Identification of Antifungal Bisphosphocholines from Medicinal Gentiana Species

Gentiana species including G. crassicaulis, G. macrophylla, G. dahurica, and G. straminea are used in traditional Chinese medicine as "Qinjiao" for the treatment of rheumatism, hepatitis, and pain. Four antifungal bisphosphocholines [irlbacholine (2) and three new analogues, gentianalines A-C (1, 3, and 4)] were identified from G. crassicaulis by a bioassay-guided fractionation and structure elucidation approach. Subsequent chemical analysis of 56 "Qinjiao" samples (45 from G. crassicaulis, five from G. macrophylla, three from G. dahurica, and three from G. straminea) showed that bisphosphocholines were present in all four Gentiana species, with irlbacholine as the major compound ranging from 2.0 to 6.2 mg per gram of dried material. Irlbacholine exhibited potent in vitro antifungal activity against Cryptococcus neoformans, Aspergillus fumigatus, Candida albicans, and Candida glabrata with minimum inhibitory concentration (MIC) values of 0.63, 1.25, 10.0, and 5.0 μg/mL, respectively. Identification of the bisphosphocholines, a rare class of antifungal natural products, in these medicinal plants provides scientific evidence to complement their medicinal use. The bisphosphocholines carrying a long aliphatic chain possess amphiphilic molecule-like properties with a tendency of retention in both normal and reversed-phase silica gel column chromatography and thereby may be neglected in natural products discovery. This report may stimulate interest in this class of compounds, which warrant the further study of other biological activities as well.