**5. Simultaneous measurements of ETR and photosynthesis in wellwatered and moderately drought-stressed cotton plants**

Electron transport rate (ETR) and photosynthesis in cotton plants of both water treatments have been measured simultaneously for revealing the role and magnitude of alternative channels for utilization of the energy of electron transport and obtaining new insights into mechanisms of adaptation of the plant photosynthetic apparatus to long-term drought stress. Indicated photosynthesis parameters have been determined at CO2 concentrations 0–400 μmol mol-1 under constant PPFD of 1000 μmol m-2 s-1 and under PPFD of 0–2000 μmol m-2 s-1 at ambient CO2 concentration in plants of genotype Navbakhor (Figure 4). It is seen that the rate

Monitoring of the Drought Tolerance of Various Cotton Genotypes Using Chlorophyll Fluorescence http://dx.doi.org/10.5772/62232 101

**Figure 4.** Response of the photosynthesis, *A*G, electron transport rate, ETR, and photorespiration, estimated as ETR/4- *A*G, to CO2 concentration in leaves of the cotton genotype Navbakhor grown in well-watered (closed symbols) and moderately drought-stressed (open symbols) conditions in the field.

the photosynthetic oxygen evolution. In experiments, before measuring photoacoustic signals, the plant leaves were adapted to dark for 10 min. After reaching the steady-state photoacoustic signal, the saturating CW light was applied, which causes decrease in the photoacoustic signal for 0.82 (Figure 3a) in the well-watered leaf and for 0.50 (Figure 3b) in the dehydrated leaf. Thus, the photoacoustic measurements have shown that photosynthetic oxygen evolution in plant leaves depresses in short time water deficiency: decrease in the relative water content for 45% causes decrease of photosynthetic activity 1.5 times. Simultaneous measurements of ФPSII in these two samples displayed decline of the operative quantum efficiencies of photo‐ chemistry in the same ratio (0.75:0.51). However, the advantage of photoacoustic measure‐ ments is evident in the case of significant level of photorespiration in plant leaves, when direct correlation between ФPSII and the net photosynthesis is disturbed (see the next section).

**Figure 3.** Induction curves of the photoacoustic signal generated in leaves of the cotton genotype Navbakhor with rela‐ tive water content 100% (a) and 65% (b). Arrows up and down show switching on and off, respectively, the measuring

Time (s)

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**(a)**

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**5. Simultaneous measurements of ETR and photosynthesis in well-**

Electron transport rate (ETR) and photosynthesis in cotton plants of both water treatments have been measured simultaneously for revealing the role and magnitude of alternative channels for utilization of the energy of electron transport and obtaining new insights into mechanisms of adaptation of the plant photosynthetic apparatus to long-term drought stress. Indicated photosynthesis parameters have been determined at CO2 concentrations 0–400 μmol mol-1 under constant PPFD of 1000 μmol m-2 s-1 and under PPFD of 0–2000 μmol m-2 s-1 at ambient CO2 concentration in plants of genotype Navbakhor (Figure 4). It is seen that the rate

**watered and moderately drought-stressed cotton plants**

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(dashed arrows) and saturating (bolt arrows) lights.

100 Applied Photosynthesis - New Progress

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of CO2 assimilation (*A*G) increases linearly with increase of intracellular CO2 concentration, *C*<sup>i</sup> , while the dependence ETR *versus C*<sup>i</sup> is non-monotonic: sharp increase of ETR with increase of CO2 concentration at *C*<sup>i</sup> < 100 μmol mol-1, further saturates on the level of ETR ~200 μmol m-2 s-1. The measurements were carried out in the field, early morning, from 7.00 to 8.00 at temperature 22–24°C.

At higher light intensities and/or low CO2 concentrations, the plant photosynthetic apparatus cannot cope with the coming light energy and a portion of this energy has to be utilized through alternative channels; photorespiration or some other processes, including Mehler reaction, may play a role of a sink for electrons transported through the photosynthetic electron transport chain [55]. In most of the cases, excluding severe drought stress, the photorespiration considered as prevailing mechanism of utilization of such an excessive light energy [56]. The magnitude of this energy utilization may be estimated by comparing ETR and photosynthesis. Assuming that assimilation of one molecule CO2 requires four electrons transported through the chain, the amount of photorespiration may be defined by dividing ETR by four and subtracting the photosynthesis [55]. By calculating this way, values of the photorespiration rate are also presented in Figure 4: photorespiration increases sharply at low concentrations up to 100 μmol mol-1, and further slowly drops with increase of CO2 concentration. The fact seems reasonable, because CO2 is a product of photorespiration. Figure 4 shows that drought stress noticeably increases ETR and slightly decreases the photosynthesis in cotton plant leaves. As a result, the photorespiration in drought-stressed leaves calculated as above is considerably higher than in well-watered plants, especially at higher CO2 concentrations. In


**Table 2.** "Dark"" respiration, RD, and operating quantum efficiency of primary photochemistry in Photosystem II, ФPSII, measured in leaves of well-watered and moderately drought-stressed cotton genotype Navbakhor.

addition, the effect of drought stress to "dark" respiration has been measured in plants of both water treatments simultaneously with the quantum efficiency of primary photochemistry (Table 2). The "dark" respiration, as an additional source of bioenergy necessary for supporting vital biochemical reactions in plants, was considerably higher in drought-stressed plants. The same occurred with the quantum efficiency of photochemistry, but with less magnitude.

The light response of ETR and photosynthesis measured in plants of Navbakhor of the two treatments was similar to the CO2 response (Figure 5). At low light intensities, most of the energy from the electron transport is utilized in photochemical reactions, and with increasing of light intensity, more and more portion of this energy is spent for photorespiration. However, the increase in ETR induced by drought stress in light response was less expressed than that in CO2 response, particularly at higher intensities. Considerable variations of photosynthesis in different replications comparable with its difference between the treatments may be attributed to diurnal changes of stomatal conductance, *g*s, which can induce relevant changes in photosynthesis [57]. In view of tightly links between stomatal conductance and photosyn‐ thesis, and efficiency of primary reactions of photosynthesis remains constant, the changes in stomatal conductivity during the day may bring about considerable changes in photosynthesis

**Figure 5.** Response of the photosynthesis, *A*G, electron transport rate, ETR, and photorespiration, estimated as ETR/4- *A*G, to light intensity (PPFD) in leaves of the cotton genotype Navbakhor grown in well-watered (closed symbols) and moderately drought-stressed (open symbols) conditions in the field.

addition, the effect of drought stress to "dark" respiration has been measured in plants of both water treatments simultaneously with the quantum efficiency of primary photochemistry (Table 2). The "dark" respiration, as an additional source of bioenergy necessary for supporting vital biochemical reactions in plants, was considerably higher in drought-stressed plants. The same occurred with the quantum efficiency of photochemistry, but with less magnitude.

**Table 2.** "Dark"" respiration, RD, and operating quantum efficiency of primary photochemistry in Photosystem II, ФPSII,

**Water treatment RD ФPSII** Drought-stressed 3.8 ± 0.5 0.67 ± 0.023 Well-watered 5.2 ± 0.6 0.62 ± 0.021

measured in leaves of well-watered and moderately drought-stressed cotton genotype Navbakhor.

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moderately drought-stressed (open symbols) conditions in the field.

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102 Applied Photosynthesis - New Progress

The light response of ETR and photosynthesis measured in plants of Navbakhor of the two treatments was similar to the CO2 response (Figure 5). At low light intensities, most of the energy from the electron transport is utilized in photochemical reactions, and with increasing of light intensity, more and more portion of this energy is spent for photorespiration. However, the increase in ETR induced by drought stress in light response was less expressed than that in CO2 response, particularly at higher intensities. Considerable variations of photosynthesis in different replications comparable with its difference between the treatments may be attributed to diurnal changes of stomatal conductance, *g*s, which can induce relevant changes in photosynthesis [57]. In view of tightly links between stomatal conductance and photosyn‐ thesis, and efficiency of primary reactions of photosynthesis remains constant, the changes in stomatal conductivity during the day may bring about considerable changes in photosynthesis

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PPFD (µmol m–2 s–1)

**Figure 5.** Response of the photosynthesis, *A*G, electron transport rate, ETR, and photorespiration, estimated as ETR/4- *A*G, to light intensity (PPFD) in leaves of the cotton genotype Navbakhor grown in well-watered (closed symbols) and

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**Figure 6.** Light response of the chlorophyll fluorescence parameters: quantum efficiency of photochemistry in Photo‐ system II, ФPSII, photochemical quenching factor, *q*p, and non-photochemical quenching, NPQ, in leaves of the cotton genotype Navbakhor grown in well-watered (closed symbols) and moderately drought-stressed (open symbols) condi‐ tions in the field.

[58]. In this case, the sum of photosynthesis and photorespiration, as measured using the ETR/ 4, is not constant, but varies during the day.

In the Figure 6 are shown the light response of the three key fluorescence parameters, operating quantum efficiency of photochemistry, ФPSII, photochemical quenching factor, *q*p, and nonphotochemical quenching, NPQ, determined in leaves of well-watered and moderately drought-stressed cotton genotype Navbakhor. As shown from the figure, at low and moderate light intensities, PPFD < 800 μmol m-2 s-1, ФPSII in drought-stressed plants was higher than in well-watered plants, whereas *q*p was the same and near to its maximum. However, with increase of light intensity, ФPSII and *q*<sup>p</sup> decrease with increments, which are higher in droughtstressed plants. And finally, at PPFD > 800 μmol m-2 s-1, both ФPSII and *qp* become lower in drought-stressed plants in comparison with well-watered plants. What concerns to NPQ, it is negligibly low at low intensities in both treatments but increases rapidly at moderate and high intensities and under drought stress. So, increasing light intensity activates photosynthetic performance of plants. At low and moderate intensities, when the plant photosynthetic apparatus copes with coming light energy, the efficiency of photosynthetic conversion of light energy is very high, when photochemical quenching factor is near to its maximum and nonphotochemical quenching is negligibly low. Long-term drought stress due to stomatal and non-stomatal limitations to photosynthesis induces enhancement of photorespiration as an alternative sink for transported electrons in reaction centers of photosynthesis. However, further increase of light intensity increases non-photochemical quenching, and in droughtstressed plants, it is higher than in well-watered ones. This causes faster decrease of ФPSII and *q*p in drought-stressed plants.

Experiments with the measurement of chlorophyll fluorescence and the gas-exchange in different cotton genotypes showed that under drought stress, CO2 uptake slightly decreases, while ETR increases considerably. Simultaneously measuring these two parameters of photosynthesis allowed us to estimate the magnitude of photorespiration in the plant leaves, assuming that changes in the ETR/4-*A*<sup>G</sup> reflect the changes in photorespiration. Photorespira‐ tion increases with increasing light intensity and decreasing CO2 concentration. Moderate drought stress noticeably increases the rate of photorespiration, which can be considered as a characteristic response of C3 plants to a drought [44].

Leaves of drought-stressed cotton plants displayed higher ФPSII and photorespiration at low and moderate light intensities, and non-photochemical quenching, NPQ, was stronger in drought-stressed plant than that in well-watered one. Obviously, higher levels of photorespi‐ ration in plant leaves during the drought stress exerts the "pressure" to the rate of electron flow and makes Photosystem II to operate with higher efficiency.

#### **6. Conclusion**

The photosynthetic apparatus of plants supports higher performance of electron transport chain through enhancement of quantum efficiency of photochemistry in Photosystem II under drought stress. The accumulated energy in this state of over-excitation may be utilized in enhanced photorespiration. This protective reaction of the plant photosynthetic apparatus to drought stress has different magnitude depending on its drought tolerance. Field measure‐ ments of the chlorophyll fluorescence parameters simultaneously with morpho-physiological indicators of the cotton genotypes studied have displayed direct correlations between these parameters under drought stress. These correlations together with possible calibration of chlorophyll fluorescence parameters by photoacoustic characteristics determined at applica‐ tion of low-frequency-modulated light to plant leaves give new opportunities in monitoring of drought tolerance of various cotton genotypes in the field.
