## 4 Discussion  
The FLD-LISP method was suggested as an effective method to measure ChlF parameters using the FLD approach. In this research, FLD-LISP was evaluated and the results were compared with ChlF parameters measured by the standard PAM method. First, the FLD-LISP ChlF yield was measured using the O\({}_{2}\)A band. To determine the saturation light pulse intensity to be used for FLD-LISP, the relationships between the actinic pulse PPFD and ChlF yields in dark-adapted and solar light-adapted paprika leaves were initially studied (Figure 3). The photosynthesis saturation PPFD of many leaves is in the range of 500-1000 \(\upmu\)mol m\({}^{-2}\) s\({}^{-1}\), which is considerably lower than the direct solar PPFD of 2000 \(\upmu\)mol m\({}^{-2}\) s\({}^{-1}\)[69]. [PERSON] et al. (2009) [18] presented that ChlF yields of the Boston fern leaf abaxial side, leaf mesophyll, and guard cell chloroplasts became constant at the saturation pulse PPFD of 800 \(\upmu\)mol m\({}^{-2}\) s\({}^{-1}\) in dark-adapted conditions and 1300 \(\upmu\)mol m\({}^{-2}\) s\({}^{-1}\) in light-adapted conditions. In this research a saturation pulse of \(>\)800 \(\upmu\)mol m\({}^{-2}\) s\({}^{-1}\) in the dark and 1800 \(\upmu\)mol m\({}^{-2}\) s\({}^{-1}\) under solar PPFD of 700 to 1300 \(\upmu\)mol m\({}^{-2}\) s\({}^{-1}\) was sufficient to saturate the photosynthetic electron transport in paprika leaves (Figure 3). This is because adequately photoactivated PSII requires stronger light; the excess light energy is dissipated as heat [7,70]. The relationship between PPFD and ChlF yield at 760.4 nm using FLD-LISP confirms the results of other studies. The saturation pulse of the red laser at the PPFD of 6000 \(\upmu\)mol m\({}^{-2}\) s\({}^{-1}\) for 0.8 s was used for all three plant species in this research, which is considerably larger than the photosynthesis saturation PPFDs of many leaves [69]. The FLD-LISP method was applied to measure the ChlF parameters of \(\Phi_{\text{PSII}}\), NPQ, and ETR in three plant species, paprika (C3 plant), maize (C4 plant), and pachira (indoor C3 plant), having different carbon fixation mechanisms and light tolerance. The relationships between PPFD and FLD-LISP derived ChlF parameters of \(\Phi_{\text{PSII}}\), NPQ, and ETR for all three plant species were studied. In general, under high PPFD, excess light energy is dissipated as heat through the NPQ process that is regulated by the acidification of the thylakoid lumen attributed to the accumulation of protons leading to a \(\Delta\)pH [4], which results in an increase in NPQ. This is an indicator of the portion of the absorbed radiation not used for electron transport in photosynthesis [4,7,71]. The regulatory protein PsbS and the conversion of violaxanthin to zeaxanthin in the xanthophyll cycle are involved in this process [4]. For both C3 and C4 plants, \(\Phi_{\text{PSII}}\) generally decreases whereas NPQ increases with increasing illumination. Also under the same level of illumination, leaves with a lower photosynthetic capacity exhibit lower \(\Phi_{\text{PSII}}\) and higher NPQ. These facts have been reported previously in various ChlF studies at leaf and cell levels [1,72,73,74,75,76,77,78,72]. The relationships between PPFD and the ChlF parameters of \(\Phi_{\text{PSII}}\), NPQ, and ETR measured using FLD-LISP in all three plant species (Figures 4-6) were similar to the relationships obtained through the PAM measurement. This confirms that the FLD-LISP method is capable of measuring ChlF parameters accurately under different PPFDs. Both \(\Phi_{\text{PSII}}/\text{PPFD}\) and NPQ/PPFD relationships were found to be linear for paprika and maize under ordinary sunlight intensity (Figures 4A,B and 5A,B). For maize, the \(\Phi_{\text{PSII}}\) values were higher and the NPQ values were lower especially in high PPFDs in comparison with paprika. These results are acceptable as paprika is a C3 plant whereas maize is a C4 plant [79,80].