The material basis and toxicity of Lithocarpus litseifolius (Hance) Chun (sweet tea), which has many health benefits, are not fully understood.In this study, total water-soluble flavonoids were extracted from sweet tea using optimized methods based on RSM.Its composition and toxicity were investigated by LC-MS and mouse models in vivo.The results showed that the optimum extraction conditions were a liquid/solid ratio of 32.98:1 mL/g, a temperature of 60.69°C, a microwave power of 633.70 W, and a microwave time of 164.08 secs.Among the 33 main components in the extract, five flavonoids constituted more than 73.80% of the total compound content.Total flavonoids were safe after oral administration and alleviated reproductive disorders but were highly toxic after i.p. injection in mice.In summary, under the optimum extraction conditions, the total water-soluble flavonoids were safe after oral administration, but the complex compounds hindered their direct injection.: LLHC: Lithocarpus litseifolius (Hance) Chun; SFD: single-factor design; RSM: response surface methodol.; CCD: central composite design; ES: extracted solution; CS concentrated solution; LP: lyophilised powder; LC-MS: liquid chromatog.-mass spectrometry; NC: neg. control; PH: procaine hydrochloride; CP: cyclophosphamide group.