エンテロトキシンの分離精製に関する研究
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概要
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In an effort to identify the active principle of enterotoxin, causing food-poisoning symptoms produced by staphylococcus group, the author performed experiments on the isolation and purification of the toxin as well as the qualitative chemical tests on the extract of the toxin. The results are summarized as follows: 1. The microorganisms used in the experiments are two toxin-producing strains of Micrococcus pyogenes var. aureus (Strain Asakusa and Strain Matsuyama), a strain which does not produce enterotoxin (Strain 209 P). The former ones were positive for all the tests M. C. N. H. G.. 2. The 18 hour cultures of the above-mentioned strains were inoculated in Dolman's mediums, to which 1 % starch had been added, and were incubated for 48 hours at 37℃ in the air containing 30 % carbon dioxide. The muslin filtrates of these cultures were heated 30 minutes at 100℃, and used as the crude toxic solutions from which the enterotoxin was to be extracted. 3. The isolation and purification of the toxin was carried out as follows: The crude solution is salted out with saturated ammonium sulphate; then the protein fraction having an isoelectric point at pH 3.5 is adsorbed at pH 6.3 by purified diatom earth, Hyflosupercel; and its resolution is obtained at pH 7.8. The zoneelectrophoresis using the veronal buffer solution of pH 8.0 is applied to separation of the toxin from other proteins, and 200 voltage current was sent at 8 miliamper 20 hours. The fraction migrating 6 cm from the original point toward the cathod is extracted by physiological saline solution and is dialized over night, precipitated with alcohol and treated with absolute alcohol, ether. The purified toxin extract was greyish white powder. 4. It was found that about 3 mg. (60 μ/kg) of this powder was capable of inducing marked signs of acute gastroenteritis, such as vomiting and diarrhea, in human adults.5. This powder proved negative for the tests of Biuret, Xantoprotein, Ninhydrin and Fehling, but it showed positive reactions to the tests of Orcin and Naphthoresorcin. Paper-chromatography of the substance obtained after the solution of this powder in sulfuric acid of 1 N had been heated and hydrolyzed showed the presence of glucose, glucosamine and glucuronic acid. The results of these qualitative tests would indicate that the active principle of the toxic fraction is mucoprotein.
- 千葉大学の論文