C18 column(4.6 mm×250 mm, 5 μm) with methanol(A)-0.1%-phosphoric acid solution(B) as the mobile phase for gradient elution was used in the HPLC to measure the contents of five flavonoids. The detection wavelength of (2R,3R)-dihydromyricetin was 290 nm, and the detection wavelength of (2S,3S)-dihydromyricetin, taxifolin, myricitrin and myricetin was 290 nm before 21 min and 255 nm from 21 to 30 min. The flow rate was 1 mL/min, the column temperature was maintained at 30 ℃, and the injection volume was 10 μL. The results showed that the linear regression equations of the contents of (2R,3R)-dihydromyricetin, taxifolin, myricitrin and myricetin were Y=0.503 1X+0.602 8, R2=0.999 2, Y=0.590 4X+ 0.072 0, R2=0.999 1, Y=0.433 4X–0.007 7, R2=0.999 5, Y=0.507 7X–0.020 7, R2=0.998 8, and the linear ranges were 10.6-106, 2.50-50.00, 2.65-53.00, 1.06-26.50 μg/mL, respectively. The single-wavelength(290 nm) method for the determination of (2R,3R)- dihydromyricetin content, and the dual-wavelengths(290, 255 nm) method for the determination of the contents of taxifolin, myricitrin, myricetin and the peak area of (2S,3S)-dihydromyricetin were superior for system application, linear relationship, repeatability, stability and standard recovery with high precision. The chromatographic peak separations of 5 flavonoids were greater than 1.5, the theoretical plate numbers of (2R,3R)-dihydromyricetin was greater than 5 000, the theoretical plate numbers of (2S,3S)-dihydromyricetin, taxifolin, myricitrin, myricetin were greater than 20 000, and the detection time of single sample was 30 min. This method could be used for the analysis and detection of flavonoids contents in vine tea.