Halpern M, Schulman N, Scribani L, Kirschenbaum D M
Pharmacol Biochem Behav. 1984 Oct;21(4):655-62. doi: 10.1016/s0091-3057(84)80053-2.
Garter snakes reliably spend more time and tongue flick more frequently at a dish containing surface washings of earthworm (EW) than at a control dish containing water or sodium chloride. In a series of experiments EW was examined to further characterize and purify the snake-attracting component(s). Earthworm wash boiled for up to two hours did not lose its snake-attracting activity, but EW boiled for four hours was not discriminated from a control substance. The active components of EW were completely precipitated by 90 and 100% saturated ammonium sulfate solutions. The molecular weight of the active components, as determined by dialysis, is larger than 67,000 daltons. Earthworm wash was separated into two peaks on either G-75 or AcA 44. The larger peak (F2) contains snake-attracting material of molecular weight greater than 67K. The smaller peak material (F4) is inactive as a snake-attractant and contains material of approximate molecular weight of 3K. Biological activity of EW covaried with Lowry (protein) assay and Dubois (carbohydrate) assay values and 280 nm absorbance. The response eliciting components of EW are not extractable from earthworms at temperatures below 60 degrees C, but can be extracted at 60 degrees C (for one minute) from the worms or the soil that forms the bedding for the earthworms.
束带蛇在装有蚯蚓体表冲洗液（EW）的盘子上花费的时间明显更多，且比在装有水或氯化钠的对照盘上更频繁地吐舌头。在一系列实验中，对EW进行了检测，以进一步表征和纯化吸引蛇的成分。蚯蚓冲洗液煮沸长达两小时仍未丧失吸引蛇的活性，但煮沸四小时的EW与对照物质无差异。EW的活性成分完全沉淀于90%和100%饱和硫酸铵溶液中。通过透析测定，活性成分的分子量大于67,000道尔顿。蚯蚓冲洗液在G-75或AcA 44上分离为两个峰。较大的峰（F2）含有分子量大于67K的吸引蛇的物质。较小峰的物质（F4）作为吸引蛇的物质无活性，且含有分子量约为3K的物质。EW的生物活性与洛瑞（蛋白质）测定法、杜波依斯（碳水化合物）测定法的值以及280nm吸光度相关。EW的引发反应成分在60摄氏度以下的温度下无法从蚯蚓中提取，但在60摄氏度（一分钟）时可从蚯蚓或构成蚯蚓垫料的土壤中提取。
