Objective:To improve the method for the determination of related substances in vidarabine monophos-phate.Methods:The analysis was performed on an ChromCore AQC18 column(4.6 mm × 150 mm,5 μm)with mobile phase A of an aqueous solution containing 0.01 mol·L-1 tetrabutylammonium hydroxide and 0.1 mol·L-1 potassium dihydrogen phosphate and mobile phase B of methanol by gradient elution at the flow rate of 1.0 mL·min-1.The column temperature was maintained at 30 ℃ and the UV detection wavelength was set at 258 nm.Results:Related substances were effectively separated from the principal component.Vidarabine mono-phosphate and its four impurities showed a good linear relationship in the self-concentration ranges(r＞0.999 9).The average recoveries and were 95.0％-99.2％with RSDs(n=9)of 1.0％-4.4％related substances in vidar-abine monophosphate and 91.8％-102.1％with RSDs of 0.5％-4.8％(n=9)for vidarabine monophosphate for injection,respectively.Conclusion:The improved method is simple,rapid and specific,and can be used for the determination of related substances in vidarabine monophosphate.