Identification and functional analysis of the autofluorescent substance in Limonium bicolor salt glands

Limonium bicolor is a typical recretohalophyte with salt glands for the secretion of excess salts into the environment. We observed that L. bicolor salt glands showed obvious blue autofluorescence under UV excitation (330-380 nm). The aim of the present study was to identify and clarify a role for this autofluorescent substance in salt secretion. Sudan IV staining showed that the autofluorescent substance was localized in the cuticle of the salt glands. Moreover, the primary autofluorescent substance was identified as ferulic acid after treatment with 0.1 M ammonium hydroxide solution, alkaline and enzymatic hydrolysis. Additional experiments using two mutants exhibiting increased (fii) and decreased (fid) salt gland fluorescence indicated that the fluorescence intensity of salt glands under UV excitation was positively correlated with the content of ferulic acid in the cuticle, strongly suggesting that the primary autofluorescent substance in the salt glands was ferulic acid. Salt gland secretion was determined using leaf discs, and the results showed that the Na+ secretion rate per single salt gland was also positively correlated with the content of ferulic acid in the cuticle, suggesting that ferulic acid in the cuticle was directly involved in salt secretion of salt gland.