The Canadian water quality guidelines for the protection of aquatic life assume that (1) the external water concentration is an effective measure of the concentration at the active site in organisms, which is ultimately responsible for a toxic response, and that (2) the safety factor accounts for any differences between laboratory and field conditions as well as the extrapolation from the effect concentration to a long-term no-effect concentration. This study examines these assumptions and assesses potential errors that environmental managers can make when applying the guidelines. The methodology is based on assessing the probability that internal concentrations of several contaminants are greater than or less than the ''safe'' concentration, assumed by the guideline, when the water concentration is at the water quality guideline. Results derived from empirical observations and a food-web bioaccumulation model show that a high probability (62-100%) exists that safe internal concentrations are exceeded for polychlorinated biphenyls, 1,2,4,5-tetrachlorobenzene, and hexachlorobenzene in Lake Ontario when the aqueous concentrations are at the water quality guideline values. This is due to field bioaccumulation factors being greater than the bioaccumulation factors in laboratory toxicity tests used for the water quality guideline development. Factors contributing to the exceedence of safe internal concentrations at the water quality guideline values are identified. Recommendations for improvement of the water quality guideline process are provided.