The partial purification and bioassay of a toxin present in extracts of the sea anemone, Tealia felina (L.).

摘要

1. Column chromatography with Agarose A50m followed by Sephadex G100 was used to separate a fraction (extract II) in the molecular weight range 12,000 to 14,000 daltons from saline extracts of the sea anemone, Tealia felina. 2. Extract II inhibited histamine-induced contractions of the guinea-pig ileum and produced haemolysis of human blood, effects on which bioassays were based. 3. The potency of extracts was assayed. A standard unit of activity (= AU) was defined such that 100 AU produced 90% inhibition of histamine-induced contractions of the guinea-pig ileum after 30 to 35 min exposure. 4. The relationship between activity of the extracts measured on the ileum and their haemolytic activity was studied, providing a second assay method based on the latter property. 5 Based on values from both methods of assay, the calculated yield in AU at the end of the separation procedure was 0.53 AU for each AU present in the original extract. In crude extract there were 5.0 AU/mg dry weight and 36.7 AU/mg protein, and after separation (extract II) there were 11.2 AU/mg dry weight and 312.2 AU/mg protein. 6 The acute LD50 values determined in mice (i.v.) were: for crude extract 124 mg/kg for extract I, 76 mg/kg and for extract II, 69 mg/kg. 7 Extract II (0.18 to 0.72 AU/ml) produced a slowly developing contraction of guinea-pig ileum. Indomethacin (2.8 x 10(-5) M) substantially reduced this response. 8 Extract II (0.03 AU/ml) reduced the contractile response of the guinea-pig ileum to acetylcholine by 39 +/- 8%, n = 6, and the response to histamine by 26 +/- 6.6%, n = 6. The response to 5-hydroxytryptamine (5-HT) was not reduced by 0.08 AU/ml of extract II, a concentration that actually increased the contractile response to KC1 by 32 +/- 11.2% n = 7. 9 It is proposed that for future work on the extract a new AU should be used. This AU is defined such that 50 AU produce 50% inhibition of histamine-induced contractions of the guinea-pig ileum after 30 to 35 min exposure.