Screening of Fresh Water Fish Extracts for Enzyme-Inducing Substances by an Aryl Hydrocarbon Hydroxylase Induction Bioassay Technique

Abstract

A sensitive biological test to detect the presence of certain contaminants, such as highly toxic halogenated dioxins, dibenzofurans, and biphenyls in foods, was applied to extracts of fresh water fish that had been prepared by a food extraction-cleanup procedure developed by the Food and Drug Administration for pesticides and industrial chemicals. Aryl hydrocarbon hydroxylase (AHH) activity in a rat hepatoma cell line was used as the biological detection system for residues that induce enzyme activity. The induction of AHH activity by the extracts was compared with a standard AHH-induction curve for the most active compound known, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and results were computed as TCDD equivalents. Several dilutions of fish extracts were used to produce AHH-induction curves from which an optimal dose-response range was determined and used to estimate TCDD equivalents. Cleaned-up extracts of fish obtained from different water bodies in the United States were examined for AHH activity. The samples which had low levels of polyhalogenated contaminants produced low biological activity, while a higher activity was obtained from fish that contained higher levels of polyhalogenated contaminants. The results suggest that the fish extracts can be screened for AHH inducers before chemical analysis.

Aryl Hydrocarbon Receptor-Mediated Toxic Potency of Dissolved Lipophilic Organic Contaminants Collected From Lincoln Creek, Milwaukee, Wisconsin, Usa, to PLHC-1 (Poeciliopsis Lucida) Fish Hepatoma Cells

“…Use of in vitro assays of CYP1A induction for assessing the biological potency of environmental samples has been described [13,15,[26][27][28]. Correlations between in vitro aryl hydrocarbon hydroxylase (AHH) induction and adverse biolog-ical effects have indicated that such assays can be used to evaluate and compare the potential toxicity of environmental samples [15,28,29].…”

Characterization of the H4IIE rat hepatoma cell bioassay as a tool for assessing toxic potency of planar halogenated hydrocarbons in environmental samples

“…A detection limit of 10 pg of TCDD was reported with the isooctane solvent carrier system, with an ED50 of 45 pg of TCDD/ plate (0.14 pmol/plate, 28 pM). A quantitation limit for this solvent system was not reported; however, in subsequent publications the limit of quantitation was 25 pg of TCDD and the linear response range was 25-500 pg of TCDD/plate (38,40). Thus, the sensitivity of this bioassay system for detection of PHHs had been established.…”

“…The first use of the H4IIE bioassay as a tool for assessing complex mixtures of PHHs in extracts was by the U.S. Food and Drug Administration. They performed the initial analytical characterization of the H4IIE bioassay as an environmental extract assay (37)(38)(39)(40). Isooctane (ISO) as the solvent carrier for extracts or pure compounds optimally enhanced bioassay sensitivity (39).…”

“…The H4IIE bioassay has been shown to be a sensitive tool for detection of PHHs in extracts of environmental samples (38)(39)(40) and much of the initial analytical characterization was performed by these scientists. However, due to improvements in PHH extractions, cleanup, and quantitation techniques in the past 10 years, studies to confirm and expand on the work already done are necessary if this bioassay technique is to be adapted as a bioanalytical tool.…”

Aryl hydrocarbon receptor‐mediated toxic potency of dissolved lipophilic organic contaminants collected from Lincoln Creek, Milwaukee, Wisconsin, USA, to PLHC‐1 (Poeciliopsis lucida) fish hepatoma cells

“…Use of in vitro assays of CYP1A induction for assessing the biological potency of environmental samples has been described [13,15,26–28]. Correlations between in vitro aryl hydrocarbon hydroxylase (AHH) induction and adverse biological effects have indicated that such assays can be used to evaluate and compare the potential toxicity of environmental samples [15,28,29].…”