Identification and Anti-Inflammatory Activity of Flavonoids from Cardiospermum Halicacabum in Murine Raw 264.7 Macrophage-A Bioassay-Guided Approach

Cardiospermum halicacabum Linn. (CH), which is commonly known as the ballon vein is used as a source of food as well as to treat various ailments, including inflammation. In Tamil cuisine, the stems and leaves of CH are used to make curries, lentil crepes, sauces and vegetable soups. Several studies have documented the anti-inflammatory activity of CH. However, the therapeutic constituents responsible for its anti-inflammatory activity remain unclear. This study aims to isolate the anti-inflammatory constituents from CH via a bioassay-guided approach. CH aerial part was extracted with water, ethanol and ethanol-water and the active ethanol-water extract was subjected to solvent-solvent partitioning to get five fractions. Among them, the methanol fraction (CH-EW-ML) showed potent anti-inflammatory activity. Further fractionation of the active methanol fraction resulted in the isolation of two flavonoids, apigenin-7-O-β-D-glucuronide (AAG) and diosmetin-7-O-β-D-glucuronide (DMG). DMG was isolated from CH for the first time. The compounds were evaluated for their nitric oxide (NO) inhibition activity in the RAW 264.7 cell line. DMG showed the most potent activity (IC50=16.72 ± 1.01 µM). Further studies showed that DMG can inhibit the production of ROS and proinflammatory cytokines as well as upregulate the expression of anti-inflammatory cytokines. This study represents the first evidence that DMG can inhibit NO, ROS, TNF-α, and IL-6 while upregulating IL-10. Our study confirmed that the infusion of CH in the food can help to alleviate inflammation and DMG could be the promising therapeutic constituent contributing to the anti-inflammatory properties observed for CH.