds of "largest volume" eluting first. The chro- matograms of several classes of petroleum products, i.e., crude oils and intermediate or heavy distillates, showed a distribution of fluorescent ACs by molecular size (Figure

702 Environ. Sci. Technol., Vol. 27, No. 4, 1993

v) 2 7 5

0 E 2 2 5

- 2'

0) 1 7 5 .

J 2 5

c

0 1 5

Y

2 1 2 5

r = 0.726,

1'

7 5

I I 1 I

5 5 0 5 1 1 5 2 2 5 3 3 5

PBCO equivalents (log1 0) from sediment screening (phenanthrene wavelengths)

3 251 t

0)

r= 0.731, p 5 0.0001 '

.5 0 5 1 1 5 2 2 5 3

PBCO equivalents (log1 0) from sediment screening (naphthalene wavelengths)

Figure 2. Correlations in sediments of (A) equivalents of Prudhoe Bay crude oil (PBCO) estimated by screening at phenanthrene wavelengths and sums from GC/MS of Co-C4 phenanthrenes (CPHNS) and (B) equivalents of PBCO estimated by screening at naphthalene wavelengths and Co-C4 naphthalenes (CNPHS) + Co-C3 dibenzothiiphenes (CDBTs) that fluoresce at naphthalene wavelengths.

3). The crude oils, includingPBC0 and weathered PBCO, showed only minor differences in their chromatographic patterns. For example, weathered PBCO and weathered Kuwaiti crude lacked a small portion of the lower molecular weight compounds in the AC fraction P8.2 min)- particularly apparent a t naphthalene wavelengths (Figure 3B). The intermediate (bp 185-345 "C) and heavy (bp > 345 "C) distillate fractions of petroleum contained nar- rower ranges of molecular sizes than did the crude oils, and as a result, their HPLC chromatograms were dissimilar to those of the crude oils (Figure 3). For example, diesel fuel and No. 2 fuel oil contained greater proportions of the lower molecular weight compounds and the marine lu- bricating oil had greater proportions of the higher mo- lecular weight compounds than were found in the crude oils (Figure 3).

HPLC Chromatographic Pattern of an Urban Sediment. The HPLC chromatogram of a sediment collected from Auke Bay, a site near Juneau, AK (Figure 31, showed high levels of ACs in screening analyses (phenanthrene equivalents, 17 000 f 3300 ng/g; n = 2). The chromatographic pattern from this sediment resem- bled those reported previously for urban sites (1). To substantiate the presence of urban-type pollutants, this

sediment was also analyzed by GC/MS. The sum of ACs (3500 f 770 ng/g, wet wt) was high in comparison to other urban sediments measured in our previous study (I). Many pyrogenic ACs (e.g., chrysene, benz[al anthracene, and benzo [a] pyrene), typically found in urban sediments, were present in substantial concentrations (sum of four- to six- ring ACs was 3300 ng/g) in the Auke Bay sediment.

HPLC Chromatographic Patterns of Sediment Extracts. The HPLC chromatograms of intertidal (0 m) sediment extracts from oiled sites in Prince William Sound, e.g., Herring Bay (Figure 4), Snug Harbor (Figure 4), and Bay of Isles (not shown), showed a molecular size distri- bution very similar to that of weathered PBCO. The intertidal sediments from Northwest Bay (Figure 41, Sleepy Bay (Figure 4), and Block Island (not shown)-heavily oiled sites that were treated with high-pressure, hot-water washes-exhibited patterns similar to that of weathered PBCO, but contained smaller proportions of some lower molecular weight compounds. In contrast, sediments from sites not known to have been oiled by the Exxon Valdez spill-MacLeod Harbor (20 m; Figure 41, Olsen Bay (100 m; Figure 4), and Port Fidalgo (100 m; not shown)-had chromatographic patterns that more closely approximated that of diesel fuel (or No. 2 fuel oil). The chromatogram of a subtidal sediment from Sleepy Bay (100 m; Figure 4) showed a somewhat wider molecular size range than did the unoiled sites, but contained a smaller proportion of the high molecular weight ACs than were found in PBCO.

Relative Concentrations of ACs in Sediments An- alyzed by GC/MS. In the samples of PBCO, weathered PBCO, and the 60 sediment extracts, concentrations of individual AC isomers determined by GC/MS were summed by groups of isomers (e.g., CZ naphthalenes). The isomer groups selected for graphing were those most characteristic of PBCO-the alkylated naphthalenes, phenanthrenes, and dibenzothiophenes. In particular, the degradation-resistant dibenzothiophenes that are found in relatively high proportion in PBCO and other North Slope crude oils are ideal marker compounds, because dibenzothiophenes are not found in substantial propor- tions in most other Alaskan (e.g., Cook Inlet crude) or continental US. crude oils (12-14). Another degradation- resistant isomer group, the C3 phenanthrenes, was chosen as the reference group and its concentration was set equal to 100%. Concentrations of the isomer groups were normalized to the C3 phenanthrenes and then graphed (Figure 5). The actual concentration of C3 phenanthrenes in each sample is indicated above the bar, e.g., 297 nglg in fresh PBCO (Figure 5A).

The relative concentrations of these groups of isomers can be compared among samples. For example, relative concentrations of the CO-C~ naphthalenes, CO-CZ phenan- threnes, and CO-C~ dibenzothiophenes were lower in weathered PBCO (Figure 5B) than in fresh PBCO (Figure 5A). The weathering of the more volatile ACs was also observed in several Prince William Sound sediments. For example, the intertidal sediments from Northwest Bay (Figure 5C), Herring Bay (Figure 5D), Snug Harbor (Figure 5E), and Sleepy Bay (Figure 5F) had proportions of petroleum-related ACs similar to each other and to weathered PBCO (Figure 5B), except for some variations in the proportions of alkylated naphthalenes. A substan- tially different profile of ACs was found in the sediments from MacLeod Harbor (Table 11; 40 m shown in Figure 5G) and Olsen Bay (Table 11; 100 m shown in Figure 5H).

Envlron. Sci. Technol., Vol. 27, No. 4, 1993 708

8.2 min

Phenanthrene wavelengths

Contaminated sedlmenl Auke Bey, Alaska

Diesel Fuel

No. 2 Fuel Oil

Fresh PBCO

Weathered PBCO

Fresh Kuwaiti crude

Weathered Kuwaiti crude

0.0 12.0

I 82

B

n Naphthalene wavelengths

Contamlnated sediment

'Urban"

i L No. 2 Fuel Oil

Intermediate distillates

A I

Marine Lubricating oil Heavy distillates

Crude oils

0.0 12.0

Flgure 3. HPLC/fluorescence chromatograms (molecular size distributions) of possible sources of contaminants found in the marine environment: a sediment from an "urban" estuary in Auke Bay, AK, lntermedlate and heavy petroleum distillates, and crude oils. Fluorescence was recorded at wavelengths where (A) the phenanthrenes (2601380 nm; phenanthrene wavelengths) and (6) the naphthalenes and dlbenzothlophenes (2901335 nm; naphthalene wavelengths) fluoresce. The retention time (8.2 mln) at which integration of the AC fraction was begun (for our HPLC system) is marked on the chromatograms. To facilitate visual comparlsons between chromatographic patterns of the Prince William Sound sedlments and possible contaminant sources, all the chromatograms within grouplngs In the flgures in this paper have been electronically adjusted to the same height.

The dibenzothiophenes were absent in these sediments, even though the proportions of alkylated naphthalenes and phenanthrenes were higher than in weathered PBCO. The profile of ACs in the 100-m Sleepy Bay sediment resembled those of MacLeod Harbor and Olsen Bay, except that the alkylated dibenzothiophenes were present.

Discussion

The degree of oiling in intertidal sediments reported by observers at sites in Prince William Sound (19) was in general agreement with the concentrations of petroleum- related ACs found in these sediments by HPLC screening

704 Environ. Sci. Technol., Vol. 27, No. 4, 1993

A Phenanthrene wavelengths I 8.2 rnln

Wealhered PBCO

Herrlna B a W m

Snug Harbor4 rn - Northwea B a y 4 rn

Sleepy B a y 4 rn

u

Macled Harbor--M q

Olsen B a r 100 rn

B Naphthalene wavelengths ,,I 8.2 rnin

Herring B a y 0 rn

Snua Harbor4 rn

Northwest Bay-0 rn

h 2 A L Diesel Fuel

MacLeod Harbor-20 rn

A Olsen Bay- 100 rn

S k e w B a r 1 00 rn

0.0 12.0 0.0 12.0

Figure 4. HPLWfluorescence chromatograms of intertidal sediments (0 m) from four oiled sites, Herring Bay, Snug Harbor, Northwest Bay, and Sleepy Bay; subtldal sediments from two UnOiled sites, MacLeod Harbor (20 m) and Olsen Bay (100 m); and a subtidal sediment from an oiled site, Sleepy Bay (100 m). These sediment samples, collected from sites in Prince William Sound after the Exxon VaMez oil splll. were screened for fluorescent aromatic compounds using the conditions listed in Figure 3.

or by GC/MS analyses; Le., the concentrations of ACs in intertidal sediments (Table 11) were consistent with the descriptions of oiling (Table I). However, several of the sites had a wide range in degree of oiling as reported by the observers (e.g., within Snug Harbor, intertidal oiling ranged from light to heavy). Although our intertidal sediment samples were collected from a relatively small area within a site, the extensive compositing of our samples should have minimized problems of nonhomogeneous distribution of the oil or small-scale patchiness within the collection area. In addition, the distribution of oil among the intertidal and the five subtidal depths within a site was highly variable (Table 11) and probably depended not only on the original magnitude of oiling but also on the redistribution of oil caused by cleanup actions and natural dispersive processes.

Concentrations of petroleum-related ACs estimated by HPLC screening of sediments collected after the Exxon Vuldez oil spill (Table 11) were highly correlated with concentrations of ACs in extracts of the same sediments determined by GC/MS (Figure 2). This correlation is in agreement with previous results showing a correlation

between HPLC screening and GUMS analyses of sedi- ments collected from urban sites (1). Thus, the utility of the rapid HPLC screening method has been extended to analyzing sediment samples for the ACs characteristic of crude oil, thereby helping to direct priorities for GC/MS analyses. As a result, the overall costs of the analyses have been reduced, while still providing the necessary detailed data in a timely fashion. Furthermore, HPLC screening analyses can be conducted "on-site", as recently demonstrated on board the NOAA R/V M t . Mitchell during the Arabian Gulf Project, to allow modification of the sampling strategy based on the screening results.

In addition to quantitative information, the size- exclusion chromatography provided information on the distribution of the ACs by molecular size. These chro- matographic patterns proved to be characteristic of classes of petroleum products, e.g., crude oils or various petroleum distillates (Figure 3). For example, the crude oils shown (PBCO, Kuwaiti, South Louisiana) exhibited similar, distinctive chromatographic patterns (Figure 3). The slight differences in the patterns of the weathered crudes compared to the fresh crude oils can be attributed to the

Environ. Scl. Technol., Vol. 27, No. 4, 1993 705

. . . . . . . . . $ 6 8 ' 8 8 . $ 6 8 8 8

3m B Weathered PBCO -

E

8- I

b'" 0

$ 6 8 8 3 ' $ 6 8 8 3 . g s c t a

z I C Northwest Bay O m I

E

3po MacLeod Harbor I

Olsen Bay

Sleepy Bay

h 2m

6 5

9 'O0 0

- I '

Flgure 5. Relative concentrations of C0-C4 naphthalenes (NPH), Co-C4 phenanthrenes (PHN), and Co-C3 dibenzothiophenes (DBT) in (A) fresh Prudhoe Bay crude oil (PBCO), (B) weathered PBCO, (C-F) intertidal (0 m) sediments from oiled sites, (G and H) subtidal sediments from unoiled sites, and (I) a subtidal sediment from an oiled site. Concentrations of ACs were determined by GC/MS, and the summed groups of isomers were normalized to the CB phenanthrenes. The number above the C3 phenanthrene bar is their summed concentration (A and B, ng/g of 011; C-I, ng/g of sediment, wet wt).

loss of some lower molecular weight ACs upon weathering (1). In contrast, the HPLC patterns from these crude oils were very dissimilar to those from the distillate fractions (e.g., diesel fuel or marine lubricating oil; Figure 3) or to that from a sediment from an "urban" estuary contam- inated primarily with pyrogenic ACs (Auke Bay, AK; see Results and Figure 3). However, because visual pattern recognition is highly subjective, GC/MS data are essential for confirmation of contaminant source.

In sediments analyzed for ACs by GUMS, the relative concentrations of groups of isomeric ACs, e.g., Cp di- benzothiophenes or CB phenanthrenes, are best illustrated in graphs similar to those used by Atlas, Boehm, and Calder (16) and Boehm, Fiest, and Elskus (17). These graphs (Figure 5) also illustrate the changes in the proportions of ACs in oiled sediments that are due to the degree of weathering of the oil. In the present study, for example, the proportions of the CO-C~ naphthalenes are much smaller and proportions of the highly alkylated (C3-c~) naphthalenes, phenanthrenes, and dibenzothiophenes are larger in weathered PBCO than in fresh PBCO (Figure 5A and B). In particular, relatively high proportions of the marker compounds for North Slope crude oils, the alkylated dibenzothiophenes, remain after the oil has

weathered. Similar findings were reported for the Amoco Cadiz reference oil compared to weathered oil extracted from sediments (16,17). Therefore, GUMS can be used to confirm the presence of PBCO in Prince William Sound sediments by identifying characteristic proportions of petroleum-related ACs in these samples.

An HPLC chromatographic pattern similar to that of weathered PBCO was identified in many Prince William Sound sediments. For example, HPLC chromatograms of the intertidal sediments from Herring Bay (Figure 4), Snug Harbor (Figure 41, and Bay of Isles (not shown) were very similar to those from weathered PBCO, providing evidence for that source of contamination. The HPLC chromatographic pattern in the intertidal sediments from Northwest Bay (Figure 41, Sleepy Bay (Figure 41, and Block Island (not shown) were similar to each other, but were more weathered than that of weathered PBCO, as indicated by the absence of some lower molecular weight compounds. The intertidal areas of these sites had been subjected to the same severe treatment to remove the oil (category 31, so their chromatograms could be expected to show similar anomalies. GC/MS analysis provided confirmation of PBCO contamination in these sediments through iden- tification of characteristic proportions of petroleum-related

706 Environ. Sci. Technol., Vol. 27, No. 4, 1993

ACs, including the dibenzothiophene marker compounds (Table 111). For example, the GUMS patterns of ACs in the intertidal sediments from Northwest Bay, Herring Bay, Snug Harbor, and Sleepy Bay were generally similar to each other (Figure 5) and to that of weathered PBCO. A combination of natural weathering and the cleanup treatments to which these sites were subjected explains the various degrees of weathering in these sediments.

MacLeod Harbor and Olsen Bay are not known to have been oiled by the Exxon Valdez spill, yet both the HPLC screening and GUMS analyses found low to moderate concentrations of ACs in these sediments (Table 11). Because of a greater abundance of the lower molecular weight compounds, the HPLC chromatographic patterns from sediment extracts from all depths in MacLeod Harbor (40 m shown in Figure 4) and Olsen Bay (100 m shown in Figure 4) were most similar to that of diesel fuel (Figure 4). Furthermore, alkylated naphthalenes and phenan- threnes were the most abundant ACs found by GUMS analyses-concentrations of alkylated dibenzothiophenes were low in these sediments (Figure 5). Although the HPLC patterns most closely resembled those of diesel fuel-a possible source due to vessel traffic from fishing and pleasure boats-other sources of ACs, including biogenic production, cannot be eliminated. However, the GUMS results are consistent only with adiesel fuel refined from a crude oil low in dibenzothiophenes (e.g., Cook Inlet crude).

HPLC patterns and GC/MS profiles resembling those of diesel fuel were also found in some subtidal sediments (20 and 40 m; Table 11) from Chenega Island, a site that was oiled in the intertidal areas but was not treated for cleanup. Apparently, the crude oil that fouled the beaches of Chenega Island was not generally transferred to the deeper sediments. In contrast, oil appears to have been transferred to the subtidal sediments of Sleepy Bay (Table 111, possibly due to the severe cleanup procedures employed at that site. Interestingly, the 100-m subtidal sediment from Sleepy Bay showed evidence of a background contribution of ACs from another source. Although the molecular weight distribution (Figure 4) and the relative concentrations of AC isomer groups from GUMS (Figure 511, in this sediment were consistent with a mixture of crude oil and diesel fuel, other sources could not be eliminated. Thus, the HPLC and GC/MS results for this subtidal sediment from Sleepy Bay demonstrated the difficulty of identifying low concentrations of PBCO at sites having a background of other ACs.

HPLC screening appears to be able to detect lower concentrations of petroleum-related ACs in samples contaminated by PBCO than does analysis by GC/MS, largely because screening measures the total fluorescence of oil components while GC/MS measures individual ACs. For example, concentrations of the isomer groups in weathered PBCO ranged from <1 to -130 pg/g (Figure 5B). The summed concentration of isomers in the C2 phenanthrene group totaled -40 pg/g (Figure 5B), and for this example, we will assume that the entire amount is attributable to a single isomer. In a hypothetical sample of sediment containing only 1 pg/g (ppm) total weathered PBCO, the concentration of the C2 phenanthrene isomer would be 0.040 ng/g of sediment. Because the limits of detection for individual ACs by GC/MS were -1 ng/g of sediment (wet wt), the CZ phenanthrene isomer would not be detectable by GUMS. The PBCO concentrations in

sediments would have to be increased at least 25-fold (to 25 pglg) to raise the concentration of the CZ phenanthrene isomer to detectable levels in this hypothetical sediment. Actually, we find approximately 12 isomers in the CZ phenanthrene isomer group. Therefore, depending upon the relative contribution of each isomer, a concentration of PBCO higher than 25 pg/g would have to be present in a sediment to detect these isomers by GC/MS. In contrast, the limits of detection for the HPLC screening method were <1 pg/g PBCO and could be lowered by a factor of 10-100 by further concentrating the extract.

In summary, analyses of 60 sediments from Prince William Sound were performed by HPLC screening and also by GC/MS to compare the methods for measuring PBCO contamination in sediments contaminated by a major oil spill. Concentrations of ACs measured by sediment screening were highly correlated with the sums of ACs determined by GC/MS, thus validating sediment screening as a tool for estimating concentrations of PBCO- related ACs. In sediments in which PBCO was the predominant source of ACs, we found HPLC screening was able to detect lower concentrations of PBCO than could be detected by GC/MS, largely because the HPLC method measures the total fluorescence of oil components whereas GUMS measures individual ACs. Moreover, differences in HPLC chromatographic patterns suggested different sources of contamination, e.g., crude oil or diesel fuel. Accordingly, the source of contamination was confirmed from GC/MS results by comparing relative concentrations of petroleum-related ACs in the sediments to those from possible sources. Thus, the HPLC screening method has an important dual role in evaluating anthro- pogenic contamination in sediments. First, sediments containing AC contaminants can be rapidly ranked by degree of contamination, and second, HPLC chromato- graphic patterns can provide a basis for suggesting possible contaminant sources. As a result, expensive GC/MS resources can be effectively allocated.

Acknowledgments

We thank Tom Ruszala, captain of the NOAA R/V Dauidson, along with the other officers and crew, for their able support in the field during sample collection. Chuck O’Clair, Terri Stinnett, and Anthony Chan of the National Marine Fisheries Service, Auke Bay Laboratory, Michael Hilley of the Alaska Department of Environmental Conservation, and Tom Yeager of the University of Alaska Institute of Water Resources provided assistance in collecting and processing sediment samples. We thank Robert Clark for collecting the sample of weathered Kuwaiti crude oil. We appreciate the technical assistance or advice of Donald Brown, Catherine Wigren, Jennie Bolton, Kristin Blair, and Sue Pierce. This work was carried out with support from projects Air/Water 6 and Technical Services 1 under the State/Federal Natural Resources Damage Assessment for the Exxon Valdez oil spill. Mention of trade names is for information only and does not constitute endorsement by the US. Department of Commerce.

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Received for review August 3, 1992. Revised manuscript re- ceived October 26, 1992. Accepted December 16, 1992.

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