OBJECTIVE： To establish a method for the simultaneous determination of juglone and regiolone in Juglans mandshurica. METHODS： HPLC was used and regiolone was used as internal reference， the relative correction factor （RCF） of regiolone and juglone wascalculated， then calculated the content of juglone based on RCF； the juglone content was used as measured value， which was determined by external standard method， angle cosine method was used to compared the difference of calculated value in quantitative analysis of multi-components by single-maker and measured value in external standard method. RESULTS： The linear range was 7.6-76.0 μg/mL for juglone （r＝0.999 0） and 8.2-82.0 μg/mL for regiolone （r＝0.999 4）； RSDs of precision， stability and reproducubility tests were lower than 3.0％； recoveries were 95.37％-97.94％（RSD＝1.04％，n＝6） and 99.13％-100.10％（RSD＝0.33％，n＝6）. The cosine of the included angle between the calculated and measured values was 0.999 84， and there was no significant difference（P＞0.05）. CONCLUSIONS： The method is simple with good precision， stability and reproducibility， and can be used for the simultaneous determination of juglone and regiolone in J. mandshurica.