of Available Bioassays for Brevetoxins in Fish and Shellfish:
Correlation with Cell Concentrations in the Water Column
August 30, 1994
National Marine Fisheries Service, 9721 Executive Center
Drive, North Koger Building, St. Petersburg, FL 33702.
PHONE: (813) 570-5324
for detecting brevetoxins in marine seafood were compared,
and correlated against analytical HPLC procedures and
the AOAC i.p. mouse bioassay. Radioligand synaptosome
binding assays, radioimmunoassays, enzyme-linked immunoassays,
Gambusia affinis fish bioassays, and electrophysiology
were utilized as potential substitutes for mouse bioassay.
Assays were performed on both natural field-collected
specimens before, during, and after a red tide, from laboratory-induced
toxic specimens, and from homogenates of seafood sources
artificially "spiked" with brevetoxins.
P. brevis cells or toxin in seawater could be
detected using any of the assays utilized. ELISA
and RIA were most sensitive in identifying low amounts
of brevetoxin. Several new formats were explored,
specifically designed for water use, for use as a "presumptive"
assay, or for the rigors of routine tissue testing Synaptosome
binding assays and electrophysiology provided the most
sensitive methods of identifying very potent toxins.
Gambusia affinis bioassay proved most useful
for rapid screening of toxic fractions, while HPLC methods
proved relatively insensitive to toxins with little carbon-carbon
unsaturation, and gave no indication of potential toxicity.
Biological matrices of shellfish or fish tissue proved
problematic for ELISA and HPLC, while RIA and mouse bioassay
could detect toxin in these matrices. Two unexpected results
were the identification of a new hemibrevetoxin (uncharacterized
structurally, Btx-H) from Mote Marine Laboratory cultures,
and the discover of irreversible brevetoxin binding to
fish tissues. Spectral results are presented for
the new hemibrevetoxin, and data is presented for the
latter irreversible binding. The ultimate disposition
of brevetoxin in fish tissues is not known. That
is to say, it is not known if toxin is metabolized to
non-detectable forms or if it is sequestered in tissues
to non-extractable forms.