Estimation of the Bioactivity of Seven Aromatic Compounds Isolated from Niihime Fruit by Simultaneously Monitoring Superoxide Generation and Intracellular Calcium Ion Levels in Neutrophils

Abstract

This study attempted to isolate bioactive compounds from niihime, a sour Citrus fruit, and to evaluate their bioactivity by simultaneously monitoring superoxide generation and intracellular calcium ion levels in neutrophils. Seven aromatic compounds were isolated from a methanol extract of niihime peel and identified as 6-demethoxynobiletin, 6-demethoxytangeretin, nobiletin, tangeretin, sinensetin, 5,7-dimethoxycoumarin, and saccharic acid 1,4-lactone 3,5-di-O-ferulate. 6-Demethoxynobiletin, 6-demethoxytangeretin, and 5,7-dimethoxycoumarin were newly shown to be contained in niihime fruit. Bioactivity assay of 10 µM saccharic acid 1,4-lactone 3,5-di-O-ferulate revealed a significant increase in both superoxide generation and intracellular calcium ion concentration (p<0.05). This result indicates that saccharic acid 1,4-lactone 3,5-di-O-ferulate has immunostimulatory activity in neutrophils.