Developmental and substrate specificity of hesperetin-7-O-glucosyltransferase activity in Citrus limon tissues using high-performance liquid chromatographic analysis

The production of glycosylated flavonoids during plant development was investigated by the analysis of glycosyltransferase activity in various citrus plant tissues. A routine method for the separation of flavonoid glucosyltransferase substrates and products using high-performance liquid chromatography (HPLC) was developed to allow their accurate assay in extracts from plant tissues down to levels of less than 0.5 mu g per injection. Cell-free extracts were prepared from developing leaves and flowers of lemon for these assays and ethyl acetate extracts of the assays were quantitatively analyzed by HPLC. The flavanones naringenin and hesperetin, the flavones apiginin and crysin, and the flavonols morin and kaempferol were shown to be glycosylated by the enzyme. The activities were slightly enhanced by the addition of metal ions. Hesperetin 7-O-glucosyltransferase activity was evaluated in young, developing flowers and leaves of lemon trees. The levels varied 30-fold from flower parts (low) to 4 cm leaves (high).