〔Abstract〕 Objective To investigate the effect of riboflavin on inactivation of pathogens under different light time conditions. Methods Plasma containingvesicular stomatitis virus(VSV) was divided into 4 groups, and the final concentrations of riboflavin were 50, 100, 150 μmol/L and 0 μmol/L was used as the control group. The growth titers at 10, 15, 20 and 25 min were observed. The control group was treated with no light. The growth titers of VSV culture before and after inactivation were calculated by Reed-Muench method, and the effect of different concentrations of riboflavin on the inactivation effect was evaluated by log reduction factor after inactivation. Results Under the condition of no light, the addition of riboflavin had no pathogen inactivation effect. With the increase of light time from 15 min to 25 min, the inactivation effect of VSV in the non-riboflavin group increased. When the light was less than 15 min, there was no significant difference in the inactivation effect of 50 to 150 μmol/L riboflavin VSV. When the light duration was 15 to 25 min, the inactivation effect of VSV was enhanced after the addition of riboflavin, but there was no difference between different concentrations of riboflavin groups. Conclusion The inactivation effect of riboflavin pathogen within 25 min is better with the increase of light time, and has little influence on riboflavin concentration.