Main conclusion Three species chosen as representatives of NADP-ME C4 subtype exhibit different sensitivity toward photoinhibition, and great photochemical differences were found to exist between the species. These characteristics might be due to the imbalance in the excitation energy between the photosystems present in M and BS cells, and also due to that between species caused by the penetration of light inside the leaves. Such regulation in the distribution of light intensity between M and BS cells shows that co-operation between both the metabolic systems determines effective photosynthesis and reduces the harmful effects of high light on the degradation of PSII through the production of reactive oxygen species (ROS). AbstractWe have investigated several physiological parameters of NADP-ME-type C4 species (e.g., Zea mays, Echinochloa crus-galli, and Digitaria sanguinalis) grown under moderate light intensity (200 µmol photons m−2 s−1) and, subsequently, exposed to excess light intensity (HL, 1600 µmol photons m−2 s−1). Our main interest was to understand why these species, grown under identical conditions, differ in their responses toward high light, and what is the physiological significance of these differences. Among the investigated species, Echinochloa crus-galli is best adapted to HL treatment. High resistance of the photosynthetic apparatus of E. crus-galli to HL was accompanied by an elevated level of phosphorylation of PSII proteins, and higher values of photochemical quenching, ATP/ADP ratio, activity of PSI and PSII complexes, as well as integrity of the thylakoid membranes. It was also shown that the non-radiative dissipation of energy in the studied plants was not dependent on carotenoid contents and, thus, other photoprotective mechanisms might have been engaged under HL stress conditions. The activity of the enzymes superoxide dismutase and ascorbate peroxidase as well as the content of malondialdehyde and H2O2 suggests that antioxidant defense is not responsible for the differences observed in the tolerance of NADP-ME species toward HL stress. We concluded that the chloroplasts of the examined NADP-ME species showed different sensitivity to short-term high light irradiance, suggesting a role of other factors excluding light factors, thus influencing the response of thylakoid proteins. We also observed that HL affects the mesophyll chloroplasts first hand and, subsequently, the bundle sheath chloroplasts.