UPLC-PDA-MS guided isolation of lindenane sesquiterpene oligomers with anti-neuroinflammatory activity from two rare Chloranthus species

A UPLC-PDA-MS guided phytochemical research on two rare varieties of Chloranthus holostegius led to the rapid isolation of eleven previously undescribed lindenane sesquiterpene (LDS) dimers [holotrichones C-K (1-9) and holoshimolides A-B (10, 11)], along with thirty-two known analogues (12-43). The previously undescribed structures with the absolute configurations were determined through comprehensive spectroscopic analysis and ECD exciton chirality method, corroborating chemical calculations. Structurally, compounds 2, 3, 10, and 11 feature an uncommon 4'α-OH group instead of the typical β-oriented configuration, which has never been encountered among LDS oligomers so far. Compounds 2, 3, and 11 also represent the first examples of LDSs bearing a saturated 2-methylbutyryloxy substituent. In the anti-neuroinflammatory assay, a series of di- and trimeric LDSs containing a common methyl (Z)-2-methyl-4-oxobut-2-enoate moiety (at positions C-7, C-8 and C-11-C-13) exhibited noticeable inhibition of NO production in LPS-stimulated BV-2 microglia cells, with compounds 5 and 23 showing the highest potency (IC₅₀s < 5 μM). Further evaluation of compounds 5, 6, and 34 revealed that they also suppressed the LPS-induced release of key inflammatory cytokines (TNF-α, IL-6, IL-1β) and reduced the transcriptional expression of iNOS in this neuroinflammation model.