Sensitive determination of brassinosteroids by solid phase boronate affinity labeling coupled with liquid...

Brassinosteroids (BRs) are regarded as the sixth plant hormone that is widely distributed in the plant kingdom. Sensitive quantification of BRs will be greatly benefit to illuminate the detail mechanisms about how BRs play crucial role in plant developmental processes such as cell division, cell expansion, cytodifferentiation, seed germination, vegetative growth and resisting biological or abiotic stress. In the current study, we developed a method for rapid and sensitive determination of endogenous BRs in plant tissues by combining LC-MS and a novel sample preparation strategy, in which the plant tissue extract was supplied to solid phase boronate affinity labeling and extraction, followed by desorption and salt-induced phase transition extraction for further purification. Under the optimized conditions, good linearity was obtained for 6 BR with correlation coefficients (r) ranging from 0.9988 to 0.9999. The limits of detection (LODs, S/N = 3) ranged from 1.4 to 2.8 pg mL^-1. The recoveries were between 93.4% and 116.2% with the relative standard deviations (RSDs) ranging from 2.8% to 15.8%. Finally, the developed method was successfully applied to the analysis of 6 endogenous BR in various plant tissues including 20 mg FW Oryza sativa shoot, 10 mg FW Oryza sativa root, 20 mg FW Arabidopsis thaliana shoot, 4 Arabidopsis thaliana flowers (2.8 mg) and one Brassica napus stamen (3.0 mg) with concentration ranging from 0.26 to 157.28 ng g^-1 FW.