白三叶叶蛋白提取及纯化工艺

A study on leaf protein extraction and purification of white clover

以白三叶(Trifolium repens)为材料探讨了叶蛋白的提取工艺和纯化方法。对白三叶叶蛋白提取中加热时间、温度、pH值、料液比、酸的种类等单因素以及纯化试剂甲醇、乙醇、丙酮、四氯化碳、水等进行了研究。结果表明,各单因素最佳参数为加热时间9min,温度90℃,pH值4.0,料液比1∶2,沉淀的酸为硝酸。对加热时间、加热温度、pH值和料液比进行4因素3水平正交试验,发现影响叶蛋白提取因素为pH值>温度>时间>料液比,最佳提取工艺为加热时间9min,温度80℃,pH值4.0,料液比1∶2。对叶蛋白纯化结果表明,纯化剂对叶蛋白纯度的影响依次为甲醇>乙醇>丙酮>四氯化碳>水,但各种纯化剂之间差异不显著(P>0.05)。

The leaf protein （LPC） of white clover （Trifolium repens） were extracted and purified by single factor experiment. Heat time, temperature, pH value, the sort of acid and the ration of fresh leaf to water as extracting factors were studied in extracting leaf protein. Methanol, ethanol, acetone, carbon tetrachloride, water as extracting solutions were studied in purifying leaf protein. The results showed that the opti- mum time, temperature, pH value, acids and the ration of fresh leaf to water were 9 minutes, 90 ℃, 4.0, nitric acid, 1 .＂ 2, respectively. The result of orthogonal experiment showed that the sequence of the factors in affecting extraction ratio of the LPC was pH value, temperature, heat time and the ratio of fresh leaf to water. The optimum combination condition was pH value 4.0, temperature 90 ℃, heat time 9 rain and the fresh leaf to water 1 ＂ 2. The result of the purification method showed that the sequence of the purifiers in affecting protein purification was methanol, ethanol, acetone, carbon tetra,chloride, water; and there were not significant differences （P〉0.05） among the factors.