OBJECTIVE：To establish a method for the simultaneous determination of uracil， hypoxanthine， xanthine， uridine and adenosine in Eisenia foetida. METHODS： HPLC was performed on the column of Venusil XBP C18 with mobile phase of 50％ methanol-0.01 mol/L potassium dihydrogen phosphate solution （gradient elution） at a flow rate of 1.0 ml/min. The detection wavelength was 254 nm with a column temperature at 30 ℃， and the injection volume was 10 μl. RESULTS：The linear range was 0.05-1.60 μg for uracil（r＝0.999 1），0.05-1.60 μg for hypoxanthine（r＝0.999 3）， 0.002-0.010 μg for xanthine（r＝0.999 1）， 0.0075-0.24 μg for uridine（r＝0.999 9） and 0.025 0-0.80 μg for adenosine （r＝0.999 0）； RSDs of precision， stability and accuracy tests were lower than 3％； recoveries were 98.30％-100.76％ （RSD＝0.8％，n＝9）， 98.68％-100.96％ （RSD＝0.8％，n＝9）， 95.16％-97.67％（RSD＝0.9％，n＝9），96.15％-99.57％（RSD＝1.5％，n＝9） and 96.39％-101.93％（RSD＝1.8％，n＝9），respectively. CONCLUSIONS：The method is simple and stable with good reproducibility， and can be used for the simultaneous determination of uracil， hypoxanthine， xanthine， uridine and adenosine in E. foetida.