Studies on anti-ulcerogenic protein in inflamed rabbit skin tissues. II. Isolation of inhibitory substance to gastric juice secretion and carrageenin edema formation from Arthus reaction site

Aonuma, S.; Kohama, Y.; Fujimoto, S.; Yoneda, M.; Kusumori, M.

Yakugaku Zasshi Journal of the Pharmaceutical Society of Japan 99(9): 889-894


ISSN/ISBN: 0031-6903
PMID: 161919
DOI: 10.1248/yakushi1947.99.9_889
Accession: 068522951

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A protein inhibitive of both gastric juice secretion and carrageenan edema formation in rats was isolated from rabbit skin tissues with Arthus reaction. The tissues obtained at 24 h of Arthus reaction were minced and dried with acetone. Dried powder was extracted with distilled water (pH 8.0), and the precipitate formed at pH 4.5 was removed from the extract. An inhibitory activity was concentrated by precipitation with 30% ammonium sulfate saturation, followed by fractionation with chromatography over Sephadex G-200 and DEAE cellulose columns. The yield of the purified active component, anti-ulcerogenic protein from rabbit skin with Arthus reaction, (APRA) was 31 mg from 100 g of tissues. APRA was a protein with a small amount of carbohydrate and free of nucleic acid, had a MW of 140,000 in the native conditions of gel filtration with Sephadex G-200 and consisted of 50,000 and 23,500 subunits with polyacrylamide gel electrophoresis in the presence of 2-mercaptoethanol and sodium dodecyl sulfate, and rabbit serum IgG. Its amino acid composition was similar to that of IgG except for histidine and cystine. But APRA did not react with guinea pig antiserum to rabbit serum IgG in precipitin test in agar. This protein (10 mg/kg, i.p.) showed the % inhibition of 46.9, 60.4, 42.2 and 46.5 in gastric juice volume, total acid output, total peptic activity and fore-stomach ulceration, respectively, in pylorus-ligated rats. APRA (20 mg/kg, s.c.) inhibited carrageenan edema formation at a degree of 40% 4, 5 and 6 h after irritant injection in rats. APRA did not represent irritant activity by subplantar injection in rats, any protease activity or any anti-papain activity in the casein digestion method.