Measuring The Potency of Pulp Mill Effluents for Induction of Hepatic Mixed-Function Oxygenase Activity in Fish

Abstract

A bioassay protocol was optimized for measuring the potency of effluents or waterborne chemicals for inducing mixed-function oxygenase (MFO) activity of rainbow trout (Oncor-hynchus mykiss). Measurements of ethoxyresorufin O-deethylase (EROD) can be made with an established endpoint assay using large volumes of reagents and tissue. However, a new kinetic microplate assay offers significant savings in time, reagents, and sample volumes. Data are distributed lognormally and must be log transformed before statistical analyses. EROD activity increases with exposure time to pulp mill effluent, and a 4-d exposure provides a near-maximal response. Optimum fish size conforms to standard practices in fish toxicology; loading rates should not exceed 1 g of fish per liter of test solution per day. Feed should be withheld from test fish 48 h before testing to reduce the variance of measured activity, and anaesthetizing fish with MS-222 does not affect their response to MFO inducers. Pulp mill effluents do not lose their potency during 2–3 wk of exposure at temperatures ranging from −20 to 13°C, whether stored in plastic or glass. Steel containers were associated with slight losses in potency. Bioassays of MFO induction in fish exposed to liquid effluents are practical and conform to standard practice for testing the lethality of waterborne chemicals. The results are sufficiently precise that differences among means based on five fish per treatment can be discriminated statistically when activity changes by threefold or more.