For 20 weeks during the growing season, cuttings of one birch clone (Betula pendula Roth.) were exposed in the Birmensdorf fumigation chambers to O₃-free air (control) or 75 nl O₃ l⁻¹. Ozone was supplied either from 1900 until 0700 h (nighttime regime), from 0700 until 1900 h (daylight regime), or all day (24-h regime). By autumn, reductions in whole-plant biomass production, root/shoot biomass and stem weight/length ratios were evident in all three O₃ regimes. The reductions in cuttings receiving the 24-h O₃ treatment were about twofold larger than in cuttings receiving the daylight O₃ treatment. Stomata were open at night, and stomatal conductance was about 50% of its maximum daytime value. We calculated that the rate of O₃ uptake into leaves in the dark approached 4 nmol m⁻² s⁻¹. Whole-plant production and carbon allocation were more sensitive to O₃ during the night than during the day; however, O₃ exposure caused similar visible leaf injury in both of the 12-h regimes, although the leaves exposed to O₃ at night exhibited delayed O₃-induced shedding. Overall, changes in production and carbon allocation were determined by the external O₃ dose rather than by the kind of O₃ exposure, indicating that, at the seasonal scale, the internal dose of ozone that was physiologically effective was a constant fraction of the external O₃ dose. We conclude that nighttime O₃ exposures should be included in the daily time period for determining critical concentrations of O₃ causing injury in trees.