In addition to the BaP-treated flake food, chemical analysis was
done in F1 fertilized eggs to assess potential BaP transfer into the
embryos. Approximately 110 F1 fertilized eggs (8 hpf) collected on
days 20 and 21 were stored at
−80 ◦C until chemical analysis. Eggs
from the four treatment groups in triplicate were extracted 3 times
with 3 mL of methylene chloride each time. A known concentration
of the surrogate standard, benzo[a]pyrene-d12, was added to each
sample to yield a final concentration of 0.2 g/mL. Samples were
vortexed for 15 s and centrifuged for 7 min at 3000 rpm (1509
×
g)
and passed over an alumina and sodium sulfate column to remove
fat and water from the extract. Samples were then blown to dryness
with N2 and brought back up with a known volume of hexane.
A known concentration of the internal standard, fluorene-d10, was
added to each sample to yield a final concentration of 0.2 g/mL to
determine extraction efficiency. Samples were run on the GC/MS
under the selected ion mode to quantitate the concentration of
BaP in each sample. BaP was not detected in the eggs despite good
recovery concentrations (50–105%). This could be due to multiple
reasons including but not limited to insufficient number of eggs,
small amount of BaP deposition in the eggs, or BaP being rapidly
metabolized by the mothers.