We have established a microanalytical method for identification of brassinolide as the bismethaneboronate derivative by the computerized mass fragmentography using GC-CIMS system, which allow us to detect as small as picogram amount. The derivatives exhibit moderate volatility and strong molecular ions(e.g. m/z 529 for brassinolide; m/z 513 for the 6-keto analogue ) as base peaks in CI-MS as shown in Fig. 3 and methane boronic acid is a specific reagent for vicinal diol derivatization effecting easy separation from other contaminants in GC. The extracts from the immature seeds and sheaths of chinese cabbage, the leaves of green tea, and the insect galls of chestnut tree were first bioassayed by the rice-lamina inclination test, a highly specific and sensitive method for brassinosteroids and the biologically active fractions were derivatized and analyzed by this microanalytical technique. Brassinolide and its analogues(II, V, VII, VIII) were positively identified as shown in Fig. 4 and 5, thus indicating the ubiquitous distribution of brassinosteroids in the plant kingdom.
