This study was conducted to determine the content of atractylenolides I and III in Atractylodes japonica. An established HPLC-DAD method was used to monitor contents of atractylenolides I and III in A. japonica obtained from Korea and China and compared with contents of A. macrocephala, A. chinensis, and A. lancea. Quantitative analysis of atractylenolides I and III was carried out on a Shiseido C18 column (S-5 ${mu}m$, 4.6 mm I.D. ${ imes}$ 250 mm) with gradient elution composed of acetonitile-water. The results show that the average content of atractylenolides I and III in A. japonica were 0.0954 and 0.1963%, respectively, and contents of A. lancea were higher than A. macrocephala, A. chinensis. In this study, we identified the differentiation of the quality of A. japonica from different species and collected locations and established content standard of atractylenolides I and III in A. japonica and this content standard was helpful to quality control of A. japonica.