GPC-HPLC-LC/MS测定植物组织中的赤霉素  

Quantitative analysis of gibberellins in plant tissues by GPC-HPLC-LC / MS

作　　者：李金克[1] 邓文红[1] 陈少良[2] 

机构地区：[1]北京林业大学实验室与设备管理处分析测试中心 [2]北京林业大学生物科学与技术学院

出　　处：《北京林业大学学报》2014年第6期171-176,共6页Journal of Beijing Forestry University

基　　金：国家自然科学基金项目(31270654;31170570);教育部科学技术研究(科学技术类)项目(113013A);人事部留学人员科技活动项目择优资助经费;高等学校学科创新引智计划(111 Project;B13007);教育部创新团队发展计划项目(IRT13047)

摘　　要：以胡杨叶片、烟草组培苗及萌发绿豆为材料,研发了利用凝胶渗透色谱（GPC）纯化、液相质谱（LC-MS）定性、高效液相色谱（HPLC）定量测定植物组织中赤霉素（GAS）的方法。植物样品用80%甲醇研磨后4℃浸提过夜,抽滤离心（5 000 r/min,20 min）、滤液加2滴浓氨水浓缩至水相,冻融离心,上清液,调p H 2.5-3.0乙酸乙酯萃取,过Sep-pak C18小柱,用含5%甲醇的CH2Cl2溶解样品,经0.22μm滤膜过滤后,通过GPC纯化、LC-MS定性,利用HPLC外标曲线法定量。经检测：胡杨叶片中GA3含量为1 162.789 8 ng/g鲜质量;烟草中GA3含量为920.906 7ng/g鲜质量;萌发绿豆中GA3含量为700.923 6 ng/g鲜质量。We developed a protocol to determine the contents of gibberellins（ GAs） in various plant tissues,such as Populus euphratica leaves,Nicotiana tabacum plantlets,and germinated green beans（ Vigna radiata）. GAs samples were purified with gel permeation chromatography（ GPC）,confirmed by liquid chromatography-mass spectrometry（ LC-MS）,and quantified by high performance liquid chromatography（ HPLC）. Plant samples were ground with 80% methanol and extracted at 4 ℃ overnight. Crude extract was filtrated and then centrifuged at 5 000 r / min for 20 min. Two drops of ammonia were added to the liquid sample which was concentrated to aqueous phase,then frozen,thawed and centrifuged. Upper liquids were re-extracted with ethyl acetate at p H 2. 5- 3. 0（ × 3）,and purified by Sep-pak C18 columns. The samples were resolved by 5% methanol-CH2Cl2 and filtrated through 0. 22 μm filter. Then samples were purified through GPC and GAs fluid was collected for subsequent quantification. GAs in the GPC-purified samples was quantified with HPLC by means of external standard curves. The gibberellins detected in plant tissues were confirmed with LC-MS. The result showed that the GA3 content was1 162. 789 8 ng / g FW in poplar leaves,920. 906 7 ng / g FW in tobacco leaves and 700. 923 6 ng / g FW in germinated green beans.

关 键 词：赤霉素 外标曲线法 凝胶渗透色谱 高效液相色谱 液相色谱-质谱 

分 类 号：S718.43[农业科学—林学] Q946.885.5[生物学—植物学]