用高效液相色谱法纯化和测量脱落酸和吲哚乙酸。
Purification and measurement of abscisic Acid and indoleacetic Acid by high performance liquid chromatography.
机构信息
United States Department of Agriculture, Agricultural Research Service, Western Cotton Research Laboratory, Phoenix, Arizona 85040.
出版信息
Plant Physiol. 1986 Aug;81(4):997-1002. doi: 10.1104/pp.81.4.997.
A procedure was selected for the simultaneous extraction and purification of abscisic acid (ABA) and indoleacetic acid (IAA). Unnecessary steps were eliminated and an accumulation of aqueous phase was avoided. The superior performance of diethyl ether (compared to ethyl acetate) for bulk purification and the superior resolution provided by 250 millimeter columns packed with 5-micrometer spherical particles of strong anion exchanger and octadecylsilane (C18) greatly facilitated the purification of samples. A fixed-wavelength (254 nanometer) ultraviolet detector and a fluorescence detector connected in series on a high performance liquid chromatograph permitted nondestructive monitoring and measurement of ABA and IAA. Derivatization was not necessary for chromatography or for detection. Isocratic elution with simple mobile phases gave sharp peaks. A few simple precautions minimized losses. Recoveries through the entire procedure averaged about 75% for ABA and about 50% for IAA. Purified ABA and IAA fractions were usually free of interfering contaminants. Identities were confirmed by gas chromatography-mass spectrometry.
一种同时提取和纯化脱落酸（ABA）和吲哚乙酸（IAA）的方法被选定。去除了不必要的步骤，并避免了水相的积累。与乙酸乙酯相比，二乙醚（diethyl ether）在大量纯化方面表现出色，而填充有 5 微米球形强阴离子交换剂和十八烷基硅烷（C18）的 250 毫米柱提供的出色分辨率，极大地促进了样品的纯化。高效液相色谱仪上串联的固定波长（254 纳米）紫外检测器和荧光检测器允许对 ABA 和 IAA 进行无损监测和测量。不需要衍生化进行色谱分析或检测。使用简单的流动相进行等度洗脱可得到尖锐的峰。只需采取一些简单的预防措施，就可以最大限度地减少损失。通过整个过程的回收率，ABA 约为 75%，IAA 约为 50%。纯化的 ABA 和 IAA 馏分通常不含干扰污染物。通过气相色谱-质谱联用确认了它们的身份。
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