of the NaPCP stock solution with filtered seawater. The concentrations of NaPCP tested 

 were: 10, 32, 56, 100, and 180 ug/L. 



Salinity, dissolved oxygen, and pH levels initially were adjusted in each container to 30 

 ppt, 8.2 mg/L and 7.8, respectively. These parameters were measured again in each container 

 at the termination of the toxicity test. 



Sediment Elutriate Toxicity Test with Embryos of the Urchin Strongulocentrotus 

 purpuratus 



Elutriates of samples from all 15 stations were tested for effects on development and 

 echinochrome pigment content of the embryos of the purple sea urchin (Strongylocentrotus 

 purpuratus). Echinochrome pigment synthesis appears to be affected by abnormal embryonic 

 development and this end-point has been shown to be sensitive, less variable, and quicker 

 than the morphological examinations (Bay et al, 1983). Embryos from one station at each of 

 the sites also were examined microscopically for the presence of cytologic and cytogenetic 

 (mitotic) abnormalities. In addition, concurrent testing of elutriates from one station at each 

 site was conducted to determine egg fertilization success. Additional 48-h tests on elutriates 

 from these five stations were conducted with embryos from two additional urchin species, the 

 white urchin (Lytechinus pictus) and the green urchin (Strongylocentrotus drobachiensis). Most 

 aspects of this toxicity test have been developed and applied in analyses of primarily water 

 or effluent samples (Oshida et al, 1981; Dinnel et al, 1982; Dinnel and Stober, 1987). 



Elutriate preparation. For each replicate tested, a 70-mL aliquot of sediment was washed 

 through a 1-mm mesh screen to remove large organisms, tubes, and debris. A total of 280 mL 

 of laboratory seawater (collected off Redondo Beach, California) was added to the sample 

 (including screening water) to produce a sample dilution of 1:4 (v/v). The sediment/ water 

 mixture then was placed in a 400-mL glass beaker and stirred overnight at 17°C with a 60- 

 rpm teflon/glass paddle. Each sample was allowed to settle for 60 min after stirring. The 

 supernatant was then poured into centrifuge bottles and centrifuged at 2,000 times gravity (G) 

 for 5 min to precipitate suspended particulates. The supernatant was carefully decanted and 

 returned to its respective 400-mL beaker. From this volume, a 10-mL aliquot was removed and 

 used in the sperm cell toxicity testing and a 220-mL aliquot was used in tests of embryos for 

 the other end-points. A total of 220 mL of the elutriates was used in the 400-mL beakers for 

 the embryo exposures. 



Animal collection. Intertidal adult urchins were collected from Point Dume, in northern 

 Santa Monica Bay, California. Release of the eggs and sperm from gravid sea urchins was 

 induced by injection of 0.5 mL of 0.5 M KC1 into the coelom of each individual. Eggs were 

 shed directly into beakers of chilled seawater and washed twice with seawater before use. 

 Sperm were collected with a minimum of seawater (dry condition) and refrigerated until 

 used. 



Test procedures. The sperm cell test followed the procedures of Dinnel et al (1987). It was 

 initiated by adding 0.1 mL of sperm stock solution to each 10-mL elutriate sample. After 60 

 min exposure, 2,000 eggs were added to each elutriate sample. Each sample was allowed 20 

 min for fertilization to occur and then preserved with formalin for later examination. The 

 formalized eggs were examined using light microscopy (100 times) and a Sedgewick-Rafter 

 counting chamber. The numbers of fertilized and unfertilized eggs in an aliquot of each sperm 

 exposure sample were counted in which the presence of a well-defined fertilization membrane 

 around the egg was the criterion defining successful fertilization. 



The embryo toxicity test methods were modified from the procedures of Oshida et al. 

 (1981). Exposure of the embryos to the elutriates was initiated by inoculating each 220-mL 

 sample with 7,500 fertilized eggs. Stirrers then were fitted to each beaker, and the sample 

 with its developing eggs was cultured for 48 hours at 17°C. After 48 hours, a 10-mL subsample 

 was removed from each beaker and preserved for later analysis of percentage normal 

 development. A duplicate sample of embryos was taken from some beakers for the 



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