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<br />Toxicity of Shale Oil to Fish and Food Chain Organisms
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<br />tions at three different times during all warmwater and coldwater
<br />studies. During the 96-hr tests, 500-ml samples were pumped
<br />from below the surface of each of the six tanks at I, 20, and 90
<br />hr. Each 500-ml sample was extracted three times with 30-ml
<br />portions of trichlorotrifluoroethane in I,OOO-ml separatory
<br />funnels. Extract volume was reduced to 10 ml under a stream of
<br />nitrogen and analyzed by infrared spectroscopy (Simard et al.
<br />1952) to determine the total oil content. Actual concentration
<br />reported was a mean of these three determinations.
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<br />Toxicity
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<br />Where possible, the standard practices for performing acute tox-
<br />icity tests were followed (ASTM 1980). At the start of each
<br />warmwater test, 20 Colorado squawfish and either 30 or 50
<br />fathead minnows were placed in each test solution, For cold-
<br />WIlter tests, 20 cutthroat trout and 3 colonized plate samplers
<br />were placed in each test solution, Dead fish were removed daily,
<br />and the concentration lethal to 50% of the organisms (LC50) and
<br />95% confidence interval were calculated for each species at 48
<br />and 96 hr (Litchfield and Wilcoxon 1949). After 96 hr of expo-
<br />lUre, live and attached invertebrates were removed from the
<br />plate samplers and stored in 70% ethyl alcohol. Organisms from
<br />each plate sampler were identified and enumerated by a private
<br />~ratory (Susswasser, P,O. Box 1255, Paso Robles, CA 93446).
<br />The three colonized plate samplers from each test solution were
<br />combined to give one value for total number of organisms. total
<br />lIImber of taxa, and the Shannon-Weaver Index (Pielou 1975).
<br />the Shannon-Weaver or diversity index was based on the lowest
<br />taXonomic classification used (either genus or species) and
<br />.'Ould be properly referred to as diversity of taxa,
<br />At the end of each 96-hr warm water test, the influence of sub-
<br />ithal exposure was measured during swimming and predator-
<br />prey tests of survivors from the three highest exposure concen-
<br />ntior.s that resulted in no mortalities, and from the control.
<br />Swimming ability of Colorado squawfish was measured in a
<br />tlImina chamber modified from Howard (1975). Individual fish
<br />were placed in the chamber, acclimated for 5 min to a water
<br />wIocity of9,2 cm/s, and then subjected to incremental increases
<br />tl6,7 cm/s in water velocity every 3 min until they failed to
<br />prim. The fish were weighed and measured for total body
<br />.. Swimming capacity was measured for each fish by dura-
<br />IDa of activity or total swimming time in seconds and by body
<br />lntcths per seconds (BLlS) as calculated by using the equation:
<br />
<br />.1;.:
<br />
<br />:r..
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<br />A + (B/180 * 6.2)
<br />BLlS =
<br />TL
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<br />*rt A is the water velocity before the failure interval, B is the
<br />..wming time during the failure interval, and TL is the total
<br />. of the fish. Both time in seconds and BLlS were analyzed
<br />.,. multiple means comparison test and least significant differ-
<br />au (Snedecor and Cochran 1967).
<br />Experiments on the predatory success of squawfish were con-
<br />flicted in 12 30-L aquaria (40 x 29 x 25 cm) with a gravel sub-
<br />_e and clusters of artificial Cabomba planted at each end.
<br />tab aquarium contained 20 L of warm water and received new
<br />IrIIer at the rate of 500 ml/min, Illumination was provided by
<br />~scent lights positioned 90 cm over each aquarium. Unex-
<br />lIIIIed fathead minnows were the prey species and either 17,23,
<br />.25 were put in each aquarium 24 hr before the experiment was
<br />lilted. The number added was constant within experiments but
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<br />varied among experiments, depending on availability. The test
<br />was begun by transferring three randomly selected squawfish
<br />from one of the three exposure concentrations or the control to a
<br />"predator success" aquarium. This procedure was repeated 3
<br />times per test solution (total of 12 tests per experiment). The
<br />experiment was terminated when 40 to 60% of the prey fish in
<br />the control tests had been captured. Predator success was mea-
<br />sured by percentage of prey captured, and comparisons were
<br />made between exposure groups and controls by using a binomial
<br />chi-square test (Snedecor and Cochran 1967).
<br />"Prey success" experiments examined the ability of exposed
<br />fathead minnows to escape capture by unexposed squawfish and
<br />were conducted in three 27-L aquaria (45 x 30 x 20 cm) con-
<br />taining 24 L of warm water and receiving 2 L of new water at
<br />5-min intervals. Other conditions were similar to those for the
<br />"predator success" aquaria, except that each prey success
<br />aquaria contained three unexposed squawfish predators
<br />(average weight, 10.5 g) maintained on a diet of fathead
<br />minnows. Predators were not fed for 4 days before the test to
<br />insure their being hungry. At 36 hr before the oil exposures,
<br />fathead minnows to be used were anesthetized with 2% tricaine
<br />methanosulfate, and the left base of the dorsal fin was injected
<br />with a dot-sized (I mm) mark of red, orange, yellow, or blue
<br />latex dye to allow differentiation among individuals from dif-
<br />ferent test solutions. The dye marks were randomized among
<br />solutions and did not affect the sensitivity of the fish to the oils
<br />or their vulnerability to predation. After the 96-hr exposure, 10
<br />fathead minnows from each of the four selected test solutions
<br />were added to each of the three prey success aquaria for a total
<br />of 40 prey fish per aquarium. Prey were initially separated from
<br />the predator squawfish by a screen, which was removed after 2
<br />hr. Prey were counted at hourly intervals until about 50% had
<br />been eaten; all fish were then removed and the surviving fathead
<br />minnows in each test solution were identified and counted. Prey
<br />success was based on percentage of prey that escaped, and a
<br />binomial chi-square test (Snedecor and Cochran 1967) was used
<br />to compare exposure groups and controls.
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<br />Results
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<br />The aromatic fraction represented 70% or more of
<br />the total compounds identified in the WSF of the
<br />crude shale oils; Geokinetics, Tosco, and Paraho
<br />(Table 1). In contrast, the aromatic fraction com-
<br />posed only 30-40% of the total WSF of the up-
<br />graded and refined products, Paraho HDT and
<br />Paraho JP-4. The components in the WSF of these
<br />two products were dominated by aliphatics, Almost
<br />all of the aromatics in Paraho HDT and JP-4 were
<br />monoaromatics, whereas heterocyclics (ketones,
<br />pyridines, quinolines, etc) dominated in the crude
<br />shale oil WSFs. Heterocyclics were not detected in
<br />the refined and upgraded products,
<br />Actual average oil concentrations were 5.6 mg/L
<br />in the highest exposure solution of the three crude
<br />oils, as compared with 2.8 mg/L in the high expo-
<br />sure solutions of the hydrotreated and refined oils.
<br />The higher test concentrations achieved with the
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