植物鲜样中游离氨基酸提取方法的比较

Comparison of Methods for Free Amino Acid Extracted from Fresh Plant Samples

为了对比不同提取液、温度、时间和纯化试剂对植物鲜样中各游离氨基酸提取效率的影响,以油菜鲜叶为例进行加标回收试验方法,通过氨基酸分析仪PF柱进行标准方法分析测定。结果表明：4种不同提取剂提取油菜鲜叶中游离氨基酸,水、0.1 mol/L盐酸和5%磺基水杨酸的提取效果都优于80%乙醇,以水提取后的颜色最为清亮,适于氨基酸分析仪直接分析;4℃时17种氨基酸平均回收率94.23%,高于80℃和100℃时的88.4%和87.4%,17种氨基酸平均回收率除缬氨酸外,其余氨基酸从1-24 h没有显著变化;80%乙醇优于5%磺基水杨酸,但两者整体回收率都较未纯化前有所下降。采用优化后的提取方法测定油菜幼苗的游离氨基酸含量,发现能有效筛选出抗逆品种。

To study the influence of different extraction solvents,temperatures,treatment times and purification reagents on extraction efficiency of the free amino acid from fresh samples,taking rape fresh leaves as samples for standard recovery test,the contents of free amino acid from rape fresh leaves were measured. The test showed that the extraction efficiencies of water,0. 1 mol / L hydrochloric acid and 5% sulfosalicylic acid were all better than that of 80% ethanol.Especially,the color of the plant extract that extracted by water was most clean and suitable for the detection of free amino acids directly. The results of the extraction efficiency of water at different temperatures showed that the average recovery of 17 kinds of amino acid at 4 ℃（ 94. 23%） was higher than that at 80 ℃（ 88. 4%） and at 100 ℃（ 87. 4%）. It indicated extraction at low temperature could significantly increase the recovery of basic amino acid. No significant changes in the average recoveries of 17 kinds of amino acid except valine were observed when extracted for different treatment times from 1 h to 24 h at 4 °C. The purification test for the plant extract that was extracted by water showed that the purification efficiency of 80% ethanol was better than that of 5% sulfosalicylic acid,but the recoveries were lower than that before purification. Therefore,the free amino acid extracted by water at 4 °C for 1 h to 24 h should be the optimized extraction method. Especially,using the optimized extraction method for determining free amino acid content,the disease-resistant rape varieties were efficiently selected from different rape varieties. The optimized extraction method was proved to be a simple,convenient and effective method for the detection of free amino acids from fresh plant sample.