Partial purification and characterization of gizzard erosion-inducing substance in heated casein-histidine mixture.

Abstract

A heated casein-histidine mixture, or the model compound of gizzard erosion-inducing fish meal, was prepared, and a gizzard erosion-inducing substance in this mixture was partially purified. The mixture was refluxed with hydrochloric acid, and the supernatant was obtained. The supernatant was then adjusted to the pH of the isoelectric point of casein and the resulting precipitate collected. The precipitate was further hydrolyzed with papain at a neutral pH. After acidification of the reaction mixture, the supernatant and the precipitate were separated. The supernatant fraction, which was toxic to induce gizzard erosion, was submitted to a gel filtration of Sephadex G-25, and resolved into five fractions. Gizzard erosion-inducing toxicity was found only in the second fraction. The non-toxic second fraction was also prepared by the same treatment from the non-toxic material of heated casein. The characters of these toxic and non-toxic second fractions were compared by using several analytical methods. Thin layer chromatography, as well as gel permeation chromatography, revealed that more than eight components having widely distributed molecular weights existed in both second fractions. By reversed phase chromatography, these fractions were found to consist of various peptides (or peptide-like compounds). Some characterizations, however, were performed. It failed to detect any difference in the components between the toxic and non-toxic second fractions by any analytical method used.