RAW264.7 cells were plated into a 96-well plate at a density of 1 × 104/well in 200 μL of DMEM medium and incubated overnight. Then the medium was replaced with various concentrations of EAE including 0.1, 0.03, and 0.01 mg/mL and incubated for another 24 h. Cell culture media were collected, following the measurement of the nitrite level using Griess reaction assay. Cytotoxicity of EAE was evaluated by MTT assay. The percentage of cell viability was calculated as a percentage of the control. The IC50 of NO inhibition rate was calculated using GraphPad (GraphPad Software, San Diego, CA, USA).
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