Clerodane diterpenoid glycosides from the tuberous roots of Paratinospora sagittata: targeted isolation, structure characterization and immunomodulatory properties

Guided by the MS/MS-based molecular networking, eight previously undescribed clerodane diterpenoid glycosides, designated tinospinosides F–M ( 1 − 8 ), along with 12 known analogues ( 9 − 20 ), were isolated from the tuberous roots of Paratinospora sagittata . Structural elucidation of the undescribed compounds was achieved through comprehensive spectroscopic analyses (NMR, HRESIMS), with their absolute configurations confirmed via single-crystal X-ray diffraction, TD-DFT/ECD computational analyses, and chemical degradation. Immunomodulation evaluation on all the isolates revealed that compounds 6 and 7 exerted significant promoting effect toward NO production in RAW264.7 macrophages. Further study demonstrated that 6 could enhance the release of immune cytokines (e.g., TNF- α ) and upregulate the protein expression of iNOS and COX-2, which was potentially mediated through the activation of NF-κB signaling pathway.