Diacetylenic Spiroacetal Enol Ethers from <i>Chrysanthemum indicum</i> Alleviates Inflammation in LPS-Induced RAW 264.7 Cells via Regulation of Nrf2 and MAPK Signaling Pathways

Five novel naturally occurring diacetylenic spiroacetal enol ethers, named chracetylespienoles A-E (<b>1</b>-<b>5</b>), along with five known analogues (<b>6</b>-<b>10</b>), were isolated from the aerial part of <i>Chrysanthemum indicum</i>. A combination of extensive spectroscopy, chemical derivatization, DP4+ NMR analysis, and ECD calculations was employed to determine the new compounds' structures and assign their absolute configurations. The anti-inflammatory activity of <b>1</b>-<b>10</b> was determined by measuring their suppression on NO production in lipopolysaccharide (LPS)-mediated RAW 264.7 cells, and <b>8</b> showed the best NO production inhibitory activity with an IC₅₀ value of 7.42 ± 0.87 μM. Furthermore, compound <b>8</b> downregulated the expression of iNOS and COX-2 proteins by inhibiting the MAPK and activating the Nrf2 signaling pathways, thereby suppressing inflammatory cascades.