Abstract: Ball mill method was used to extract flavone C-glycosides from bamboo leaves using deep eutectic solvents(DESs) as extraction solvent. The contents of four flavone C-glycosides(isoorientin, orientin, isovitexin and vitexin) in bamboo leaves were determined by liquid chromatography quadrupole time-of-flight mass spectrometry(LC-Q-TOF-MS). By single factor experiments and orthogonal design, the parameters including water content, extraction time and liquid-solid ratio of DESs were investigated. The results showed that the optimized extraction process of flavone C-glycosides from bamboo leaves was as follows: DESs(composed of choline chloride and ethylene glycol) water content of 50%, extraction time of 150 s, and liquid-solid ratio of 6∶1(mL∶g). Under these conditions, the total extraction yield of four flavone C-glycosides from bamboo leaves was 1.55 mg/g, including isoorientin of 0.99 mg/g, orientin of 0.23 mg/g, vitexin of 0.04 mg/g and isovitexin of 0.29 mg/g. The extraction yield of this method was 7.6% higher than that of immersion extraction, and the time and organic solvent were saved. This optimized technology combined with LC-Q-TOF-MS detection was successfully applied to the extraction and determination of flavone C-glycosides in bamboo tea samples made from bamboo leaves.