2.3. Chlorophyll a fluorescence measurements and investigation of non-photochemical quenching (NPQ) parameters

Chlorophyll a fluorescence was analysed on intact leaves using a PAM-101–102-103 fluorometer (Walz, Effeltrich, Germany) after 20 min of dark adaptation. The maximum quantum efficiency of PSII (F_v/F_m) was calculated from the values of minimal (F₀) and maximal fluorescence (F_m) as F_v/F_m = (F_m-F₀)/F_m. F₀ was determined by switching on the measuring light, with a modulation frequency of 1.6 kHz and PPFD of less than 1 μmol m⁻² s⁻¹, after 3 s illumination with far-red light in order to eliminate reduced electron carriers. In the light-adapted state the actual quantum efficiency (ΔF/F_m') of photosystem II (PSII) was calculated from the values of the steady-state value (F_s) and maximal fluorescence (F_m’), as ΔF/F_m' = (F_m’-F_s)/F_m’. The F_m and F_m’ values were measured using a 0.7 s flash of white light at PPFD 3500 μmol mmol m⁻² s⁻¹ (light source: KL 1500 electronic, Schott, Mainz, Germany; Solti et al. 2008). For quenching analysis the leaves were exposed to actinic white light (PPFD of 100 mmol photons m⁻²s⁻¹) for 20 min. Excitation energy allocations were calculated according to Hendrickson et al. (2005), where the parameters were: Φ_f,D – fluorescence and constitutive thermal dissipation; Φ_NF – thermal dissipation in non-functional PSII; Φ_NPQ – light-dependent and ΔpH- and xanthophyll-mediated thermal dissipation; Φ_PSII – PSII electron flow. The distribution of the parameters was calculated according to the following equation: 1 = Φ_PSII + Φ_f,D + Φ_NPQ + Φ_NF.