Abstract The phototoxic activity of xanthotoxin and visnagin plus NUV was considerably increased in arg-1 cells of Chlamydomonas reinhardii by additional dark periods. Pre-treatment in darkness before NUV irradiation, as well as NUV treatment, interrupted by several dark periods, had a greater toxic effect than a treatment of equal doses of NUV without dark periods. The effectiveness of the toxocity depended on the number of dark periods and/or on the length of the dark period. Prolonged NUV irradiation was less effective than additional dark periods compared with equal NUV doses. Interruption of NUV irradiation with dark periods or pre-treatment in darkness also clearly enhanced the mutagenic effectiveness of xanthotoxin plus NUV. On the other hand, a prolonged NUV irradiation without dark periods resulted in a slight reduction of Arg + revertants. In contrast with its toxicity, the weak photomutagenicity of visnagin was less influenced by additional dark periods than the stronger photomutagenicity of xanthotoxin. Based on these results we assume that the synergistic effect of dark periods was due to the absence of NUV. When xanthotoxin and visnagin are irradiated with NUV, dimers and polymers are formed. Dark periods may interrupt or retard this photoreaction. Therefore, the number of biologically active monomers may be higher in experiments with additional dark periods than in experiments without dark periods. It might therefore be concluded that, during treatment periods of equal length, more reversible bonds were formed with nucleic acids. These additional reversible bonds can be changed into irreversible bonds by radiation following the dark periods and thus enhance the activity of the drugs.