Fluorescence cystoscopy (FC) efficiently enhances the detection and improves the therapeutic management of early bladder cancer. During an FC, about 150 ml of water is needed to inflate the bladder. The water is quickly diluted by urine which can be fluorescent. If this bladder washout fluid (BWF) becomes fluorescent, the FC images are frequently degraded. Unfortunately, it is unclear which elements of the diet may contribute to this background fluorescence. We propose to start this exploration with over-the-counter (OTC) vitamin supplements. To this end, we measured excitation–emission matrices of urine samples and the kinetics of modifications of urine fluorescence obtained from nine healthy volunteers before, during, and after intake of a commercially available OTC vitamin supplement. The pharmacokinetics shows that the BWF fluorescence values reach a maximum 8 to 10 h after vitamin intake. They decrease in the half-day that follows and reach values close to baseline day afterward. Based on these results, we conclude that, in order to avoid degradations of fluorescence images, it is likely best that the intake of OTC vitamin supplements be avoided during the week preceding an FC.