Study on anti-inflammatory activity of the main component of Girald Daphne Bark in vitro

Abstract

Objective To investigate the anti-inflammatory activity of the main active ingredients in thedried stem bark of Daphne giraldii Nitsche. Methods Severialchemical compounds like vladinol D, pinoresinol, daphneticin, daphnoretin, daphnetin, giraloid A and giraldoid B were isolated from the stem barks. The CCK-8 experiemnts were analyzed for the cytotoxicity study. The cells were divided into the control group, the model group and the treatment group according to random number table method. The control group and the model group were added with 50 μl culture medium. Moreover, treatment group was added with different concentrations (50.00, 25.00, 12.50, 6.25, 3.12 μg/ml) of the solutions of giraloid A, giraldoid B and daphneticin. Then, RAW264.7 cells were treated with 50 μl LPS (4 μg/ml) for 24 h in the model group and treatment group. Griess reagent was used to determine the amount of NO release, and the secretion of TNF-α was detected by ELISA kit. Results Cytotoxicity test indicated that giraldoid A (50.00 μg/ml), giraldoid B (50.00 μg/ml) and daphneticin (50.00 μg/ml) showed noobvious cytotoxicity. Giraldoid B (12.50, 25.00, 50.00 μg/ml) could inhibit the production of NO (271.86% ± 20.92%, 256.48% ± 20.92%, 199.31% ± 15.16% vs. 358.62% ± 28.64%) and TNF-α (647.87% ±115.79%, 618.42% ± 87.52%, 588.33% ± 87.94% vs. 1 035.06% ± 58.29%) in RAW264.7 induced by LPS compared with the model group. Giraldoid A (25, 50 μg/ml) could inhibit the production of NO (234.99% ± 34.28%, 167.36% ± 25.76% vs. 358.62%±28.64%) and TNF-α (691.76% ± 60.37%, 534.01% ± 41.60% vs. 1 035.06% ± 58.29%) in RAW264.7 induced by LPS compared with the model group. Daphneticin (12.5, 25, 50 μg/ml) could inhibit the production of NO (283.89% ± 36.69%, 243.08% ± 48.19%, 225.92% ± 33.67% vs. 358.62% ± 28.64%) and TNF-α (713.77% ± 121.96%, 670.62% ± 18.70% vs. 1 035.06% ± 58.29%) in RAW264.7 induced by LPS compared with the model group. Conclusions Giraldoid A, giraldoid B and daphneticin exhi bited anti-inflammatory effect through inhibiting the release of NO and the production of TNF-α in RAW264.7 induced by LPS. Key words: Daphne giraldiiNitsche; Inflammation; Lipopolysaccharides; Nitric oxide; Tumor necrosis factor-alpha