Pyranochromones with Anti-Inflammatory Activities in Arthritis from Calophyllum membranaceum .

Eleven new pyranochromones, calomembranone A-K ( 1 - 11) , two new pyranocoumarins, calopolyanolide E and F ( 12 and 13 ), together with six known analogues ( 14 - 19 ) were isolated from the leaves of Calophyllum membranaceum . Their structures and absolute configurations were elucidated by analysis of spectroscopic data, computational calculations, as well as X-ray crystallography of 4 and 9 . The anti-inflammatory activities of all the isolates were evaluated by measuring their NO inhibitory effects in LPS-stimulated RAW 264.7 cells. Structure-activity relationships are also discussed. Compound 7 showed the strongest NO inhibition (IC 50 = 0.92 μM). Oral administration of 7 dose-dependently reduced the paw swelling and downregulated neutrophil-to-lymphocyte ratio in the carrageenan-induced acute arthritis mice model. Molecular dynamics simulation and cellular thermal shift assay results indicated that 7 participated in a robust and stable interaction with the active site of TLR4. Compound 7 also suppressed the inflammation in arthritis through the regulation of TLR4 mediated signal transduction via IKK/NF-κB signaling pathway and the consequent reduction of IL-2, IL-4, and IL-5.