234 
Fishery Bulletin 95(2), 1997 
nations estimated potential rather than actual fecun- 
dity. However, we chose to collect animals at this ear- 
lier stage of development to ensure that specimens came 
from resident subpopulations at sites with known sedi- 
ment contaminant concentrations. This selection would 
not have been possible if animals had been collected on 
their spawning grounds. 
Fecundity determinations were carried out on 5 to 
10 animals from each site at each sampling time. 
Fish sampled in 1986-87 were collected as part of a 
study on gonadal development in English sole 
(Johnson et al., 1988); ovary samples were preserved 
for fecundity determination and archived, but not 
analyzed, at that time. Fish collected in 1989-90 were 
sampled specifically for fecundity determination. 
Fish were caught by otter trawl in 5-min tows and 
held in aerated saltwater in holding tanks on the 
deck of the research vessel until they could be pro- 
cessed. Within an hour of capture, fish were weighed 
and measured. From each animal, a 1-mL blood 
sample was collected with a heparinized syringe from 
the caudal vessel. Blood samples for measurement 
of plasma estradiol concentrations were centrifuged 
at 3,000 xg, and the plasma was stored at -20°C. 
Fish were sacrificed by severing the spinal cord. 
Ovaries from vitellogenic females were removed and 
weighed; one ovary was slit longitudinally and pre- 
served in modified Gilson’s fluid (Simpson, 1951) for 
later determination of fecundity and egg weight. 
Ovarian tissues for histological examination were 
preserved in Davidsons’ fixative (Mahoney, 1973). 
Additionally, tissue samples for determination of PCB 
concentrations were collected from the liver and 
ovary and stored at -20°C. Bile for measurement of 
fluorescent aromatic compounds (FAC’s) was col- 
lected and stored at -0°C. 
Analysis of samples 
Ovaries collected for histology were embedded in 
paraffin, stained with hematoxylin and eosin (Luna, 
1968), and examined microscopically to confirm their 
developmental stage and to record ovarian atresia 
and related lesions by using criteria outlined in 
Hunter and Macewicz (1985b) and Johnson et al. 
( 1991). Ovarian lesion severity was ranked on a sub- 
jective scale of 1 to 7, with 1 being minimal and 7 
being severe. 
Fecundity was determined by using the gravimet- 
ric method described by Bagenal and Braum (1971). 
Ovaries were preserved in Gilson’s fluid for at least 
3 months to allow eggs to harden and ovarian con- 
nective tissue to disintegrate. Preserved eggs were 
washed with water, filtered to separate them from 
residual ovarian connective tissue fragments, and 
dried at 60°C for 24 hours. All eggs were weighed, 
and then 3 subsamples of 200 eggs each were 
weighed. Fecundity, relative fecundity, and reproduc- 
tive rate were subsequently determined by using the 
formulas below: 
>• (total weight of eggs) (#of eggs in subsample)] 
Fecundity = - 
(mean weight of eggs in subsample) 
Relative fecundity = (Fecundity/gutted body weight 
(g)]; and 
Reproductive rate (g of eggs/year) = [Fecundity x egg 
weight in (g)]. 
Additionally, gonadosomatic index (GSI), hepato- 
somatic index (HSI), and condition factor were cal- 
culated as follows: 
GSI = 
ovary weight (g) 
gutted body weight (g) 
x 100 
HSI = 
liver weight (g) 
gutted body weight (g) 
x 100 
Condition factor = gutted body weight (g)/length 3 (cm). 
Levels of fluorescent aromatic compounds (FAC’s) 
in bile were measured according to the method of 
Krahn et al. (1987), which provides a semiquanti- 
tative determination of the concentrations of metabo- 
lites of PAH’s (Krahn et al., 1993). Bile sampled from 
fish was injected directly into a Spectra-Physics 
Model 8800 high performance liquid chromatograph 
(HPLC ) equipped with a Phenomenex reversed-phase 
C18 analytical column. The polar analytes (prima- 
rily metabolites of AH’s) in bile were eluted with a 
linear gradient from 100% water containing 5 mL of 
acetic acid/L to 100% methanol and monitored by two 
fluorescence detectors connected in series. Fluores- 
cence of metabolites was measured at two wave- 
lengths: 290/335 nm, where metabolites of naphtha- 
lene (NPH ) and related two-ring aromatic compounds 
from petroleum fuels fluoresce; and 380/480 nm, 
where metabolites of benzo[a]pyrene (BaP) and re- 
lated multi-ring AH’s from combustion sources fluo- 
resce (Krahn et al., 1987). Levels of biliary FAC’s were 
reported as equivalents of known concentrations of BaP 
or NPH standards on the basis of biliary protein be- 
cause recent studies (Collier and Varanasi, 1991) have 
shown that such normalization can account for varia- 
tion in FAC levels associated with the feeding status of 
