Photosynthetic parameters, growth, and pigment contents were determined during expansion of the fourth leaf of in vitro photoautotrophically cultured Nicotiana tabacum L. plants at three irradiances [photosynthetically active radiation (400-700 nm): low, LI 60 µmol m-2 s-1; middle, MI 180 µmol m-2 s-1; and high, HI 270 µmol m-2 s-1]. During leaf expansion, several symptoms usually accompanying leaf senescence appeared very early in HI and then in MI plants. Symptoms of senescence in developing leaves were: decreasing chlorophyll (Chl) a+b content and Chl a/b ratio, decreasing both maximum (FV/FM) and actual (ΦPS2) photochemical efficiency of photosystem 2, and increasing non-photochemical quenching. Nevertheless, net photosynthetic oxygen evolution rate (PN) did not decrease consistently with decrease in Chl content, but exhibited a typical ontogenetic course with gradual increase. PN reached its maximum before full leaf expansion and then tended to decline. Thus excess irradiance during in vitro cultivation did not cause early start of leaf senescence, but impaired photosynthetic performance and Chl content in leaves and changed their typical ontogenetic course. and B. Radochová, I. Tichá.