FISHERY BULLETIN: VOL. 81, NO. 3 



TABLE 3. — Percent of California grunion yolk-sac larvae 

 with developmental abnormalities when exposed to in- 

 creasing concentrations of BaP. 



1 x±SD(n) 



2 No eggs hatched in one replicate of these series, therefore n = 



Notochord Length 



Notochord lengths (NL) of embryos and yolk-sac 

 larvae are shown in Figure 4. Larvae which hatched 

 after exposure to initial levels of BaP up to 361 ppb 

 were not significantly different from controls in 

 length (P>0.05). Shorter notochord lengths were ob- 

 served in embryos exposed to BaP concentrations 

 >7 ppb (P<0.05). Upon hatching, the mean 

 notochord length of the control larval group was 5.8 

 mm, while embryos of the same age exposed to 24 

 ppb BaP averaged 4.0 mm NL. 



containing 297 ppb BaP, 42% of the yolk-sac larvae 

 were abnormal. 



Gross abnormalities observed in yolk-sac larvae ex- 

 posed to 24 ppb BaP included lateral foldings of the 

 posterior one-fourth of the tail, absence of caudal fin 

 folds, and hemorrhagic lesions or congested vascula- 

 ture in the caudal region (Table 4). 



In contrast, development of embryos exposed to 24- 

 361 ppb BaP for 14 d was retarded and resembled 

 that of the normal embryo at 2.5 and 5.5 d of age (Fig. 

 3a, b) (David 1939). Abnormalities included 1) 

 malformed tails with congested vessels or 

 hemorrhage, 2) sporadic heartbeat resulting in inter- 

 mittent blood flow, 3) head displacement in relation 

 to the yolk sac, and 4) lack of melanophores near the 

 lateral line above the intestinal tract. 



Embryos exposed to 869 ppb BaP for 14 d 

 resembled normal embryos at 1.2-2.5 d of age (David 

 1939). However, affected embryos had a lateral cur- 

 vature midbody with occasional melanophores found 

 on the trunk. In general, those embryos with shorter 

 notochord lengths were observed to have yolk sacs 

 much larger than those of the controls. 



TABLE 4. — Abnormalities observed in California grunion yolk-sac 

 larvae and embryos exposed to increasing concentrations of BaP. 



BaP 



exposure 



(PPb) 



Abnormality 



Yolk-sac 

 larvae 



Embryos 



24 Lateral folding of posterior fourth of tail 



Absence of caudal finfold. 

 Congested vasculature on caudal region. 



24-361 14-d-old embryo retarded in growth (resembled nor- 



mal embryo growth at 1 .5-5.5 d of age). 

 Sporadic heart beat. 



Displacement of head in relation to yolk sac. 

 Absence of melanophores near lateral lines. 

 Absence of lens formation. 

 Lesions as in larvae (above). 



869 14-d-old embryo retarded in growth (resembled nor- 



mal embryo growth at 1 .2-2.5 d of age). 

 Lateral curvature midbody. 



Absence of melanophores (except in trunk region). 

 Unused yolk sac. 

 Lesions as in larvae (above). 



DISCUSSION 



Toxic and teratogenic effects of the carcinogen BaP 

 on developing fish were studied by incubating em- 

 bryonic stages of California grunion to increasing 

 concentrations of BaP. Eggs were hatched in 

 seawater alone, although California grunion eggs are 

 normally incubated in sand. David (1939) concluded 

 that there was probably no special adaptation of em- 

 bryo metabolism to sand incubation, and speculated 

 that spawning in the sand was a mechanism to protect 

 eggs from predation. Incubation in seawater without 

 sand permitted optimal observation of embryonic 

 development with minimum disturbance. Also, ex- 

 clusion of sand reduced the possibility of fungal and 

 bacterial overgrowth and eliminated any possible 

 contamination and influence by sand and/or sand- 

 absorbed materials, as well as eliminating a large, 

 potentially adsorptive surface for BaP. Preliminary 

 trials resulted in a hatching rate of 90-100 ( /< in our 

 laboratory, comparable to values previously report- 

 ed (David 1939; Hubbs 1965). 



Because of the low solubility of BaP in seawater, it 

 was necessary to select a solubilizing or dispersing 

 agent to create a uniform distribution (Davis et al. 

 1942; Wilk and Schwab 1968; Neff 1979). Such an 

 agent could affect early development by acting in an 

 additive or synergistic manner with BaP. Solvents 

 examined in preliminary studies were benzene, 

 Triton X-100, trioctanoin, and acetone, all of which 

 caused observable alterations except for acetone in 

 low concentrations. Triton X-100 and trioctanoin 

 proved lethal, whereas the carcinogen benzene in- 

 duced tail malformations. The validity of utilizing a 

 solvent to distribute high levels of BaP in seawater 

 can be compared with the not-uncommon situation in 

 nature whereby lipophilic compounds are solubilized 

 by contaminating substances such as detergents or 

 oils. Also, certain solvents in which BaP is soluble, 

 such as benzene, toluene, and xylene, are present in 

 varying quantities in crude and refined oil. However, 



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