The use of glyphosate in the desiccation of Urochloa brizantha cv. Marandu is a common practice in Brazilian agriculture; however, the effect of night application in this species is not clearly known. Thus, this study aimed to evaluate the influence of light availability on glyphosate efficiency in the control of U. brizantha. Seven herbicide doses (0, 90, 180, 360, 720, 1,440, and 2,160 g ha-1) in plants submitted to daytime application, followed by 24, 48, or 72 h of dark and night were evaluated. The accumulation of dry matter of U. brizantha, the visual evaluation of injuries, the doses that provided 50 and 80% of control (C50 and C80), and the doses that provided 50 and 80% reduction of dry matter (GR50 and GR80) at 7, 14, 21, 28 days after application (DAA) and at 30 days after cutting (regrowth) were determined. Satisfactory control (>80%) and the absence of regrowth were established from 1,440 g ha-1 in all treatments. However, injury reduction was observed in plants that remained 72 h in the dark relative to daytime treatment in all doses. The highest dose of glyphosate (2,160 g ha-1) reduced U .brizantha by 41, 15, and 3%. For the dose of 1,440 g ha-1 the reduction was 37, 16, and 6%, at 7, 14, and 21 DAA, respectively. The values of GR50 and GR80 decreased from regrowth (37 and 28% less relative to daytime treatment) for plants that were 72 h in the dark. The light favors the behavior of glyphosate in the plant, and therefore this herbicide should not be applied at night for the control of U. brizantha.