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This thesis is concerned with the development of new methodologies in the field of Light-induced Pulsed Electron Spin Resonance Dipolar Spectroscopy (LiPDS), with the aim of applying them to the structural investigation of complex biological macromolecular systems with endogenous or artificially-added chromophores that can be photoexcited to form triplet states. The orientational effects in previously reported LiPDS techniques and how to exploit them to extract additional molecular conformational information are explored in Chapter 2. The ReLaserIMD and LiDEER techniques are combined to fully characterise the relative orientation of the two spin centres in porphyrin-nitroxide model peptides, and a new two-dimensional LiPDS technique (frequency-correlated ReLaserIMD) is introduced to capture the complete orientational dependence of the dipolar interaction with a single experiment. These novel methodologies are extended to non-porphyrin chromophores. Chapter 3 goes a step further to tackle the challenge of measuring the dipolar interaction between two photogenerated triplets, removing the need for having any permanent spin centre in LiPDS. The new Light-Induced Triplet-Triplet Electron Resonance spectroscopy (LITTER) technique is developed using bis-porphyrin model peptides and an in-depth conformational study of a spectroscopic molecular ruler is performed exploiting the orientational effects. LITTER is also compared to conventional DEER spectroscopy, showing superior results, and is applied to the structural study of a dimeric heme protein and a supramolecular porphyrin tetramer. Chapter 4 addresses some of the limitation of the LITTER technique by using non-identical chromophores and two-colour photoexcitation in bis-labelled model peptides, improved performance and direct combination with F¨orster Resonance Energy Transfer. This thesis constitutes an important step forward in the knowledge and development of the novel field of LiPDS, by introducing new triplet spin labels, improving existing techniques, developing new experiments and analysis methods, and combining them with other experimental tools for integrative structural biology.

Many animals, and in particular migratory birds, have the ability to use the geomagnetic field to navigate. The leading hypothesis to explain the origin of their compass sense is based on magnetosensitive radical pairs (RPs) formed in cryptochrome flavoproteins. Numerous studies have demonstrated that the reactivity of light-induced RPs within flavin-based model systems and cryptochromes could be influenced by the magnitude of a magnetic field, giving rise to magnetic field effects (MFEs). Yet, many questions remain unanswered, especially regarding the ability of these systems to detect the direction of the field, which has been shown to provide critical information for navigation. This work examines different chemical and spectroscopic approaches for their poten- tial to yield or elucidate directionally sensitive magnetic field responses in flavin-based RPs. Chapter 1 explores whether selectively manipulating the magnetic properties of cryptochromes in vivo could disturb the bird's migratory behaviour. If it did, it would demonstrate that cryptochromes are i) sensitive to the field direction and ii) the primary magnetic sensor for avian magnetoreception. The potential of this ambitious experiment is assessed by examining the impact of isotopic substitution (which changes the hyperfine interaction driving the RPs' magnetosensitivity) on the MFE of model systems. In vitro, the directionally dependent response of RPs to a field (anisotropic MFEs) can only be detected if the radicals are immobilised and oriented. In Chapters 2-3, several matrices are investigated as potential immobilisation media and the unexpected behaviour of flavins in glycerol solutions is further examined. Chapter 4 explores direct molecular alignment via protein crystallisation and introduces confocal microscopy as a powerful technique for the study of MFEs in crystals. By exploiting the capabilities of this technique, the MFEs of several cryptochromes, including avian cryptochromes, are investigated and contrasted, leading up to the examination of anisotropic MFEs in cryptochrome crystals (Chapter 5).

Many animal species have the ability to navigate using the Earth's magnetic field. This is particularly true of migratory birds, who are able to fly thousands of kilometres only to nest in exactly the same place year on year. The best current hypothesis as to the method by which this magnetoreception - the ability to sense magnetic fields - takes place is that of magnetically sensitive radical pairs in cryptochromes - blue light photoreceptors found in the retinae of birds and other animals. Chapter 1 explores the theory needed for understanding the phenomenon of magnetoreception, especially spin chemistry. The study of magnetically sensitive processes requires the ability to detect tiny changes in the concentrations of the chemical species involved. This has driven the development of state-of-the-art and high-sensitivity techniques, with a particular focus on optical spectroscopy. One such method is the use of cavity-based absorption measurements which increase the optical path length by passing the probe light through the sample multiple times. In this work, a spectrometer that employs the use of an optical cavity - cavity ring-down spectroscopy (CRDS) - was entirely redesigned, rebuilt and recoded. The work to improve the spectrometer, along with an introduction to the optical technique, is described in Chapter 2. In Chapter 3, the performance of the spectrometer is scrutinised using a model system - flavin mononucleotide (FMN) and hen egg-white lysozyme (HEWL) - allowing for exploration of the detection sensitivity and stability of the apparatus. Time-resolved and magnetically altered reaction yield (MARY) experiments yield results similar to those reported previously, particularly by Dr D. Sheppard. The sensitivity of the rebuilt spectrometer is highlighted, revealing the importance of storage conditions of this widely used "calibration sample" with a slight deviation in the time- and wavelength-resolved spectra obtained when compared to previously reported results. Once shown that the spectrometer is fully functional, the effect of pump laser power on ring-down time is also explored with the hope of improving the signal-to-noise ratio by compensating for fluctuations from the laser. The core of the thesis describes the measurements performed on cryptochrome 4 from three avian species: the migratory European robin (Erithacus rubecula); the pigeon (Columba livia), which is non-migratory, but known to orient using magnetic cues; and the chicken (Gallus gallus), which is not known to use the geomagnetic field for any purpose. The cryptochromes from the three species share strongly conserved amino acid sequences, but should, to support the cryptochrome hypothesis of magnetoreception, show differences that are based on the function. A marked difference in the spectroscopic results is shown between the migratory robin and the other two species. Additional measurements are performed on a point-mutant of the three cryptochromes in which the final electron transfer partner - a tryptophan residue at position 369 - is replaced so that it cannot be involved in forming the radical pair. It is shown that, while all cryptochrome proteins show some sensitivity of their radical concentrations to applied magnetic fields, that of the migratory robin is most remarkable - both in its magnitude and lifetime. The results of this thesis, presented in Chapter 4, hence provide further support of the radical pair hypothesis of magnetoreception and the data and conclusion formed a substantial part of a Nature publication in 2021.

The research presented in this thesis is concerned with the applications of Electron Paramagnetic Resonance (EPR) spectroscopy to the study of porphyrin systems and cryptochrome proteins. In particular, the fine and hyperfine interactions measured using EPR are exploited to further our understanding of both the intricate electronic and spatial structure of these systems as well as their potential future applications. In a first project, the 1H and 14N hyperfine couplings in the radical cations of oligoporphyrin molecular wires were determined using continuous-wave EPR (cwEPR) as well as pulsed Electron-Nuclear Double Resonance (ENDOR). The degree of spin delocalisation, which quantifies the viability of these systems as nano-wires, can be elucidated via these hyperfine couplings. A comparison of a previously investigated butadiyne-linked oligoporphyrin series with a novel series of ethyne-linked analogues revealed that the trends in the cwEPR spectral envelopes of the latter deviate from the trends expected for complete spin delocalisation, in contrast to the former series which follows the theoretical trend neatly. The consequences of these cwEPR envelope trends to the spin delocalisation in both series is explored in detail from the perspective of Density Functional Theory (DFT) calculations, complementary ENDOR measurements, and a re-examination of the original theory behind the cwEPR envelope analysis method. The applicability of trEPR and ENDOR to studying complex spin density trends was further exploited in the investigation of the photo-excited triplet states of palladium(II), zinc(II), free-base, and mesoper fluoro oligoporphyrin systems. For these triplet states, the information obtained from ENDOR about the hyperfine couplings is complemented by the fine-structure parameters, i.e. the g-tensor and zero- field splitting (ZFS) tensor, and the non-Boltzmann (spin polarised) populations of the triplet spin sublevels, all of which dominate their trEPR spectral signature. In the palladium(II) systems, the spin-orbit interaction due to this heavy metal ion is shown to dominate all the fine-structure and spin polarisation parameters. However, ENDOR and electronic structure calculations reveal that the spin density in the Pd2+ systems is conserved relative to the Zn2+ and free-base porphyrins. This means that the Pd2+ ion could replace the Zn2+ in molecular wires when a particular type of spin polarisation pattern is desired without compromising on spin delocalisation. In view of the potential to control spin delocalisation in nano-wires, the trends in the fine-structure parameters of meso-perfluoro/acceptor substituted porphyrin systems was also investigated. The sigma- acceptor groups are shown to disturb the planarity of the porphyrin backbone, causing significant orthorhombic distortions in the symmetry of the spin density. The implications of these distortions on the future use of meso-perfluoro groups in molecular wires, as well as on the continuing role of EPR spectroscopy in their study, is reviewed in light of the experimental observations. The last chapter continues the exploitation of light induced spin polarisation but turns its attention to spin-correlated radical pairs in a series of cryptochrome proteins, which could be responsible for the ability of songbirds to sense and exploit the Earth's magnetic field for navigation. The fine and hyperfine interactions obtained by new simulation methods will be linked to the all-essential spin-correlation and to the intimate spatial structure of the radical pairs.

The study of spin chemistry and, in specific, magnetosensitivity has been growing in importance in recent years. Not only has there been great advances in the field of magnetic field effects (MFEs) on biological reactions, but also on their applications in semiconductor and polymer science. Many chemical systems exhibit sensitivity to comparatively weak magnetic fields by a variety of mechanisms. The most prevalent of such magnetic field effect (MFE) mechanisms in solution is the radical pair (RP) mechanism, which will be investigated in this work. Chapter 1 will introduce ideas behind the RP-based magnetosensitivity: the quantum mechanics of spin, the theory of long-range electron transfer, chemical kinetics and finally the RP-mechanism itself. The chapter will conclude by describing the magnetosensitivity of flavin-based systems investigated in this work. A significant part of this work revolved around the understanding, using and development of the methodology involved. Chapter 2 will discuss some of the mathematical and computational methods used throughout this work. The methods covered are the time-frequency analysis, fitting, numerical solutions to differential equations and analysis of uncertainty. The examples used throughout this chapter have been selected to illustrate important points of their respective subjects, but also to be relevant to understanding other parts of this work. Chapter 3 will make use of some of the methodology described in the previous chapter and talk about the development and theoretical treatment of a modulated fluorescence experiment - ModMARY. The chapter will conclude by showcasing the methodology developed to measure a fluorescent exciplex-based system traditionally investigated by ModMARY pyrene/1,3-dicyanobenzene. The work done in this chapter is the basis of a publication led by the author. The flavin-based system often used in magnetosensitivity studies of biological systems is the solution of flavin mononucleotide (FMN) and hen egg-white lysozyme (HEWL). This system is well known, but not fully understood. The interactions of freely diffusing FMN within a crystal lattice of HEWL has been investigated in Chapter 4, by confocal laser-scanning microscopy (CLSM). The principle of CLSM, as well as the data analysis methodology developed, will be discussed. Then, the spatiotemporal evolution of the MFE of FMN in HEWL crystals will be explored. The work in this chapter is part of a joined publications with Dejéan et al. MFEs can be probed optically not only by fluorescence, as investigated in the aforementioned chapters, but also by absorption techniques. The understanding and development of broadband cavity-enhanced absorption spectroscopy (BBCEAS), and its application to measurements of solution-based cryptochrome samples will be discussed in Chapter 5. The systems of most interest in the field of magnetoreception are the cryptochrome molecules - flavin-binding proteins proposed as the source of the avian magnetic sense. The methodology from previous chapter will be used in Chapter 6 to measure the samples of cryptochrome proteins from five different animal species and their selected mutants. This data will be compared and its implications will be discussed. The work is a part of a joined publications with Xu et al.

Some animals, such as migratory birds, are known to have the ability to sense and use magnetic fields for orientation. At the heart of this remarkable ability is believed to be a photoinduced chemical reaction taking place within a blue-light receptor protein called cryptochrome (Cry). It is hypothesised that magnetic fields can play a decisive role in determining reaction rates (and, hence, yields) by acting on the spin states of spin correlated radical pairs (RPs) formed between a flavin chromophore and a tryptophan (Trp) electron-transfer (ET) chain within Crys. In this work, cavity ring-down spectroscopy (CRDS) has been used for the detection of magnetic field effects (MFEs) via small changes in the differential absorbance (ΔΔA~10−6) of aqueous protein samples, with sub-microsecond time resolution. The photochemistry and effects of magnetic fields on European robin (Er) Cry, fruit fly Cry, and a plant Cry are characterised. In a key result for this field of research, an MFE in the photochemistry of ErCry is revealed for the first time, lending considerable support to its proposed involvement in bird magnetoreception. Using Cry mutants with modified Trp ET chains, the role of individual Trps in the manifestation of MFEs was explored. Replacing the terminal (i.e. fourth) Trp of the ET chain in ErCry with a redox inert amino acid is shown to significantly increase the optically-detected MFE, indicating that the RP formed between the flavin and the third Trp might be the primarily magnetically sensitive species. To better understand the relevant biochemical processes in Cry, de novo designed artificial flavoproteins (maquettes) have been used to model the photochemistry of Cry (Chapter 4). The comparatively simple and adaptable design of these model proteins facilitates detailed MFE studies by reducing the complexity of natural Cry. In these maquettes, photoinduced ET leads to the formation of a RP which exhibits MFEs analogous to those in Crys. The profound effect of the donor-acceptor distance on the MFE is demonstrated using flavomaquettes with varying flavin-Trp distances. The application of cavity enhanced spectroscopy to both natural cryptochromes and artificial flavoproteins, opens up new pathways for the detection and characterisation of MFEs in biologically relevant environments. The enhanced sensitivity of these approaches constitutes a significant step forward in understanding the underlying biochemical processes of animal magnetoreception.

The studies presented in this thesis deal with the investigation of inter- and intramolecular interactions in porphyrin nanostructures and molecular triad systems using a variety of different electron paramagnetic resonance (EPR) based techniques as well as optical spectroscopy. Dipolar and exchange couplings were investigated in a range of linear and cyclic copper-containing porphyrin nanostructures by continuous wave (cw) and pulse EPR. The magnitude of the exchange interaction, J, and trends in this property could be determined by numerical simulations of the cw EPR spectra or pulse dipolar evolution data. Using double electron-electron resonance (DEER), through-bond exchange coupling in a bis-copper six-porphyrin nanoring could directly be measured over a distance of ~4 nm. By comparison of the results with those obtained for a singly connected but otherwise identical system, the J-coupling in this ring structure containing two identical, parallel paths was further shown to be the result of constructive quantum interference. In a ten-membered porphyrin nanoring containing two copper and eight zinc centres, the interaction between the central metals of the porphyrins and axial nitrogen ligands of different molecular templates was investigated by cw EPR, electron nuclear double resonance (ENDOR) and DEER. The weak coupling between copper and axial nitrogen ligands could be detected and it was found that the difference in binding affinity between copper and zinc to axial nitrogen ligands influences the structure of the formed complexes and preferred location of the copper centres. We were able to show that the template design allows control over the complex geometry on a molecular level. Triplet state delocalisation was investigated in a series of ladder complexes, formed between linear zinc porphyrin oligomers and bidentate nitrogen ligands, by transient cw EPR and pulse ENDOR techniques. It was found that even though the porphyrin units are held in an approximately co-planar arrangement within the ladder complex, which should favour long-range electronic communication, complete delocalisation is restricted to two porphyrin units at low temperatures and no delocalisation across the bridging ligand was observed. In a different project on symmetric and asymmetric zinc porphyrin oligomers with varying side and end groups, the influence of electronic symmetry on triplet state delocalisation was determined to be the major factor governing and limiting the extent of triplet state delocalisation in linear porphyrin oligomers. Triplet states of zinc porphyrins with different side groups have also been investigated in liquid crystal solvents by EPR. The alignment of the porphyrins is observed to depend strongly on the side groups of the porphyrin core. The orientation and conformations of the porphyrins could be obtained by simulation of the transient cw EPR spectra and order parameters were determined. The influence of small (< 10 G) magnetic fields on the recombination kinetics of spin-correlated radical pairs formed upon photo-excitation of molecular triad systems was studied by transient absorption techniques. Molecular triads serve as model compounds for the study of anisotropic magnetic field effects, which could previously only be detected at comparatively high fields of >30 G. Here it is shown for the first time that hyperfine-driven magnetic field effect anisotropy is observable at magnetic fields comparable to that of the Earth (~0.5 G), underlining the feasibility of a chemical compass for avian magnetoreception.

Magnetic fields as weak as the Earth's can change the yields and/or kinetics of radical pair reactions even though the interaction energies involved are orders of magnitude smaller than kT at room temperature. Proposed as the source of the light-dependent magnetic compass in migratory birds, Magnetic Field Effects (MFEs) on chemical reactions can be understood by the radical pair mechanism (Chapter 1) and are thought to operate in flavin-containing protein photoreceptors in the retina. Numerous studies on flavin-based model systems of biological importance have shown MFEs under physiological conditions. In many instances, these effects are small and are only likely to be of any importance to the postulated chemical compass if the primary effects of the magnetic fields on the geminate radical pairs are further enhanced via yet unknown amplification mechanisms. In this thesis, a novel technique is introduced which measures MFEs on the prompt fluorescence of continuously photoirradiated flavin/electron donor model systems (Chapter 2). By exploiting the sensitivity of this method it is possible to probe the effects of the magnetic interaction characteristics within the radical pair as well as those of the embedding media. In Chapter 3, the flavin-ascorbic acid system and rare examples of Low Field Effects relevant for magnetosensitivity to weak magnetic fields under biologically relevant conditions are explored. The possibility for the chemical amplification of MFEs is demonstrated for cryptochromes and related model systems through experiment and simulation (Chapter 4-5). Under conditions of continuous photoexcitation, any change in the kinetics of radicals formed downstream to the geminate radical pair can significantly enhance the prompt field effect over the timescale of milliseconds and longer. Given the efficiency of amplification and the simplicity of its implementation, it is not inconceivable for nature to have adopted these effects for improved magnetoreception.

The research described in this thesis employs a variety of Electron Paramagnetic Resonance (EPR) techniques for the study of the electronic and structural properties of artificial supramolecular porphyrin systems and of protein complexes of biological relevance. The electron delocalisation in the cationic radical and photoexcited triplet states of linear and cyclic Π-conjugated multiporphyrin arrays was investigated. In the radical cations, information on the extent of delocalisation can be inferred from the measurement of hyperfine couplings, either indirectly from the continuous wave EPR spectrum or directly using pulsed hyperfine EPR techniques. The results of room temperature EPR experiments showed complete delocalisation of the electron on the timescale of the EPR experiments, but frozen solution EPR measurements revealed localisation onto mainly two to three porphyrin units in the larger porphyrin systems. Information on the delocalisation of the triplet state in the same porphyrin systems is contained both in the hyperfine couplings and in the zero-field splitting (ZFS) interaction. The results outlined in this thesis show that the hyperfine couplings provide a much more accurate estimate of the extent of delocalisation. The trends in proton and nitrogen hyperfine couplings with the size of the porphyrin systems indicate uneven spin density distributions over the linear arrays, but complete delocalisation in the cyclic systems. Time-resolved EPR and magnetophotoselection experiments have shown a reorientation of the zero-field splitting tensor between a single porphyrin unit and longer linear arrays, resulting in alignment of the main optical transition moment and the Z axis of the ZFS tensor. Continuous wave and pulsed dipolar EPR techniques were employed for the determination of the structure of two different protein complexes, the homomultimeric twin-arginine translocase A (TatA) protein channel and the ferredoxin-P450 complex of the electron transport chain in Novosphingobium aromaticivorans. The interaction between nitroxide spin labels introduced at different positions of the TatA monomer was investigated in the complex reconstituted in detergent micelles by analysing the dipolar broadening of the EPR spectra and the results of three- and four-pulse Double Electron-Electron Resonance (DEER) measurements. In combination with results from NMR and molecular dynamics calculations, a structure for the channel complex was proposed. The structure of the ferredoxin-cytochrome P450 complex was investigated by orientation-selective DEER between nitroxide labels introduced on the cytochrome P450 protein and the iron-sulfur cluster of the ferredoxin. The distance and orientation information contained in the experimental DEER data was interpreted in terms of a structural model of the protein complex by orientation-selective DEER simulations combined with a modelling approach based on protein-protein docking.

Molten globules are compact intermediates in the protein folding process which retain native-like secondary structure, but have a fluctuating ensemble of tertiary folds. Human alpha-lactalbumin is a Ca2+ binding protein which forms a molten globule at pH 2. Although much is known about the secondary structure of the alpha-lactalbumin molten globule, the tertiary structure has so far not been elucidated. Presented in Chapter 2 is a double electron-electron resonance study of alpha-lactalbumin. The C-helix was found to be fully folded in the molten globule, in agreement with previous studies. Furthermore, it is also shown that both the N- and C-termini are in close proximity to the C-helix, forming part of the hydrophobic core of the protein. Data collected in the presence of 8M urea show that the protein is unfolded under these conditions, and distinguishes the molten globule state from this. These data provide quantitative information about the tertiary structure of the alpha-lactalbumin molten globule, and may provide important insight into the protein folding process. Although a wide variety of chemical and biological systems are known to be sensitive to the strength of an applied magnetic field, no biological systems, such as the cryptochrome proteins believed to be the magnetoreceptors in migratory birds, have yet been shown to be sensitive to the field direction. Traditionally, the anisotropic response of such systems to the magnetic field has been probed using photoselection or alignment with liquid crystals. The experimental sensitivity to anisotropy could, however, be improved by increasing the degree of molecular alignment, such as by crystallisation. Presented in Chapter 3 is a study of hen egg white lysozyme crystals doped with FMN, which together form a magnetically sensitive system. The crystals were studied using confocal microscopy, with the magnetic field effect being spatially resolved. The diffusion of FMN through the crystals was also studied, with the spatial variation of the magnetic field effect being explained with reference to the diffusion of FMN. Although no anisotropy of the magnetic field effect was observed in this system, the use of crystallisation as a method of controlling molecular orientation is potentially useful in systems which are expected to show anisotropy (such as cryptochrome). The behaviour of many chemical systems differs between the bulk solution and at the interface. These interfacial processes can be difficult to probe using conventional absorption spectroscopy due to the relatively low absorbance of species at the interface as compared to the bulk. Cavity ring down and cavity enhanced absorption spectroscopy are two techniques which increase the sensitivity of absorption measurements, and evanescent wave variants of both have been applied to interfacial studies. Presented in Chapter 4 is a polarisation sensitive, broad band, evanescent wave cavity enhanced absorption experiment, which allows the study of the spectra of species at the silica/solvent interface across the visible region. A variety of systems which show interesting inter- facial behaviour have been studied, including crystal violet at the silica/water interface, and with polyelectrolyte films. The data could be fitted to extract spectra, kinetics, and average orientations of three individual species. These studies demonstrate the power of the polarisation sensitive, broad band nature of this experiment in determining complex interfacial behaviour involving more than one species.

This thesis discusses, employs and develops further applications of both Electron Paramagnetic Resonance (EPR) and Nuclear Magnetic Resonance (NMR) methodology in the study of a variety of biological systems. Chemically Induced Dynamic Nuclear Polarization (CIDNP) is used to study the surface accessibility of Tryptophan and Tyrosine residues in the Equine Lysozyme-Oleic Acid complex (ELOA). It is believed that these amino acids may play a vital role in the interaction of this partially structured complex with cell walls leading to the apoptosis of the cell. Double Electron-Electron Resonance (DEER) is an EPR technique that can be used to study the distance and, in some cases, the orientation between two paramagnetic centres on a nano-meter length scale. Attaching spin labels, such as Methane Thiosulphonate Spin Label (MTSL), to proteins, or by using spin active cofactors in proteins as innate spin labels, allows the measurement of distances within proteins or their complexes. In this thesis, model copper systems were initially studied in order to gain a better understanding of the orientational information obtainable from such spectra. The methodology developed from this project was applied to the protein−protein complexes of ferredoxin reductase−ferredoxin and ferredoxin−P450, three proteins that form a vital electron transport chain, leading to the catalytic hydroxylation of many small molecules. The structure of the docked P450−ferredoxin complex is the first experimentally determined structure of its type and thus is of considerable biochemical interest. As part of the structural determination of the ferredoxin complexes, it was necessary to measure the relative orientation of the g-matrix with respect to the molecular structure. This was achieved using a combination of orientationally selective proton Electron Nuclear Double Resonance (ENDOR) and HYperfine Sublevel CORrElation (HYSCORE) spectroscopy. The expansion of the DEER technique to measure an ever-increasing variety of systems has shown the limitations of the pulse sequences most frequently employed at present, particularly in relation to the measurement of many of the metal centres, which could be used as innate spin probes in protein systems. Methodological work has also been undertaken to examine possible solutions to circumvent some of these problems, namely low modulation depths and fast relaxation times. This has resulted in the development of two modifications to the existing techniques: Dead-time free three-pulse DEER and Repeated Excitations IN DEER (REINDEER). These techniques have been used to study of both synthetic model systems and also spin labelled proteins.

Magnetic fields influence the rate and/or yield of chemical reactions that proceed via spin correlated radical pair intermediates. The field of spin chemistry centres around the study of such magnetic field effects (MFEs). This thesis is particularly concerned with the effects of the weak magnetic fields B₀ ~ 1mT relevant in the ongoing debates on the mechanism by which animals sense the geomagnetic field and on the putative health effects of environmental electromagnetic fields. Relatively few previous studies have dealt with such weak magnetic fields. This thesis presents several new theoretical tools and applies them to interpret experimental measurements. Chapter 1 surveys the development and theory of spin chemistry. Chapter 2 introduces the use of Tikhonov and Maximum Entropy Regularisation methods as a new means of analysing MARY field effect data. These are applied to recover details of the diffusive motion of reacting pyrene and N,N-dimethylaniline radicals. Chapter 3 gives a fresh derivation and appraisal of an approximate, semiclassical approach to MFEs. Monte Carlo calculations allow the elucidation of several "rules of thumb" for interpreting MFE data. Chapter 4 discusses recent optically-detected zero-field EPR measurements, adapting the gamma-COMPUTE algorithm from solid state NMR for their interpretation. Chapter 5 explores the role of RF polarisation in producing MFEs. The breakdown in weak fields of the familiar rotating frame approximation is analysed. Chapter 6 reviews current knowledge and landmark experiments in the area of animal magnetoreception. The origins of the sensitivity of European robins Erithacus rubecula to the Earth’s magnetic field are given particular attention. In Chapter 7, Schulten and Ritz’s hypothesis that avian magnetoreception is founded on a radical pair mechanism (RPM) reaction is appraised through calculations in model systems. Chapter 8 introduces quantitative methods of analysing anisotropic magnetic field effects using spherical harmonics. Chapter 9 considers recent observations that European robins may sometimes be disoriented by minuscule RF fields. These are shown to be consistent with magnetoreception via a radical pair with no (effective) magnetic nuclei in one of the radicals.