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Biotransformation in Fishes 205

(Gaworecki et al., unpublished data). In addition, a polyclonal antibody that was generated against
channel catfish phenol-type SULT did not recognize any SULT protein in mummichog samples. Despite
the low expression of phenol-type SULT in mummichog from clean reference sites, animals from the
Atlantic Wood site in the Elizabeth River in Virginia, a site heavily contaminated with PAHs and other
contaminants, showed significantly increased SULT activity (Gaworecki et al., 2004). Treatment with
single doses of 3-methylcholanthrene or benzo(a)pyrene failed to induce the protein; thus, contrary to
AhR-mediated CYP1A and UGT induction, SULT upregulation does not appear to be linked to AhR
activation. This is not unusual, as GST induction also requires chronic exposure (James et al., 1979).

Inhibition of SULT
Sulfotransferase is subject to inhibition from a variety of sources. In vitro, PAP derived from the
cosubstrate or sulfonate donor molecule PAPS inhibits SULT activity. Many bulky substrates inhibit
their own sulfonation; for example, in catfish intestinal cytosol 3-hydroxybenzo(a)pyrene inhibited its
own metabolism at concentrations above 0.5 µM (Tong and James, 2000). The nonsubstrate β-naph-
thoflavone inhibited SULT activity with 3-, 7-, and 9-hydroxybenzo(a)pyrene in channel catfish intestinal
and hepatic cytosol, with an IC of 48.0 ± 3.0 µM (James et al., 1997). A natural flavonoid, quercetin,
50
was a potent inhibitor of sulfation of 3-, 7-, and 9-hydroxybenzo(a)pyrene in channel catfish intestinal
cytosol, with an IC of less than 1 µM (van den Hurk and James, 2000). Although pentachlorophenol
50
was reported to inhibit phenol sulfation in mammals, it was a poor inhibitor of the sulfonation of
hydroxylated benzo(a)pyrenes in catfish and had an IC of about 100 µM (van den Hurk and James,
50
2000). Recent studies showed that several hydroxylated PCBs inhibited estrogen SULT in mammalian
systems with nanomolar IC values. Studies of the effects of hydroxylated PCB on the sulfonation of
50
hydroxylated benzo(a)pyrenes in catfish intestinal cytosol showed that several of these PCB metabolites
inhibited SULT, with low to high micromolar IC values (van den Hurk et al., 2002). Several metal
50
cations, including mercury, cobalt, zinc, cadmium, copper, and lead, were shown to inhibit zebrafish
SULT, although quite high concentrations (5 mM) were tested (Sugahara et al., 2003c).
Sulfonation of thyroid hormones was studied in liver cytosol from rainbow trout (Oncorhynchus
mykiss) (Finnson and Eales, 1998). Based on optimum pH, thermal stability, and specific inhibitors, the
investigators concluded that it is most likely that only one isozyme form is responsible for the sulfonation
of thyroid hormones in trout, in contrast to the multiple isozyme forms found in mammals. No information
is available yet on the inhibiting effects of hydroxylated PCBs on thyroid hormone sulfonation, as has
been reported for mammalian systems (Schuur et al., 1998).

Activation of SULT
2+
In vitro, SULT activity with some substrates can be increased by the addition of Mg and others by
incorporation of BSA into the assay vial. This phenomenon was also observed with the isolated catfish
SULT (Tong and James, 2000) and zebrafish SULT (Sugahara et al., 2003a). The in vivo significance
of this activation is unclear (Konishi-Imamura et al., 1995).

Amino Acid Conjugation

Overview
A relatively little studied pathway of phase II metabolism is the conjugation of carboxylic acids with
amino acids. This pathway results in the formation of an amide bond between the carboxylate and the
amino group of the amino acid (Figure 4.20). The resulting conjugate is an anion at physiological pH
and is generally a better substrate for organic anion transporter proteins in kidney than the unconjugated
xenobiotic; thus, amino acid conjugation facilitates the excretion of xenobiotic carboxylic acids (James
and Pritchard, 1987). The amino acid utilized for this pathway in all fish species studied to date is taurine
(James, 1976). It has been suggested that glycine conjugates (the major metabolites of benzoic acids in
mammals) are metabolites of p-aminobenzoate in fish (James, 1986), but no solid evidence supports this
pathway in fish. The enzymes responsible for amino acid conjugation are found in liver and kidney