Spatial Profiling of Gibberellins in a Single Leaf Based on Microscale Matrix Solid-Phase Dispersion and Precolumn...

A spatial-resolved analysis method for profiling of gibberellins (GAs) in a single leaf was developed on the basis of microscale sample preparation and precolumn derivatization coupled with ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The proposed microscale sample preparation was based on modified matrix solid-phase dispersion (MSPD) method, in which the plant sample (<1 mg) and C18 sorbent were ground together in one microcentrifuge tube, and then extraction solvent was added followed by centrifugation. In this protocol, the grinding, extraction, and purification were performed in one microcentrifuge tube without any sample transfer step, resulting in an obvious decrease in sample loss. Moreover, a new derivatization reagent, 3-bromopropyltrimethylammonium bromide (BPTAB), was used to further enhance the signal intensities of GAs on MS by 3-4 orders of magnitude, which was much higher than the reported derivatization reagents for GAs such as bromocholine bromide and 3-bromoactonyltrimethylammonium bromide. The present method showed high sensitivity (minimum detectable amount (MDA) of 10.1-72.3 amol for eight GAs) and low sample consumption (down to 0.30 mg FW). Under the optimized conditions, the distribution of GA₁₉ in a single Arabidopsis thaliana leaf was profiled with a spatial resolution of 2 × 2 mm².