The thyroid hormone (TH)-disrupting activity of effluents and environmental water samples in Thailand was surveyed by three in vitro bioassays with different endpoints. These assays test the potency of competitive binding with the active form of TH, 3,3',5-[(125)I]triiodo-l-thyronine (T(3)), to the plasma transport protein transthyretin (TTR) and TH receptor (TR; the TTR assay and TR assay, respectively) and the interference with the cellular T(3)-signaling pathway through TR-mediated luciferase gene activation (the luc assay). The TH-disrupting activity in water samples collected from paper manufacturing plants (PMPs), the canal Khlong U-Taphao, and a sewage-treatment plant (STP) was detected predominantly in the dichloromethane/methanol or methanol fractions of solid-phase extraction, suggesting a similar hydrophobic nature of the causative contaminants. The TR assay was relatively more sensitive than the TTR assay to the competitively potent contaminants. The luc assay indicated that the dichloromethane/methanol fractions of most water samples contained anti-T(3)-like activity. Our assays demonstrated that wastewater treatment effectively removed the TH-disrupting contaminants from wastewater in the PMP and the STP. The potencies for TH disruption at the three sampling points of the STP exhibited positive correlations among the three bioassays, whereas those from the canal and PMP water were not correlated among the three bioassays. Furthermore, the influent contaminants that were competitively potent in the TTR assay partially affected the luc assay. These bioassays are useful monitoring tools that give results relevant for evaluating the health of amphibian populations.