Micro bioassay for biomarkers analysis in fish

LóPEZ ACA, V.; GONZALEZ, P.V.; CARRIQUIRIBORDE, P.

Guaraparí

Congreso; XIII Congresso Brasileiro de Ecotoxicología; 2014

Sociedad Brasilera de Ecotoxicología-SETAC-Brasil

The aim of the present study was to develop a bioassay to assess EROD (ethoxyresorufin-O-deethylase) and AchE (acetylcholinesterase) activities in fish at microscale. Experiments were performed in 200 ml jars using pools with different numbers of 15 (8.14 ± 1.52 mg) and 30 (14.55 ± 2.93 mg) day old pejerrey (Odontesthes bonariensis) larvae. Chlorpyrifos (0-0.1-0.5-1.0-5.0-10 µg/L) and â-naphthoflavone (0-0.1-1.0-10-100 µg/L) were used as reference toxicant prepared in dechlorinated tap water and using DMSO (dimethyl sulfoxide) as vehicle. Exposure time varied from 48 to 96h. EROD activity was assessed in whole body homogenate, while the head and the trunk were homogenized separately for AchE. Both activities were normalized again total protein content measured by Lowry. Results showed that pools of at least 10 individuals are need to detect basal EROD activity. Enzyme was dose-response induced, showing 35% and 56% or 26% and 62% increased activity in fish exposed to 10 and 100 µg â-BNF/L after 48h or 96h, respectively. This data indicate that induction plateau was reached quickly before 48h. Measurement of AchE activity was more sensitive and just one larvae was needed to detect basal activity separately in head and trunk. Two separated experiments confirmed that dose-response curves for this enzyme are characterized for an induction, both in the head (25%) and the trunk (61%) at low (0.1 µg CHL/L) concentrations followed by inhibition at higher concentrations in both sectors. The activity decrease observed in the head was 36, 52, 88 and 67%, while in the head was 27, 75, 94, and 86% for fish exposed to 0.5, 1.0, 10, 5 and 10 µg CHL/L, respectively. Inhibition plateau seemed to be reached beyond 5 µg CHL/L. A simple, practical and fast micro bioassay was developed for ?in vivo? detecting of neurotoxicants and phase I biotransformation inducers pollutants in fish.