Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), Department of Chemistry , Wuhan University , Wuhan 430072 , P.R. China.
State Key Laboratory of Hybrid Rice, Department of Cell and Developmental Biology, College of Life Sciences , Wuhan University , Wuhan 430072 , P.R. China.
Anal Chem. 2019 Mar 5;91(5):3492-3499. doi: 10.1021/acs.analchem.8b05087. Epub 2019 Feb 18.
Phytohormones play crucial roles in every aspect of plant life, and their regulatory functions rely on complex crosstalk networks among the different classes of phytohormones in an antagonistic or synergistic manner. Therefore, the simultaneous determination of multiclass phytohormones is important for studies of phytohormone functions and networks. However, due to the heterogeneity of the sensitivity resulting from structural diversity and the low content in the plant samples, simultaneous determination of multiclass phytohormones is challenging, especially in very tiny plant tissues or organs. Here, we describe a novel method for the simultaneous determination of 31 phytohormones from different classes in a single run. This uses a one-pot multifunctional derivatization coupled to liquid chromatography-tandem mass spectrometry (LC-MS/MS). N, N-Diethyl ethylenediamine (DEED) and 2-methyl-4-phenylaminomethyl-benzeneboronic acid (2-methy-4-PAMBA) were used for derivatization of carboxylated phytohormones and brassinosteroids (BRs), respectively, to simultaneously improve detection sensitivities of carboxylated phytohormones and BRs. The method was fully validated for the 31 targeted phytohormones and could quantify multiclass endogenous phytohormones in small amounts of fresh plant samples (0.02-5 mg, FW). Finally, we used this method to investigate the spatial-temporal distribution of multiple phytohormones in reproductive organs of a single flower of Arabidopsis thaliana for the first time. This method could be a powerful auxiliary tool for studies of phytohormone functions and regulatory networks.
植物激素在植物生命的各个方面都起着至关重要的作用，它们的调节功能依赖于不同类别的植物激素之间以拮抗或协同方式进行的复杂串扰网络。因此，同时测定多类植物激素对于研究植物激素功能和网络非常重要。然而，由于结构多样性导致的敏感性异质性和植物样品中含量低，同时测定多类植物激素具有挑战性，特别是在非常小的植物组织或器官中。在这里，我们描述了一种新的方法，可在单次运行中同时测定来自不同类别的 31 种植物激素。该方法使用一锅多功能衍生化，结合液相色谱-串联质谱（LC-MS/MS）。N，N-二乙基乙二胺（DEED）和 2-甲基-4-苯氨基甲基苯硼酸（2-甲基-4-PAMBA）分别用于衍生化羧基化植物激素和油菜素内酯（BRs），以同时提高羧基化植物激素和 BRs 的检测灵敏度。该方法对 31 种目标植物激素进行了全面验证，并能够定量测定少量新鲜植物样品（0.02-5mg，FW）中的多种内源性植物激素。最后，我们首次使用该方法研究了拟南芥一朵花生殖器官中多种植物激素的时空分布。该方法可以成为研究植物激素功能和调控网络的有力辅助工具。
