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JournalofApplied Phycology 8: 325-333, 1996.
© 1996 Kluwer Academic Publishers. Printedin Belgium.
Microscopic green algae and cyanobacteria in high-frequency intermittent light Ladislav Nedbal*, Vladimir Tichy1 , Fusheng Xiong' & Johan U. Grobbelaar2 1
NationalResearch Centerfor Photosynthesis and Global Climate Change, Institute of Microbiology AVCR, Opatovicky' mlin, 37981 Tiebon, Czech Republic 2 Departmentof Botany and Genetics, UOFS, Bloemfontein 9300, South Africa (*Author for correspondence;e-mail
[email protected]) Received 21 April 1996; revised 6 August 1996; accepted 28 August 1996
Key words: algae, cyanobacteria, growth rate, intermittent light, oxygen evolution, photoinhibition
Abstract The effects of fluctuations in the irradiance on Scenedesmus quadricauda, Chlorella vulgaris and Synechococcus elongatus were studied in dilute cultures using arrays of red light emitting diodes. The growth rate and the rate of photoinhibition were compared using intermittent and equivalent continuous light regimes in small-size (30 ml) bioreactors. The CO 2 dependent photosynthetic oxygen evolution rates in the intermittent and continuous light regimes were compared for different light/dark ratios and different mean irradiances. The kinetics of the electron transfer reactions were investigated using a double-modulation fluorometer. The rates of photosynthetic oxygen evolution normalized to equal mean irradiance were lower or equal in the intermittent light compared to the maximum rate found in the equivalent optimal continuous light regime. In contrast, the growth rates in the intermittent light can be higher than the growth rate in the equivalent continuous light. Photoinhibition is presented as an example of a physiological process affecting the growth rate that occurs at different rates in the intermittent and equivalent continuous lights. The difference in the dynamics of the redox state of the plastoquinone pool is proposed to be responsible for the low photoinhibition rates observed in the intermittent light. Abbreviations Fo, constant fluorescence measured with all QA oxidized; LED, light emitting diodes; PSII, Photosystem II; QA, primary quinone acceptor of the Photosystem II Introduction The capacity to adjust and operate in a fluctuating irradiance environment is one of the photosynthesis hallmarks that has been perfected during the evolution of plants. The elaborate physiological response to the seasonal and diurnal cycles, to the changing meteorological conditions, to the canopy motion or to the very rapid irradiance changes due to the refraction and reflection on a moving air-water surface is highly relevant in nature (Schenck, 1957; Dera & Gordon, 1968; Gordon et al., 1971) and, in the controlled cultivation and production of algal species, it is carried over from the natural environment to the bioreactor.
The individual algal cells are exposed to the fluctuating light environment in mixed bioreactors due to the transitions between the photic and aphotic layers of the bioreactors and, in some systems, also between the sun exposed cultivation area and the dark retention tank. In order to quantify the effects of an intermittent light regime on the yield of algal bioreactors, it is necessary to identify the frequency distribution of the light experienced by an individual cell, to separate the effects of the light fluctuations from those of mixing and to identify the physiological response of the specific algal strain to the dominant light frequencies (Richmond & Vonshak, 1978; Laws et al., 1983; Grobbelaar, 1994). With this information, the volume or areal productivity of the bioreactor can be modeled
326 as a function of the fluctuating light frequency and, eventually, the frequency distribution of the bioreactor can be optimized for maximal yields. In order to provide partial information aimed at this goal, the physiological response of two eukaryotic green algae and one cyanobacterial species to a well defined square-wave intermittent light was studied. The frequency of the applied intermittent light was higher than 0.1 Hz and slower than 10 kHz. The high frequency limit was chosen to be about one order of magnitude faster than the maximal operating frequency (rate constant) of the photosynthetic apparatus in saturating light (e.g., Whitmarsh, 1992). It can be expected that an intermittent light of this or higher frequency will be sensed by the algae as equivalent to the continuous light because the mobile electron carriers will be unable to respond to the fluctuations of this frequency range and will smooth the photochemical response. The frequencies close to the low frequency limit (10-0.1 Hz) are common both in nature and in the thin layer bioreactors (Setlik et al., 1970; Doucha & Livansky, 1995; Tichy et al., 1995; Grobbelaar et al. 1995). The high frequency limit can become relevant in space-based algal bioreactors using frequency modulated fluorescence tubes or the solid state light emitting diodes (Bula et al., 1991; Barta et al., 1992; Tennessen et al., 1995). Most of the existing information about the physiological response of algae to the high-frequency intermittent light has been accumulated in historical experiments contributing greatly to our present understanding of photosynthesis. The work of Warburg (1919) and Emerson & Arnold (1932) are only few examples of achievements reached using the intermittent light and algae (see also Myers, 1994 for review). It is well established that the physiology of plants and algae are different in fluctuating and continuous light (e.g. Weller & Franck, 1941; Walsh & Legendre, 1983; Falkowski, 1984; Grobbelaaret al., 1992; Kroon, 1994; Tennessen et al., 1995). Intermittent light has been suspected for a long time to be beneficial in reaching faster photosynthesis rates, higher yields and/or faster growth rates (e.g., Warburg 1919; Kok, 1953; Phillips & Myers, 1954; Laws et al., 1983; Terry, 1986). On the other hand, the notion of enhanced photosynthesis rates in the intermittent light over continuous light was critically evaluated by Sager and Giger (1980) and it should also be noted that photosynthetic rates on the individual cell level should not be confused with the increased rates measured in intermittent regime per area of the
bioreactor. The elevated rates in rapidly mixed bioreactors are in fact due to a light integration effect described in detail e.g. by Terry (1986). In spite of decades of intensive research, the operation of an individual algal cell in the intermittent light environment is not yet fully described. Namely, the photosynthesis rate should be defined as a function of the frequency, of the light/dark ratio and of the mean irradiance. The relation between photosynthetic and growth rates should be identified for various light regimes. The physiological mechanisms causing differences between the frequency dependence of photosynthetic and growth rates should be revealed.
Material and methods Test algae and the bioreactor The green algae Chlorella vulgaris and Scenedesmus quadricauda and the thermophilic cyanobacterium Synechococcus elongatus were grown in a turbidostat at about 500 imol photon m -2 s- l , for several days before being diluted to less than 20 M chl a ( 0
80
O _
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0 o
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TIME (h) Figure 8. Kinetics of photoinhibition of the Hill reaction activity in Scenedesmus quadricaudain the continuous (O) and intermittent light (0). The culture conditions are as in Figure 7.
LU
S. elongatus 100
Icn
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0 E C,
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0O
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TIME (h) Figure 9. Kinetics of photoinhibition of the Hill reaction activity in Synechococcus elongatus in the continuous (0) and intermittent light (0). The culture conditions are as in Figure 7.
1986). Invariably, the maximal rates in the intermittent light regime were reached at very short L/D periods of
