New Derivatization Approach for the Analysis of Brassinosteroids by Ultra High Performance Liquid Chromatography-Quadrupole Time of Flight Mass Spectrometry

摘要

Brassinosteroids (BRs) were plant hormones of great importance and have been confirmed as the sixth class of plant hormones. BRs exhibit remarkable influence on plants. However, their extremely low concentrations along with co-existent intricate matrix in plants aggravate the difficulties for the isolation and the determination, which induces the ambiguous understanding of their biological synthesis, degradation, metabolic pathways and bio-functions. Many techniques have been employed for the detection and identification of BRs in various plant species. In recent years, LC-MS has been an applicable approach for the analysis of BRs and several boronic acids have been tried for the derivatization of BRs to improve the signal response under electrospray ionization (ESI) source. However, the derivatization reactions were relatively complicated and some of them were not cost-efficient. Due to its increased resolution, higher sensitivity, excellent peak shapes and enhanced reproducibility, ultra-high-performance liquid chromatography (UHPLC) holds advantages over HPLC. In this study, based on UHPLC-MS approach, a new labeling reagent was first introduced to label BRs. Compared to the previous boronic acids reported for BRs derivatization, the new derivatization reagent is chemically stable and commercially inexpensive. The nitrogen atom in the labeling reagent can greatly enhance the ionization efficiency in positive mode of ESI MS. Moreover, the derivatization process is easy to be performed. Based on the new labeling approach, the UHPLC-MS method developed here is highly sensitive and selective with the LOD improved to sub-femtomolar range.