Effects of physical binding ofo-nitroanisole with feed upon its systemic availability in male F344 rats

Physical binding of test chemicals to feed often occurs in dose formulations prepared for uses in toxicology studies. The tenacity of this binding appears to increase with storage time of the feed formulations. The effects of this binding on systemic bioavailability are unknown, o-Nitroanisole (ONA) which is a typical example of this phenomenon, has been widely used in the dye industry as a dyestuff intermediate (Kirk-Othmer, 1963). Because of potential worker exposure, its chronic toxicity and carcinogenicity was evaluated by the National Toxicology Program in a 2-year bioassay with ONA in dosed feed formulations. Routine analysis of ONA concentrations in feed using a standard extraction procedure showed that the apparent concentration decreased with storage time. However, after the aged ONA feed samples were digested with strong acid (8.3 N HC1), the determined concentrations were the same as those obtained from freshly prepared feed formulations. This finding was indicative of physical binding between ONA and feed components. In this fgaper, we report the results of a study in which two groups of male F344 rats were dosed for 7 consecutive days with freshly prepared ONA feed formulations or 30 day aged ONA feed formulations followed by an 18 hour urine collection. To evaluate the systemic availability the urinary concentrations of ONA's main metabolites, free and conjugated o-nitrophenol (ONP), as well as the levels of creatinine were determined. The influence of ONA physical binding upon its systemic bioavailability was evaluated by comparing the total (free and conjugated) ON-P urinary excretion between the two groups. MATERIALS AND METHODS o-Nitroanisole( lot #1517AM/04) was obtained from Aldrich Chemical Co. and the purity was found to be 99%. m-Dinitrobenzene (Lot # 107F-5011) was obtained from Sigma Co. and was used as the internal standard for rat urine ONP analysis. o-Nitrophenol (Lot #88-75-5) was obtained from Aldrich Chemical Company Inc. and used as a reference standard. HPLC grade acetonitrile (lot # A998-4, Fisher Solvents), reagent grade ethyl ether (lot # E138-1, Fisher Solvents) and reagent Send reprint requests to C.W. Jameson.