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Light-induced paramagnetism in chloroplasts. Proc Natl Acad Sci USA
... More recently, electromagnetic (low Hz to kHz frequencies) radiation was successfully applied in bone healing and osteoporosis treatment, with several FDA approved treatments, Tendons and muscles after sports injuries [16] also seem to heal faster, with less pain and swelling. ...
... Journal of Al-Nahrain UniversityVol.16 (4), December, 2013, pp.152-160Science ...
... 100X.Journal of Al-Nahrain UniversityVol.16 (4), December, 2013, pp.152-160Science ...
The aim of the study was to investigate the effect of magnetized water on the histological structure of the heart, lung, and spleen.
... Continuous-wave (CW) and pulsed electron paramagnetic resonance (EPR) spectroscopy have been employed to study the molecular structure and function of the Chl cofactors (Commoner et al., 1956;Kä b et al., 1995Kä b et al., , 1996Kä b and Lubitz, 1996). These methods have made it possible to probe the unpaired electron spin that is formed upon photo-oxidation or reduction of Chl in photosynthetic RCs. ...
... These methods have made it possible to probe the unpaired electron spin that is formed upon photo-oxidation or reduction of Chl in photosynthetic RCs. In fact, one of the earliest applications of EPR spectroscopy by Commoner et al. detected the steady-state photoaccumulation of the primary donor cation, P 700 $+ , of PS I (Commoner et al., 1956). Subsequently, the signal was attributed to a strongly electronically coupled Chl a dimer (Chl 1A and Chl 1B in Figure 1B) (Norris et al., 1971;Davis et al., 1993;Mac et al., 1998;Kä ss et al., 2001), given that the line width was narrower than that of a monomeric Chl a (Norris et al., 1971). ...
This research addresses one of the most compelling issues in the field of photosynthesis, namely, the role of the accessory chlorophyll molecules in primary charge separation. Using a combination of empirical and computational methods, we demonstrate that the primary acceptor of photosystem (PS) I is a dimer of accessory and secondary chlorophyll molecules, Chl2A and Chl3A, with an asymmetric electron charge density distribution. The incorporation of highly coupled donors and acceptors in PS I allows for extensive delocalization that prolongs the lifetime of the charge-separated state, providing for high quantum efficiency. The discovery of this motif has widespread implications ranging from the evolution of naturally occurring reaction centers to the development of a new generation of highly efficient artificial photosynthetic systems.
Video abstract:
... In the initial experiments [1], the leaves were lyophilized before insertion into the cavity, and hence kinetic studies on the specimens were not possible. In a more recent series of experiments, however, Commoner, Heise and Townsend [36] were able to study aqueous suspensions of chloroplasts, in situ in the cavity, under different conditions of illumination. This was achieved by the use of very small specimen tubes in conjunction with a high-sensitivity 100-kc/s field modulation spectrometer. ...
... The variation of radical concentration with intensity and the wavelength of the incident illumination were also studied [36]. The concentration was found to rise with an increasing intensity but reached a saturation value at high levels, and this is also true of the photosynthetic activity of both chloroplasts and whole cells. ...
... EPR signals belonging to Z/D intermediates were observed for the first time in spinach chloroplasts already in 1956 [27]. They are long living radicals which could be recorded at room temperature by continual wave (cw) EPR spectrometer. ...
... The EPR spectrum of oxidized core of PS 1 (P700 + ) belongs to the first observed EPR signals. Commoner and Heise [27] called it as signal I with g = 2.0026 and line width 0.8 mT. This signal is well visible in Fig. 3C (both lines). ...
... The above observations can be explained by assuming that some organelles of plant cells (i.e. mitochondria) possess paramagnetic properties as found in chloroplasts by Commoner et al. (1956). This is consistent with findings that mitochondria are sensitive to magnetic fields (Belyavskaya, 2004). ...
... This should not be surprising because both mitochondria and chloroplasts have electron transport chains imbedded in their membranes generating an electrochemical potential gradient for protons across the membranes. Commoner et al. (1956) suggested that metabolically active tissues of plant cells contain free radicals and they play an important role in electron transfer and in the kinetics of the chemical reactions. 6. ...
Difficult-to-germinate seeds are a common plant conservation, plantation and biotechnological problem. Oil palm (Elaeis guineensis) seed germination takes 6 months to 1 year with ≈40% failure rate. We investigated the affect of various electromagnetic DC field strengths on oil palm germination. Kernels imbibed water more effectively when exposed to a magnetic field. The kernels were divided into three groups. The first group (50) was soaked (imbibed) in distilled water (2 h) with no exposure to applied magnetic fields (blank control). The second group was exposed to magnetic fields with intensities of 2.5 mT, 5.0 mT, 7.0 mT, 9.0 mT and 11.0 mT for 1, 2, 3, 4 and 5 h (Dry Treated Kernels, DTK). The same electromagnetic protocol was conducted on a third group but with kernels which were immersed in water (Magnetically-Treated-Water-Kernels, MTWK). 96% germination was achieved by day-30 for the MTWK treatment using 9.0 mT for 4 h: slightly lower results were found for DTK kernels. None of the controls germinated within 30 days. Young oil palm seedlings kept in a shade house (≈110 mol quanta m −2 s −1 PPFD) watered every day with MTW grew 3 times faster (3.1 mm day −1) than controls watered using unmagnetised water.
... The essence of van Niel's formulation, illustrated in Fig. 1, is that neither CO2 nor the H-donor substrate (H2A in a genera] formulation) participates directly in a photochemical 1 This symposium was held at the Annual Meeting of the American Society for Microbiology, Chicago, Ill., 24 April 1961, under the sponsorship of the Division of Agricultural and Industrial Bacteriology, with Robert L. Starkey as convener. 2 An abundance of very recent information will necessitate reappraisal of many areas in this mobile field of investigation. ...
... [23] 2.2 In-cell EPR EPR studies in cellular contextsd ate back to more than 60 years. The light-induced paramagnetism of chloroplast observed by Commoner et al. [24] or the investigation of nitroxide behavior in cells, tissues, and whole animals for EPR and magnetic resonance imaging (EPRI and MRI, respectively)a re some examples. [25] However,i nt he past decade, the floweringo f structurals tudies of biological macromolecules in their native environment was associated with the introduction of the concept of "in-cell spectroscopies", such as in-cell fluorescence resonance energy transfer (FRET), in-cell NMR, and in-cell EPR spectroscopy.T hus,i nt his paper we adopted the "in-cell" definition for structurals tudies of proteins or nucleic acids, inside cells. ...
Exploring the structure and dynamics of biomolecules in the context of their intracellular environment has become the ultimate challenge for structural biology. As the cellular environment is barely reproducible in vitro, investigation of biomolecules directly inside cells has attracted a growing interest. Among magnetic resonance approaches, site‐directed spin labeling (SDSL) coupled to electron paramagnetic resonance (EPR) spectroscopy provides competitive and advantageous features to capture protein structure and dynamics inside cells. To date, several in‐cell EPR approaches have been successfully applied to both bacterial and eukaryotic cells. In this review, the major advances of in‐cell EPR spectroscopy are summarized, as well as the challenges this approach still poses.
... Two years later, Commoner reported that EPR signals consistent with free radicals could be detected in isolated tobacco-leaf chloroplasts upon irradiation with an automobile headlamp. [6] The EPR signals reached a maximum value after about 20 s of irradiation, then decayed exponentially with a 45-s time constant when the lamp was switched off. ...
Prior to 1950, the consensus was that biological transformations occurred in two-electron steps, thereby avoiding the generation of free radicals. Dramatic advances in spectroscopy, biochemistry, and molecular biology have led to the realization that protein-based radicals participate in a vast array of vital biological mechanisms. Redox processes involving high-potential intermediates formed in reactions with O2 are particularly susceptible to radical formation. Clusters of tyrosine (Tyr) and tryptophan (Trp) residues have been found in many O2-reactive enzymes, raising the possibility that they play an antioxidant protective role. In blue copper proteins with plastocyanin-like domains, Tyr/Trp clusters are uncommon in the low-potential single-domain electron-transfer proteins and in the two-domain copper nitrite reductases. The two-domain muticopper oxidases, however, exhibit clusters of Tyr and Trp residues near the trinuclear copper active site where O2 is reduced. These clusters may play a protective role to ensure that reactive oxygen species are not liberated during O2 reduction.
The study analyses the impact of alternating (magnetic induction B = 30 mT for t = 60 s) and constant magnetic fields (B = 130 mT for t = 17 h) and alternating electric fields (electric current E = 5 kV/cm for t = 60 s) on various growth parameters of soy plants: the germination energy and capacity, plants emergence, the fresh mass of seedlings, protein content (Kjeldahl’s method), and photosynthetic parameters (with MINI-PAM 2000 WALTZ Photosynthesis Yield Analyser and a SPAD-502 Chlorophyll Meter). Four cultivars were used: MAVKA, MERLIN, VIOLETTA, and ANUSZKA. Moreover, the advanced Machine Learning processing pipeline was proposed to distinguish the impact of physical factors on photosynthetic parameters. The use of electromagnetic fields had a positive impact on the germination rate in MERLIN seeds. The best results in terms of germination improvement were observed for alternating magnetic field stimulation in all cultivars (p > 0.05). For the VIOLETTA cultivar an increase (p > 0.05) in the emergence and overall number of plants as well as fresh mass was observed after electromagnetic field stimulation. For the MAVKA and MERLIN cultivars, the concentration of proteins in the leaves was noticeably higher in plants grown from seeds stimulated using a constant magnetic field.
Depicting how biomolecules move and interact within their physiological environment is one of the hottest topics of structural biology. This Feature Article gives an overview of the most recent advances in Site-directed Spin Labeling coupled to Electron Paramagnetic Resonance spectroscopy (SDSL-EPR) to study biomolecules in living cells. The high sensitivity, the virtual absence of background, and the versatility of spin-labeling strategies make this approach one of the most promising techniques for the study of biomolecules in physiologically relevant environments. After presenting the milestones achieved in this field, we present a summary of the future goals and ambitions of this community.
Light-induced reactions in photosynthetic reaction centers are initiated by the absorption of a photon, which results in the transfer of a single electron and the generation of radical ions in the donor and acceptor molecules involved in the charge-separated state. Electron paramagnetic resonance (EPR) spectroscopy is the ideal method for the study of such reactions. In addition to measuring spectra of the electron transfer cofactors in continuous light, reactions can be initiated by brief flashes of light, thereby allowing the kinetics of forward electron transfer as well as recombination reactions to be obtained. Because the donor and acceptor pairs are so closely spaced and because light induced charge separation is so rapid, the donor and early acceptors are in a quantum mechanically spin entangled state, which confers properties such as increased sensitivity, the ability to measure reactions on the nanosecond timescale, and the determination of bond angles between cofactors. Additionally, distances between pairs of cofactors can be measured by detecting the modulation of a phase shifted “out-of-phase” electron spin echo signal. In this methods article, we will describe how continuous wave EPR, time resolved EPR, and pulsed EPR can be used to measure these properties in Type I photosynthetic reaction centers. Methods of analysis are described for the bound electron transfer cofactors in the heterodimeric Photosystem I reaction center of plants and cyanobacteria and in the homodimeric reaction centers found in phototrophic members of the phyla Bacillota, Chlorobiota, and Acidobacteriota.
Electron spin resonance measurements of normal tissue at 77°K indicate the presence of two types of resonances which can be identified by varying the incident microwave power: (i) an intense, easily saturable, organic free-radical component and (ii) a weak but relatively nonsaturating component probably due to paramagnetic trace elements.
Chlorophylls (Chl)s exist in a variety of flavors and are ubiquitous in both the energy and electron transfer processes of photosynthesis. The functions they perform often occur on the ultrafast (fs – ns) time scale and until recently, these have been difficult to measure in real time. Further, the complexity of the binding pockets and the resulting protein-matrix effects that alter the respective electronic properties have rendered theoretical modeling of these states difficult. Recent advances in experimental methodology, computational modeling, and the emergence of the new X-ray crystal structures of reaction centers (RC) have renewed interest in these processes and allowed researchers to elucidate previously ambiguous functions of Chls and related pheophytins. This is complemented by the wealth of experimental data obtained from decades of prior research. Studying the electronic properties of Chl molecules has advanced our understanding of both the nature of the primary charge separation and subsequent electron transfer processes of RCs. In this review, we examine the structures of primary electron donors in Type I and Type II RCs in relation to the vast body of spectroscopic research that has been performed on them to date. Further, we present density functional theory calculations on each oxidized primary donor to study both their electronic properties and our ability to model experimental spectroscopic data. This allows us to directly compare the electronic properties of hetero- and homodimeric RCs.
In 1944, electron paramagnetic resonance (EPR) was discovered by Evgenii Konstantinovich Zavoisky in the USSR (Union of the Soviet Socialist Republics). Since then, magnetic resonance methods have contributed invaluably to our knowledge in many areas of Life Sciences and Chemistry, and particularly in the area of photosynthesis research. However, the road of the magnetic resonance methods, as well as its acceptance in Life Sciences and Chemistry, was not smooth and prompt in the (former) USSR. We discuss the role played by many including Jakov K. Syrkin, Nikolai N. Semenov, Vladislav V. Voevodsky, Lev A. Blumenfeld, Peter L. Kapitza, and Alexander I. Shalnikov during the early stages of biological and chemical EPR spectroscopy in the USSR.
Barry Commoner’s scientific career is best characterized by his insistent commitment to holistic (as opposed to reductionist) approaches to understanding how living things function and his alertness in bringing the most modem tools from physics and chemistry to bear on the properties of living systems. The pioneering work of his laboratory on the life history of tobacco mosaic virus was widely admired. In addition, his was the first work which used a magnetic resonance technique to investigate biological phenomena. Characteristically, these studies utilized whole, living, functioning organisms. He pushed the limits of sensitivity for measuring small differences in the nitrogen isotopic composition of drainage water to investigate the relative contributions of fertilizer N to the high nitrate levels found in an agricultural watershed. While the conclusion that N applied as fertilizer was responsible for about half of the nitrate pollution initially met with fierce resistance, the methods which he conceived are now widely used. His ideas about the multiple roles of DNA in inheritance were sti1lless warmly received. While being involved in all of this, the Barry whom many of you know best found the time and energy to be a major figure in bringing the dangers of radioactive fallout to public attention, to be a vigorous and effective opponent of the war in Vietnam and to play a leading role in establishing the environmental movement.
Die den Stoffwechsel bedingende molekulare Struktur und Mikro-Architektur der Zelle zeigt in den höheren Tieren und Pflanzen wie auch in den Mikroorganismen eine große Einheitlichkeit. Bei aller Verschiedenheit der Gestalt ist diese grundlegende Übereinstimmung einer der stärksten Beweise für die Einheit des Lebendigen, so wie wir es von unserer Erde her kennen. Zu erörtern, ob es eine andere Art von chemischer Organisation des Lebens gibt, liegt zur Zeit völlig im Bereich der Spekulation. Die Frage, warum sich der aller Gestaltung zugrunde liegende Stoffwechsel so einheitlich ausgebildet hat, kann zur Zeit wissenschaftlich nicht beantwortet werden. Sind solche universal verwendeten Erfindungen wie die des Porphyrinsystems wegen eines einzigartigen Nutzeffektes selektiv immer wieder zur Herrschaft in verschiedensten Stoffwechselbereichen gekommen, oder ist eine solche Erfindung nur einmal im Zuge des Lebens durch einen Zufall gemacht worden, oder entspricht ihr Dasein einer Notwendigkeit, die in den thermodynamischen Vorteilen solcher Moleküle begründet liegt, so daß Leben, so oft es auch entstehen mag, immer in der gleichen Weise zu solchen optimalen Organisationsformen gelangen müßte? Diese Fragen vermögen wir heute noch nicht zu entscheiden. Aber Einheitlichkeit des Grundstoffwechsels und die erstaunliche Ähnlichkeit der Feinstrukturen aller Zellen machen heute eine allgemeine Biologie zu einer Wirklichkeit und nicht nur zu einem Wunschbild. Deshalb ist es auch möglich, Reaktionen des Grundstoffwechsels und allgemeine Prinzipien der Physiologie an den verschiedensten Organismen zu studieren. Das in dem einen Bereich des Lebendigen Erkannte kann auch in anderen Bereichen angewandt werden. Der Biochemiker sucht sich von Fall zu Fall Objekte aus ganz verschiedenen Bezirken des Lebendigen. Biochemie und Genetik geben die Mittel in die Hand, die alle Wissenschaft gefährdende Sammlung von Vielfalt durch übergeordnete Gesichtspunkte zu überwinden. Dadurch wird der Mannigfaltigkeit erst der richtige Rang eingeräumt als Ausdruck der Variabilität durchgehender Gesetzmäßigkeiten.
Summary This document is part of Volume 1 ‘Magnetic Properties of Free Radicals’ of Landolt-Börnstein - Group II Molecules and Radicals.
We present historic discoveries and important observations, related to oxygenic photosynthesis, from 1727 to 2003. The decision to include certain discoveries while omitting others has been difficult. We are aware that ours is an incomplete timeline. In part, this is because the function of this list is to complement, not duplicate, the listing of discoveries in the other papers in these history issues of Photosynthesis Research. In addition, no one can know everything that is in the extensive literature in the field. Furthermore, any judgement about significance presupposes a point of view. This history begins with the observation of the English clergyman Stephen Hales (1677–1761) that plants derive nourishment from the air; it includes the definitive experiments in the 1960–1965 period establishing the two-photosystem and two-light reaction scheme of oxygenic photosynthesis; and includes the near-atomic resolution of the structures of the reaction centers of these two Photosystems, I and II, obtained in 2001–2002 by a team in Berlin, Germany, coordinated by Horst Witt and Wolfgang Saenger. Readers are directed to historical papers in Govindjee and Gest [(2002a) Photosynth Res 73: 1–308], in Govindjee, J. Thomas Beatty and Howard Gest [(2003a) Photosynth Res 76: 1–462], and to other papers in this volume for a more complete picture. Several photographs are provided here. Their selection is based partly on their availability to the authors (see Figures 1-15). Readers may view other photographs in Part 1 (Volume 73, Photosynth Res, 2002), Part 2 (Volume 76, Photosynth Res, 2003) and Part 3 (Volume 80, Photosynth Res, 2004) of the history issues of Photosynthesis Research. Photographs of most of the Nobel-laureates are included in Govindjee, Thomas Beatty and John Allen, this volume. For a complementary time line of anoxygenic photosynthesis, see H. Gest and R. Blankenship (this volume).
Before discussing the many and varied applications of E.S.R., it may be just as well to mention the very first application of this technique by Zavoiskii1 in 1945, as this gives the underlying principle used, namely the study of energy level separations between the Zeeman levels in paramagnetic materials.
Oil palm (Elaeis guineensis) is widely grown in southern Thailand as an important economic crop for bioenergy, human consumption and industrial use. Reliable production of young oil palm trees is limited and the plants are expensive because it takes up to 8-12 months for germination to occur. In our study, control seed kernels imbibed with normal water (NW) had zero germination even after 56 days. Kernels were immersed in magnetically-treated water (MTW) for 48 hours to imbibe. The treated kernels were placed in germinators and sprayed with MTW or NW. Oil palm kernels imbibed and watered with MTW had 100% germination within 35 days. Comparison of seeds (a) imbibed with MTW and watered with MTW, (b) imbibed with MTW and then watered with NW and (c) imbibed with NW and then watered with MTW showed that the critical treatment stage was the imbibition step. Simply soaking seeds or kernels in MTW for an imbibition treatment is a simple technique to apply in nurseries. Any gains of applying MTW after the imbibition step lead to only modest increases in germination rate.
Oil palm (Elaeis guineensis) is widely been grown in southern Thailand as an important economic crop for bio-energy, human consumption and industrial uses. Reliable production of young oil palm trees is limited and the plants are expensive because it takes up to 8-12 months for germination to occur. The slow germination rate and the high failure rate (approximately 40% failure rate). In our study, control seed kernels imbibed with normal water (NW) had zero germination even after 56 days. Kernels were immersed in magnetically-treated water (MTW) for 48 hours to imbibe the seed kernels. The treated kernels were placed in germinators and sprayed with MTW or NW. Oil palm kernels imbibed and watered with MTW had 100% germination within 35 days. Comparison of seeds (a) imbibed with MTW and watered with MTW, (b) imbibed with MTW and then watered with NW and (c) imbibed with NW and then watered with MTW showed that the critical treatment stage was the imbibition step. Simply soaking seeds or kernels in MTW for an imbibition treatment is a simple technique to apply in nurseries. Any gains of applying MTW after the imbibition step leads to only modest increases in germination rate.
Durch die Photosynthese bzw. Kohlensäureassimilation werden in der pflanzlichen Zelle unter Wirkung des im Chorophyll absorbierten Sonnenlichts aus Kohlendioxyd und Wasser energiereiche Kohlenhydrate aufgebaut. Die Pflanze wandelt somit über diesen wichtigsten photochemischen Prozeß, den die Natur kennt, Lichtenergie in chemische Energie um, die dann beim Kohlenhydratabbau dem Organismus zugeführt wird.
The influence of frequent magnetic field stimulation (MFS) on plants is the subject of intense research. The effects of MFS on plants vary depending on its intensity, time of exposure or application form. The effects of low-frequency magnetic field in two doses, 0.2 mT, 16 Hz (MFS-1) and 0.2 mT, 50 Hz (MFS-2) on the mitotic activity and selected physiological and biochemical parameters in narrow-leafed lupin (Lupinus angustifolius L.) were evaluated. Non-exposed plants were used as control (C). It was noted that after the exposure of plants to MFS-1, the biometric parameters, mitotic activity, BSA and GPOX activity remained at the control level. However, a significant decrease in the assimilation pigment content was observed. On the other hand, the exposure of plants to MFS-2 was manifested by a decrease in the biometric parameters, mitotic activity and the assimilation pigment content, but an increase in GPOX activity in roots was noted.
This historical minireview describes basic lines of progress in our understanding of the functional pattern of photosynthetic water oxidation and the structure of the Photosystem II core complex. After a short introduction into the state of the art about 35 years ago, results are reviewed that led to identification of the essential cofactors of this process and the kinetics of their reactions. Special emphasis is paid on the flash induced oxygen measurements performed by Pierre Joliot (in Paris, France) and Bessel Kok (Baltimore, MD) and their coworkers that led to the scheme, known as the Kok-cycle. These findings not only unraveled the reaction pattern of oxidation steps leading from water to molecular oxygen but also provided the essential fingerprint as prerequisite for studying individual redox reactions. Starting with the S. Singer and G. Nicolson model of membrane organization, attempts were made to gain information on the structure of the Photsystem II complex that eventually led to the current stage of knowledge based on the recently published X-ray crystal structure of 3.8 Å resolution in Berlin (Germany). With respect to the mechanism of water oxidation, the impact of Gerald T. Babcock’s hydrogen abstractor model and all the considerations of electron/proton transfer coupling are outlined. According to my own model cosiderations, the protein matrix is not only a ‘cofactor holder’ but actively participates by fine tuning via hydrogen bond networks, playing most likely an essential role in water substrate coordination and in oxygen-oxygen bond formation as the key step of the overall process.
The physical and chemical mechanisms of primary photon capture and utilization for energy storage in photosynthesis, although enigmatic, are of paramount significance for maintenance of life on Earth. Photosynthesis by green plants and algae is an essential component of the food chain for higher mammals and man. Furthermore, a primary source for molecular oxygen is photosynthesis performed by marine microorganisms and in the tropical rainforests. The agricultural importance of photosynthesis cannot be disputed in that expanding population pressures on the world’s limited food sources urgently necessitate the development of more efficient crops. Increased comprehension of the photosynthetic process and its regulation by the plant will permit eventual genetic selection or engineering for food crops exhibiting high yield, rapid maturation, and resistance to climatic variations.
During the last two years significant advances have been made towards an understanding of biophysical aspects of photosynthesis. Only part of these will be discussed in this review, which will be confined to primary photoreactions and associated electron transport. The first part will deal with recent results concerning photosynthesis of oxygenevolving organisms (algae and higher plants); the second part with bacterial photosynthesis, a subject that was not treated in the previous review (AMESZ).
The first publication on the use of ESR techniques in connection with biological systems appeared only nine years after the first demonstration of paramagnetic resonant absorption.† In that first report, a variety of biological systems, such as leaves, seeds, and tissue preparations, were shown to contain free radicals. A definite correlation was found between the concentration of the radicals and the metabolic activity of the material. This work appears to confirm earlier ideas that free radicals are involved as intermediates in metabolic processes. However, the question as to which free radicals are involved in a given metabolic process proved to be a much more difficult question to answer.
For magnetic resonance studies of paramagnetic species electron paramagnetic resonance (EPR) is a well-established method. However, when trying to elucidate the electronic structure of large and lowsymmetry radicals, as they typically occur in biological systems, one is often hampered by problems of spectral resolution. It was as early as 1956 when Feher (1956) demonstrated that by electron nuclear double resonance (ENDOR) the spectral resolution can be greatly improved. ENDOR signals are obtained by monitoring the changes of the amplitude of a saturated EPR line that occur when sweeping the frequency of an additionally applied rf field through the nuclear (NMR) region. This first ENDOR experiment was technically feasible only because the sample—phosphorus doped silicon—was studied at low temperature, where all the relaxation times are sufficiently long to easily obtain saturation. For radicals in liquid solution, however, these relaxation times are much shorter—on the order of 10-5-10-7 sec—and, consequently, ENDOR-msolution experiments are technically much more sophisticated since much larger saturating microwave and rf fields have to be applied. This probably explains why the first ENDOR-in-solution experiments required many more years before they could be successfully performed by Cederquist (1963) and by Hyde and Maki (1964).
The chapter discusses electron paramagnetic resonance in photosynthetic studies and the discussion is primarily concerned with the observations which have been made on the photo-induced, unpaired electrons in photosynthetic materials, and with the correlations which have been made between the resonance observations and biological parameters. The various kinds of electrons which can be studied using electron paramagnetic resonance techniques have been mentioned. A discussion is made on the nature of the electron paramagnetic resonance (EPR) experiment by which these various types of unpaired electrons are observed, how this type of experiment has been applied to photosynthetic systems, and the conclusions concerning photosynthesis that can be drawn from the experiments. The parameters associated with the observation of unpaired electrons are described, with emphasis on those aspects of the observation which become important when the sample material is polycrystalline or amorphous. The types of photosynthetic materials on which resonance experiments have been performed have been listed. Then in varying degrees of detail, discussion on some of the experiments which have been performed has been presented. The treatment deals with those experiments that seemed to be most pertinent. Finally, a brief appendix is attached in which several terms are defined, and various experimental details are described.
This chapter discusses quantum conversion in chloroplasts. The primary quantum conversion process involves the most important case of radiation catalysis. The reduction of the carbon dioxide by a reducing agent to produce carbohydrate is a separate and independent reaction, independent at least of the primary light absorption and conversion process. The other half of the photosynthesis process—the primary quantum conversion—has been associated with the photolysis of water. In addition, the primary quantum conversion act of photosynthesis in chemical terms is an ionization occurring in a charge transfer complex. However, this cannot occur in isolated charge transfer molecules in solution or in a homogeneous single-phase crystal because the products cannot escape from each other. The primary quantum conversion act, as it occurs in modern photosynthesis, can only take place in a laminated structure where the electrons and holes can escape from each other by electron migrations and not by atomic migration.
1.1. Procedures are described for the preparation of chloroplasts capable of carrying out three photochemical reactions, each representing an increasingly complex phase of photosynthesis: photolysis of water (Hill reaction), esterification of inorganic phosphate into adenosine triphosphate (photosynthetic phosphorylation) and the reduction of carbon dioxide to the level of carbohydrates with a simultaneous evolution of oxygen.2.2. The three photochemical reactions were separable by variations in the technique for preparation of chloroplasts and by differential inhibition by several reagents. Inhibition of a more complex phase of photosynthesis does not affect the similar one which precedes it and, conversely, the inhibition of a simpler phase of photosynthesis is paralled by an inhibition of the more complex phase which follows.3.3. Reversible inhibition of CO2 fixation and photosynthetic phosphorylation, but not of photolysis, by sulfhydryl group inhibitors suggests that sulfhydryl compounds (enzymes, cofactors, or both) are involved in phosphorylation and CO2 fixation, but not in the primary conversion of light into chemical energy as measured by the Hill reaction.4.4. Evidence is presented in support of the conclusion that the synthesis of ATP by green cells occurs at two distinct sites: anaerobically in chloroplasts, by photosynthetic phosphorylation, and aerobically in smaller cytoplasmic particles, presumably mitochondria, by oxidative phosphorylation independent of light.5.5. A general scheme of photosynthesis by chloroplasts, consistent with these findings, is presented.
HIGHLY purified `malic' enzyme from pigeon's liver1 mediates a fixation of carbon dioxide through catalysis of reaction (1). When, by coupling with a suitable dehydrogenase system, triphosphopyridine nucleotide is continuously reconverted to its reduced form, malate is readily synthesized from pyruvate and carbon dioxide2. Should illuminated chloroplast preparations be able to effect a reduction of pyridine nucleotides, then the photochemical reaction could be coupled to reductive carbon dioxide fixation by a well-defined enzyme system2,3. Such a mechanism might operate in photosynthesis. A `malic' enzyme specific for triphosphopyridine nucleotide is present in the tissues of higher plants4.
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