Abstract: OBJECTIVE To establish an HPLC-DAD method for simultaneously determination of cytosine, cytidine, uracil, adenine, guanine, uridine, thymine, guanosine, thymidine and adenosine in Tibetan medicine of flower of Gentiana stramiinea and flower of Gentiana farreri. Determination and comparison the ten nucleoside compounds of the two Tibetan medicine samples which come from six different origins of Qinghai province. METHODS A Bonus RP column(250 mm×4.6 mm, 5 μm) was used, methanol and 0.04% acetic acid solution was used as the mobile phase, the flow rate was 1.0 mL·min^-1, the column temperature was 35℃ and the detector wavelength was set at 260 nm with injection volumn of 20 mL. RESULTS Ten nucleoside compounds had good linear relationship, precision, stability, and repeatability according to the requirements of the quantitative analysis methodology. The recoveries of ten nucleoside compounds were 95.1%-98.5%. Furthermore, the total contents of the ten nucleoside compounds in different origins were 849.38-1 014.37 μg·g^-1 and 256.27-822.52 μg·g^-1 for the flower of Gentiana farreri and Gentiana straminea respectively. Adenosine, guanosine, uridine and cytidine were the main nucleoside compounds in the two Tibetan medicine samples which were higher than others in content, while uracil, guanine and thymidine were lower. CONCLUSION The developed HPLC-DAD method is simple, available, and has good repeatability, which is suitable for the determination of above ten nucleoside compounds of flower of Gentiana straminea and Gentiana farreri. Moreover, the quantitative data of ten nucleoside compounds in the two Tibetan medicine samples origin from Qinghai province can provide helpful information for the quality control and evaluation.