| Abstract |
The interactions of the photosynthetic process and in general the cellular metabolism of microalgae with the extracellular water environment (exchange of Η+ and e- - Chapters I and II) and their gaseous environment (exposure to extreme atmospheres, oxygenic and anoxic - Chapters III and IV), are presented and thoroughly analyzed in the four chapters of this Doctoral Thesis:
CHAPTER I: The microalgae photosynthetic process and the extracellular proton environment interact with each other. The photosynthetic process of microalgae causes an increase in pH in the culture medium as a result of cellular proton pumping rather than the effect of CO2 fixation. Photosynthetic water photolysis and the plastoquinone reduction/oxidation cycle provide protons in the lumen. Biological weak bases, such as polyamines, act as "permeable buffers" in the lumenduring the photosynthetic process, converting ΔpH to Δψ. This is probably the main reason for the continuous light-driven uptake of protons from the culture medium, through the cytoplasm and stroma, into the lumen. The rate of proton uptake and thus microalgal growth is proportional to light intensity, cell concentration, and extracellular proton concentration. Low pH in microalgal cultures, without light and nutrient limitations, strongly induces photosynthesis (and proton uptake) and, consequently, growth. In contrast, the mitochondrial respiratory process, in the absence of photosynthetic activity, does not substantially change the pH of the culture. The intensification of the respiration process by exogenous supply of glucose leads to significantly reduced pH values in the culture medium, almost exclusively through the production of protons. Increased concentrations of atmospheric CO2 dissolves in the water causing the "ocean acidification" effect by inhibiting the calcification process, very important for many phytoplankton and zooplankton organisms, but also for corals. The proposed new approach to the interaction of microalgal photosynthetic activity with proton concentration in the aquatic environment, independent of CO2 concentration, may be the Ariadne's thread to solve in the near future central environmental problems related to ocean acidification.
CHAPTER II: Given the above approach (of chapter I) we then raised the following question: If cells are able to produce a potential difference through photosynthesis, what would happen if we provided an electric current to photosynthesis (photosynthetic electron flow)? The idea to supply electrons of artificial origin (electric current) to the photosynthetic electron transport chain was realized by placing photosynthetic microalgae in an electrolytic assembly of an electro-photobioreactor [E(P)BR]. The results after a series of experimental approaches for the regulation of the concentration of nutrients and cells (regulation of electron conductivity) in relation to the supply of a constant intensity of electric current in microalgae culture highlighted the compatibility of channeling electric current in a biological chain of electron transport, such as that of photosynthesis process (and the respiratory process), inducing it significantly. Not only was the photosynthetic activity accelerated by the supply of appropriate current intensity, but also the production rate of bio-hydrogen (H2) was significantly increased under selected electric field conditions, proving that the additional (beyond electrolysis) hydrogen is of biological origin. The use of a special E(P)BR reactor with individual compartmentalization of the culture allows us to monitor the production of bio- hydrogen at the anode electrode, the cathode electrode and the space between the two electrodes. That allows us to discuss a possible bio-hydrogen production mechanism of hydrogen in an electric field. Combined human technology (electricity) with biology (photosynthetic and respiratory electron flow) opens up a new field of research with two main perspectives. The first concerns the use of photosynthetic microalgae to increase the efficiency of the electrolytic process in terms of hydrogen production (through the significant additional production of bio-hydrogen) making it energetically advantageous, and the second concerns a new perspective of rapid activation of the photosynthetic mechanism aiming to increase the biomass for its use for biotechnological purposes.
CHAPTER III: The interaction of protons (Chapter I) and electrons (Chapter II) with the photosynthetic mechanism of microalgae in a closed system indirectly highlighted the role of gas phase composition in photosynthesis. Therefore, in this chapter we examined the strategy of microalgae to cope with different atmospheres. The present study examines the survival strategy of microalgae in closed systems with anoxic atmospheres and the possibility of creating an oxygenic atmosphere through the photosynthetic management of solar radiation. The complete absence of CO2 appears to be dealt with by the microalga initially by catabolizing cellular organic matter through the respiration process (probably using NO2 instead of O 2), which produces CO2. This CO2 supports the photosynthetic process, which produces O 2, part of which is mainly used to enrich the atmosphere with O 2 and the rest for the respiration process and biomass production. Microalgae showed tolerance to exogenously supplied extreme CO2 concentrations (1%-40%) in an anoxic atmosphere and exhibited significantly higher photosynthetic activity compared to that of microalgae cultures grown in an oxygenic atmosphere. This response of microalgae to extremely high CO2 concentrations under anoxic conditions immediately enhances the atmospheric O2 level as well as culture growth without signs of stress. Since this microalgae strategy is particularly efficient at sequestering CO2 in a CO2 -rich atmosphere, it could help combat the greenhouse effect by incorporating microalgae cultures into gas detoxification systems with extremely high CO2 concentrations. The ability of microalgae to rapidly convert hostile atmospheres into O 2 -rich atmospheres could be used in the future to continuously recycle the atmosphere of human facilities on other planets. Furthermore, the increase of microalgae biomass under these conditions is an ideal combination for important (astro)biotechnological applications.
CHAPTER IV: As an extension of the results of chapter III was the investigation of the effect of an absolute H2 atmosphere on the photosynthetic organisms. The specificity of the effect of the H2 atmosphere on the photosynthetic organism and by extension on the entire cellular metabolism, forced us to check it separately. The exposure of microalgae and other (micro)organisms to an absolute hydrogen atmosphere stops both the photosynthetic and respiratory processes and, by extension, the complete cellular metabolism, stopping cellular growth for the desired period of time, extending their survival for long periods of time intervals and at the same time making them resistant to any stress conditions(metabolic arrest). The addition of oxygen or air restores completely the metabolic rate almost immediately and cell growth resumes normally from where it had stopped. Metabolism is the sum total of all the chemical reactions in life-sustaining organisms. There is an ongoing effort to control metabolic rate, which correlates with maximum lifespan potential and is one of the oldest scientific questions. In the present approach, we report for the first time the complete cessation of cellular metabolism and consequently cellular growth (metabolic arrest) of a range of organisms from microalgae to yeasts upon exposure to a 100% hydrogen atmosphere. Metabolic recovery is immediate with minimal O2 addition. Molecular dynamics simulations were used to decipher this phenomenon at the atomic scale. Several protein complexes, related to both photosynthetic and respiratory processes (LHCII, cytochrome c5, alanine dipeptide) were investigated for interaction with a 100% H2 atmosphere. Exposure to hydrogen, unlike oxygen, reduces protein residue fluctuations indicating thermostability. The whole phenomenon is not only limited to microorganisms, such as microalgae, but can also be extended to other (micro)organisms, but also to the preservation of a range of plant and other products (e.g. vegetables, fruits, ... etc.), opening new innovative paths of biotechnological applications.
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