Search

Photosensitization by drugs is directly related with the excited species and the photoinduced processes arising from interaction with UVA light. In this context, the ability of gefitinib (GFT), a tyrosine kinase inhibitor (TKI) used for the treatment of a variety of cancers, to induce phototoxicity and photooxidation of proteins has recently been demonstrated. In principle, photodamage can be generated not only by a given drug but also by its photoactive metabolites that maintain the relevant chromophore. In the present work, a complete study of O-desmorpholinopropyl gefitinib (GFT-MB) has been performed by means of fluorescence and ultrafast transient absorption spectroscopies, in addition to molecular dynamics (MD) simulations. The photobehavior of the GFT-MB metabolite in solution is similar to that of GFT. However, when the drug or its metabolite are in a constrained environment, i.e. within a protein, their behavior and the photoinduced processes that arise from their interaction with UVA light are completely different. For GFT in complex with human serum albumin (HSA), locally excited (LE) singlet states are mainly formed; these species undergo photoinduced electron transfer with Tyr and Trp. By contrast, since GFT-MB is a phenol, excited state proton transfer (ESPT) to form phenolate-like excited species might become an alternative deactivation pathway. As a matter of fact, the protein-bound metabolite exhibits higher fluorescence yields and longer emission wavelengths and lifetimes than GFT@HSA. Ultrafast transient absorption measurements support direct ESPT deprotonation of LE states (rather than ICT), to form phenolate-like species. This is explained by MD simulations, which reveal a close interaction between the phenolic OH group of GFT-MB and Val116 within site 3 (subdomain IB) of HSA. The reported findings are relevant to understand the photosensitizing properties of TKIs and the role of biotransformation in this type of adverse side effects., Financial support from the Spanish Ministry of Science and Innovation (RYC-2015-17737, BEAGAL 18/00211, PID2020-115010RB-I00/AEI/10.13039/501100011033, FPU19/00048, PID2019-105512RB-I00/AEI/10.13039/501100011033), the Xunta de Galicia (ED431C 2021/29), the Centro singular de investigación de Galicia accreditation 2019–2022 (ED431G 2019/03), and the European Regional Development Fund (ERDF) is gratefully acknowledged. All authors are grateful to the Centro de Supercomputación de Galicia (CESGA) for use of the Finis Terrae computer.

[EN] Four new dyes that derive from borylated arylisoquinolines were prepared, containing a third aryl residue (naphthyl, 4-methoxynaphthyl, pyrenyl or anthryl) that is linked via an additional stereogenic axis. The triaryl cores were synthesized by Suzuki couplings and then transformed into boronic acid esters by employing an Ir(I)-catalyzed reaction. The chromophores show dual emission behavior, where the long-wavelength emission band can reach maxima close to 600 nm in polar solvents. The fluorescence quantum yields of the dyes are generally in the range of 0.2¿0.4, reaching in some cases values as high as 0.5¿0.6. Laserflash photolysis provided evidence for the existence of excited triplet states. The dyes form fluoroboronate complexes with fluoride anions, leading to the observation of the quenching of the long-wavelength emission band and ratiometric response by the build-up of a hypsochromically shifted emission signal., Funding by the Spanish Ministry of Economy, Industry, and Competitiveness (CTQ2014-54729-C2-1-P for U.P., CTQ2013-48164-C2-1-P and CTQ2013-48164-C2-2-P for A.R., Ramon y Cajal contracts RYC-2013-12585 for A.R. and RYC-2015-17737 for I.V.), the Spanish Ministry of Science, Innovation, and Universities (CTQ2017-89832-P for U.P., CTQ2016-78875-P for M.C.J., and CTQ2017-89416-R for I.V.), the European Research and Development Fund, and the Andalusian Government (2012/FQM-2140 for U.P., 2009/FQM-4537 and 2012/FQM-1078 for A.R) is gratefully acknowledged.

[EN] Gefitinib (GFT) is a tyrosine kinase inhibitor currently used for the treatment of metastatic non-small cell lung cancer. Although it has been suggested that GFT can be phototoxic, there are no systematic studies on this issue. Here, the photosensitizing potential of GFT has been assessed by means of NRU assays and protein photooxidation. In addition, a thorough photophysical study is presented based on ultrafast transient absorption spectroscopy, fluorescence and laser flash photolysis. Transient species generated after excitation of GFT have been characterized in solution and in biological environments (i.e. HSA and HaCaT cells) to gain insight into the mechanisms involved in photodamage. The photobehavior of GFT was strongly medium-dependent. Excitation of the drug resulted in the formation of locally excited (LE) singlet states ((1)GFT*), which were found to be the main emissive species in non-polar solvents and also within HSA and HaCaT cells. By contrast, in polar solvents, LE states rapidly evolved (similar to 1 ps) towards the formation of longer-lived intramolecular charge transfer (ICT) states. The triplet excited state of GFT ((3)GFT*) can be formed through intersystem crossing from (1)GFT* in non-polar solvents and from ICT states in the polar ones, or in the particular case of ethanol, by photosensitization using 2-methoxyacetophenone as an energy donor. In the HSA environment, (3)GFT* was hardly detected due to quenching of its LE (1)GFT* precursor by Trp through an electron transfer process. Accordingly, HSA photooxidation by GFT was demonstrated using the protein carbonylation method. In summary, a good correlation is established between the photophysical behavior and the photobiological properties of GFT, which provides a mechanistic basis for the observed phototoxicity., Financial support from the Spanish Government (RYC-2015-17737, CTQ2017-89416-R, BEAGAL 18/00211, FPU19/00048 and PID2020-115010RB-I00; the Carlos III Institute (ISCIII) of Health, grants: PI16/01877, and CPII16/00052) and Conselleria d'Educacio, Cultura i Esport (PROMETEO/2017/075) is gratefully acknowledged.

[EN] The epidermal growth factor receptors EGFR and HER2 are the main targets for tyrosine kinase inhibitors (TKIs). The quinazoline derivative lapatinib (LAP) is used since 2007 as dual TKI in the treatment of metastatic breast cancer and currently, it is used as an oral anticancer drug for the treatment of solid tumors such as breast and lung cancer. Although hepatotoxicity is its main side effect, it makes sense to investigate the ability of LAP to induce photosensitivity reactions bearing in mind that BRAF (serine/threonine-protein kinase B-Raf) inhibitors display a considerable phototoxic potential and that afloqualone, a quinazoline-marketed drug, causes photodermatosis. Metabolic bioactivation of LAP by CYP3A4 and CYP3A5 leads to chemically reactiveN-dealkylated (N-LAP) andO-dealkylated (O-LAP) derivatives. In this context, the aim of the present work is to explore whether LAP and itsN- andO-dealkylated metabolites can induce photosensitivity disorders by evaluating their photo(geno)toxicity through in vitro studies, including cell viability as well as photosensitized protein and DNA damage. As a matter of fact, our work has demonstrated that not only LAP, but also its metaboliteN-LAP have a clear photosensitizing potential. They are both phototoxic and photogenotoxic to cells, as revealed by the 3T3 NRU assay and the comet assay, respectively. By contrast, theO-LAP does not display relevant photobiological properties. Remarkably, the parent drug LAP shows the highest activity in membrane phototoxicity and protein oxidation, whereasN-LAP is associated with the highest photogenotoxicity, through oxidation of purine bases, as revealed by detection of 8-Oxo-dG., This study was funded by the Carlos III Institute (ISCIII) of Health (Grants: PI16/01877, CPII16/00052, ARADyAL RD16/0006/0030) co-funded by European Regional Development Fund, the Spanish Government (RYC-2015-17737, CTQ2017-89416-R,) and Generalitat Valenciana (Prometeo/2017/075). We would also like to thank IIS La Fe Microscopy Unit for technical assistance.

[EN] Benzophenone (BP) is present in a variety of bioactive molecules. This chromophore is able to photosensitize DNA damage, where one of the most relevant BP/ DNA interactions occurs with thymine (Thy). In view of the complex photoreactivity previously observed for dyads containing BP covalently linked to thymidine, the aim of this work is to investigate whether appropriate changes in the nature of the spacer could modulate the intramolecular BP/Thy photoreactivity, resulting in an enhanced selectivity. Accordingly, the photobehavior of a series of dyads derived from BP and Thy, separated by linear linkers of different length, has been investigated by steady-state photolysis, as well as femtosecond and nanosecond transient absorption spectroscopy. Irradiation of the dyads led to photoproducts arising from formal hydrogen abstraction or Paterno-Buchi (PB) photoreaction, with a chemoselectivity that was clearly dependent on the nature of the linking bridge; moreover, the PB process occurred with complete regio- and stereoselectivity. The overall photoreactivity increased with the length of the spacer and correlated well with the rate constants estimated from the BP triplet lifetimes. A reaction mechanism explaining these results is proposed, where the key features are the strain associated with the reactive conformations and the participation of triplet exciplexes., Financial support from the Spanish Government (RYC-2015-17737 and CTQ2017-89416-R) and from the Conselleria d'Educació Cultura i Esport (PROMETEO/2017/075 and GRISOLIAP/2017/005) is gratefully acknowledged. The authors would like to thank the use of RIAIDT-USC analytical facilities for the X-ray crystallography analysis.

[EN] The photoinduced cycloreversion of oxetanes has been thoroughly investigated in connection with the photorepair of the well-known DNA (6-4) photoproducts. In the present work, the direct photolysis of the two regioisomers arising from the irradiation of benzophenone (BP) and 1,3-dimethylthymine (DMT), namely the head-to-head (HH-1) and head-to-tail (HT-1) oxetane adducts, has been investigated by combining ultrafast spectroscopy and theoretical multiconfigurational quantum chemistry analysis. Both the experimental and computational results agree with the involvement of an excited triplet exciplex(3)[BPMIDLINE HORIZONTAL ELLIPSISDMT]* for the photoinduced oxetane cleavage to generate(3)BP* and DMT through an adiabatic photochemical reaction. The experimental signature of(3)[BPMIDLINE HORIZONTAL ELLIPSISDMT]* is the appearance of an absorption band atca.400 nm, detected by femtosecond transient absorption spectroscopy. Its formation is markedly regioselective, as it is more efficient and proceeds faster for HH-1 (similar to 2.8 ps) than for HT-1 (similar to 6.3 ps). This is in line with the theoretical analysis, which predicts an energy barrier to reach the triplet exciplex for HT-1, in contrast with a less hindered profile for HH-1. Finally, the more favorable adiabatic cycloreversion of HH-1 compared to that of HT-1 is explained by its lower probability to reach the intersystem crossing with the ground state, which would induce a radiationless deactivation process leading either to a starting adduct or to a dissociated BP and DMT., Financial support from the Spanish Government (RYC-2015-17737, CTQ2017-89416-R, RYC-2015-19234, CTQ2017-87054-C2-2-P, and MDM-2015-0538), from the Conselleria d'Educacio, Investigacio, Cultura i Esport (PROMETEO/2017/075 and GRISOLiAP/2017/005) and from the Universitat de Valencia (postdoctoral grant within the "Atraccio de Talent 2019" Program for A. G.) is gratefully acknowledged. This work was also supported by a 2019 Leonardo Grant for Researchers and Cultural Creators, BBVA Foundation. The Foundation takes no responsibility for the opinions, statements, and contents of this project, which are entirely the responsibility of its authors.

[EN] Lapatinib (LAP) is an anticancer drug, which is metabolized to theN- and O-dealkylated products (N-LAP andO-LAP, respectively). In view of the photosensitizing potential of related drugs, a complete experimental and theoretical study has been performed on LAP,N-LAP andO-LAP, both in solution and upon complexation with human serum albumin (HSA). In organic solvents, coplanar locally excited (LE) emissive states are generated; they rapidly evolve towards twisted intramolecular charge-transfer (ICT) states. By contrast, within HSA only LE states are detected. Accordingly, femtosecond transient absorption reveals a very fast switching (ca. 2 ps) from LE (lambda(max)=550 nm) to ICT states (lambda(max)=480 nm) in solution, whereas within HSA the LE species become stabilized and live much longer (up to the ns scale). Interestingly, molecular dynamics simulation studies confirm that the coplanar orientation is preferred for LAP (or to a lesser extentN-LAP) within HSA, explaining the experimental results., Financial support from the Spanish Government (RYC-2015-17737, CTQ2017-89416-R, ISCIII grants RD16/0006/0004, PI16/01877 and CPII16/00052, SAF2016-75638-R, RYC-2015-19234, CTQ2017-87054-C2-2-P, and MDM-2015-0538), Conselleria d'Educacio Cultura i Esport (PROMETEO/2017/075), the Xunta de Galicia [ED431B 2018/04 and Centro singular de investigacion de Galicia accreditation 2019-2022 (ED431G 2019/03)] and the European Regional Development Fund is gratefully acknowledged. We thank the Centro de Supercomputacion de Galicia (CESGA) for use of the Finis Terrae computer.

International audience; The electronic excited states populated upon absorption of UV photons by DNA are extensively studied in relation to the UV-induced damage to the genetic code. Here, we report a new unexpected relaxation pathway in adenine–thymine double-stranded structures (AT)n. Fluorescence measurements on (AT)n hairpins (six and ten base pairs) and duplexes (20 and 2000 base pairs) reveal the existence of an emission band peaking at approximately 320 nm and decaying on the nanosecond time scale. Time-dependent (TD)-DFT calculations, performed for two base pairs and exploring various relaxation pathways, allow the assignment of this emission band to excited states resulting from mixing between Frenkel excitons and adenine-to-thymine chargetransfer states. Emission from such high-energy long-lived mixed (HELM) states is in agreement with their fluorescence anisotropy (0.03), which is lower than that expected for p–p* states (0.1). An increase in the size of the system quenches p–p* fluorescence while enhancing HELM fluorescence. The latter process varies linearly with the hypochromism of the absorption spectra, both depending on the coupling between p–p* and charge-transfer states. Subsequently, we identify the common features between the HELM states of (AT)n structures with those reported previously for alternating (GC)n : high emission energy, low fluorescence anisotropy, nanosecond lifetimes, and sensitivity to conformational disorder. These features are also detected for calf thymus DNA in which HELM states could evolve toward reactive p–p* states, giving rise to delayed fluorescence.

[EN] Through-bond triplet exciplex formation in donor-acceptor systems linked through a rigid bile acid scaffold has been demonstrated on the basis of kinetic evidence upon population of the triplet acceptors (naphthalene, or biphenyl) by through-bond triplet-triplet energy transfer from benzophenone., Financial support from the Spanish Government (Grants SEV-2012-0267, CTQ2012-38754-C03-03, CTQ2013-47872-C2-1-P and JCI-2011-09926), EU (PCIG12GA-2012-334257), Generalitat Valenciana (Prometeo Program), and Technical University of Valencia (VLC/Campus, ASIC-UPV for computational facilities and Predoctoral FPI fellowship for P. Miro) is gratefully acknowledged.

International audience; The study concerns the relaxation of electronic excited states of the DNA nucleoside deoxycytidine (dCyd) and its methylatedanalogue 5-methyldeoxycytidine (5mdCyd), known to be involved in the formation of UV-induced lesions of the genetic code. Due to the existence of four closely lying and potentially coupled excited states, the deactivation pathways in these systems are particularly complex and have not been assessed so far. Here, we provide a complete mechanistic picture of theexcited state relaxation of dCyd/5mdCyd in three solvents water, acetonitrile, and tetrahydrofuran by combining femtosecond fluorescence experiments, addressing the effect of solvent proticity on the relaxation dynamics of dCyd and 5mdCyd for the first time, and two complementary quantum mechanical approaches (CASPT2/MM and PCM/TD-CAM-B3LYP). The lowest energy ππ* state is responsible for the sub-picosecond lifetime observed for dCyd in all the solvents. In addition, computed excited state absorption and transient IR spectra allow one, for the first time, to assign the tens of picoseconds time constant, reported previously, to a dark state (nOπ*) involving the carbonyl lone pair. A second low-lying dark state, involving the nitrogen lone pair (nNπ*), does significantly participate in the excited state dynamics. The 267 nm excitation of dCyd leads to a non-negligible population of the second bright ππ* state, which affects the dynamics, acting mainly as a “doorway” state for the nOπ* state. The solvent plays a key role governing the interplay between the different excited states; unexpectedly, water favors population of the dark states. In the case of 5mdCyd, an energy barrier present on the main nonradiative decay route explains the 6-fold lengthening of the excited state lifetime compared to that of dCyd, observed for all the examined solvents. Moreover, C5-methylation destabilizes both nOπ* and nNπ* dark states, thus preventing them from being populated.

[EN] The properties of singlet and triplet excited states are strongly medium-dependent. Hence, these species constitute valuable tools as reporters to probe compartmentalised microenvironments, including drug@protein supramolecular systems. In the present review, the attention is focused on the photophysical properties of the probe drugs (rather than those of the protein chromophores) using transport proteins (serum albumins and 1-acid glycoproteins) as hosts. Specifically, fluorescence measurements allow investigating the structural and dynamic properties of biomolecules or their complexes. Thus, the emission quantum yields and the decay kinetics of the drug singlet excited states provide key information to determine important parameters such as the stoichiometry of the complex, the binding constant, the relative degrees of occupancy of the different compartments, etc. Application of the FRET concept allows determining donor-acceptor interchromophoric distances. In addition, anisotropy measurements can be related to the orientation of the drug within the binding sites, where the degrees of freedom for conformational relaxation are restricted. Transient absorption spectroscopy is also a potentially powerful tool to investigate the binding of drugs to proteins, where formation of encapsulated triplet excited states is favoured over other possible processes leading to ionic species (i. e. radical ions), and their photophysical properties are markedly sensitive to the microenvironment experienced within the protein binding sites. Even under aerobic conditions, the triplet lifetimes of protein-complexed drugs are remarkably long, which provides a broad dynamic range for identification of distinct triplet populations or for chiral discrimination. Specific applications of the laser flash photolysis technique include the determination of drug distribution among the bulk solution and the protein binding sites, competition of two types of proteins to bind a 3 drug, occurrence of drug-drug interactions within protein binding sites, enzymatic-like activity of the protein or determination of enantiomeric compositions. The use of proteins as supramolecular hosts modifies the photoreactivity of encapsulated substrates by providing protection against oxygen or other external reagents, by imposing conformational restrictions in the binding pockets, or by influencing the stereochemical outcome. In this review, a selected group of examples is presented including decarboxylation, dehalogenation, nucleophilic addition, dimerisation, oxidation, Norrish type II reaction, photo-Fries rearrangement and 6 electrocyclisation, Financial support from the Spanish Government (CTQ2010-14882, JCI-2011-09926, RyC-2007-00476), from the EU (PCIG12-GA-2012-334257), from the Universitat Politènica de València (SP20120757) and from the Consellería de Educació, Cultura i Esport (PROMETEOII/2013/005, GV/2013/051) is gratefully acknowledged.

The interaction dynamics between the drug flurbiprofen (FBP) and human serum albumin (HSA) has been investigated by time-resolved fluorescence spectroscopy, combining femtosecond fluorescence upconversion and picosecond time-correlated single photon counting. In order to obtain additional information on the drug/ protein interaction, several covalently linked model dyads, composed of FBP and tryptophan or tyrosine, were also studied. For all systems, the main feature was a remarkable dynamic FBP fluorescence quenching, more prominent in the dyads than in the protein complex. All systems also displayed a clear stereoselectivity depending on the (S)- or (R)-form of FBP, that was strongly influenced by the conformational arrangement of the investigated chromophores.

[EN] The electrochemical and photophysical properties of a family of conjugated ligands and their iridium(III) cyclometallated complexes are described. They consist of a series of monocationic Ir-III bis-2-phenylpyridine complexes with p-phenylethynyl-1,10-phenanthroline ligands of different length. The structure of these ligands includes terminal acetylthiol or pyridine groups, which can provide good electrical contacts between metal electrodes. Cyclic voltammetry, absorption and emission spectroscopy, laser flash photolysis and density functional theory calculations reveal that the high conjugation of the diimine ligand affords small energy gaps between the frontier orbitals. Nevertheless, the nature of the terminal substituents and the extent of the conjugation in the diimine ligand have little influence on the photophysical features at room temperature. The spectroscopic data and theoretical calculations agree that the charge-transfer nature of the emitting excited state is maintained along the series at room temperature, whereas in rigid matrices ligand-centred states also contribute to the low-temperature emission. The good conducting features of the diimine ligands, the small dependence of the HOMO-LUMO (HOMO = highest occupied molecular orbital, LUMO = lowest unoccupied molecular orbital) gaps of these complexes on the ligands and the charge-transfer nature of the emitting excited state make these complexes promising test beds for the study of photo-conducting phenomena in molecular junctions., The authors would like to especially acknowledge Prof. Hans U. Gudel for his valuable inspiration and friendship. The present work has been funded by the European Union (EU), (ERC Advanced Grant SPINMOL), the Spanish Ministerio de Economia y Competitividad (MINECO) (projects MAT2014-56243, CTQ2015-71154, and Unidad de Excelencia Maria de Maeztu MDM-2015-0552), and the Generalitat Valenciana (Prometeo/2012/053, PrometeoII/2013/006 and ISIC-Nano). J. P., I. V. and S. T. thank the Ministerio de Ciencia e Innovacion (MICINN) for their predoctoral fellowships and JdlC contracts. J. A., I. V., and S. T. also thank the European Union (EU) for their FP7-PEOPLE-2012-IEF-329513, PCIG12GA-2012-334257 and MSCA-IF-657465, and FP7-PEOPLE-2012-CIG-321739 grants, respectively.

International audience; The effect of metal ions on the excited states of guanine nanostructures, short d(TG4T)4 quadruplexes and long G4-wires, are studied by fluorescence spectroscopy. The steadystate emission spectra show that both systems exhibit a strong cation effect. Fluorescence decays and fluorescence anisotropy decays, recorded from the femtosecond to the nanosecond timescale, reveal the following picture. In the presence of K+, emission arises mainly from delocalized ππ* states (excitons), whose decay spans several decades of times. In contrast, the fluorescence in the presence of Na+ is dominated by emission from charge transfer excited states decaying essentially on the subnanosecond time-scale. Such a difference is not due to the initially populated (Franck−Condon) states. The interproton distances derived from two-dimensional NMR measurements on the ground state of d(TG4T)4 show that the geometrical arrangement of guanines, governing the electronic coupling, is the same for both cations, in line with the UV absorption spectra. The observed cation effect is correlated with the excited state relaxation: the increased mobility of Na+ ions within the quadruplex favors trapping of ππ* excitons by charge transfer excited states, whereas such a process is hindered for the larger K+ ions. This is rationalized by quantum calculations on two stacked guanine tetrads.

In the present work, a systematic fluorescence study on supramolecular systems using two serum albumins (HSA or BSA) as hosts and the nonsteroidal antiinflammatory drugs carprofen (CPF) or naproxen (NPX) as guests has been undertaken. In parallel, model dyads containing Tyr or Trp covalently linked to CPF or NPX have also been investigated. In HSA/(S)-CPF and BSA/(S)-CPF ( λ exc = 266 nm), at 1:1 M ratio, an important degree (more than 40%) of singlet–singlet energy transfer (SSET) was observed to take place. The distance ( r ) calculated for energy transfer from the SAs to (S)-CPF through a FRET mechanism was found to be ca. 21 A. In the case of HSA/(S)-NPX and BSA/(S)-NPX, energy transfer occurred to a lower extent (ca. 7%), and r was determined as ca. 24 A. In order to investigate the possible excited state interactions between bound ligands and the relevant amino acids present in the protein binding sites, four pairs of model dyads were designed and synthesised, namely ( S , S )-TyrCPF, ( S , R )-TyrCPF, ( S , S )-TrpCPF, ( S , R )-TrpCPF, ( S , S )-TyrNPX, ( S , R )-TyrNPX, ( S , S )-TrpNPX and ( S , R )-TrpNPX. A complete SSET was observed from Tyr or Trp to CPF, since no contribution from the amino acids was present in the emission of the dyads. Likewise, a very efficient Tyr or Trp to NPX energy transfer was observed. Remarkably, in ( S , S )-TrpNPX and ( S , R )-TrpNPX a configuration-dependent reduction in the emission intensity was observed, revealing a strong and stereoselective intramolecular quenching. This effect can be attributed to exciplex formation and is dynamic in nature, as the fluorescence lifetimes were much shorter in ( S , R )- and ( S , S )-TrpNPX (1.5 and 3.1 ns, respectively) than in (S)-NPX (11 ns).

[EN] The photoreactivity of diastereomeric dyads containing (S)- or (R)-flurbiprofen (FBP) and (S)-Tyr, either directly linked (1) or separated by a cyclic spacer (3) has been investigated. The main feature is a remarkable intramolecular quenching of FBP fluorescence, especially in 1. The process is clearly configuration dependent, being more efficient for the (R,S)- diastereomer in 1 and for the (S,S)-analogue in 3. Noteworthy, exciplex emission is detected in the 380-500 nm region in the case of 3. Fluorescence decay kinetics from the femtosecond to the nanosecond time-domains provides evidence for the dynamic nature of the quenching. In agreement with the steady-state and time-resolved observations, molecular modelling points to a more favourable geometric arrangement of the two interacting chromophores in 1 than in 3., Financial support from the Spanish Government (CTQ2013-47872-C2-1-P), EU (PCIG12GA-2012-334257, LASERLAB-EUROPE grant agreement no. 284464, EU FP7, and MSCA- 657465) and Generalitat Valenciana (PROMETEOII/2013/005) is gratefully acknowledged.

[EN] The photoreactivity of fenofibric acid (FA) in the presence of human and bovine serum albumins (HSA and BSA, respectively) has been investigated by steady-state irradiation, fluorescence, and laser flash photolysis (LFP). Spectroscopic measurements allowed for the determination of a 1:1 stoichiometry for the FA/SA complexes and pointed to a moderate binding of FA to the proteins; by contrast, the FA photoproducts were complexed more efficiently with SAs. Covalent photobinding to the protein, which is directly related to the photoallergic properties of the drug, was detected after long irradiation times and was found to be significantly higher in the case of BSA. Intermolecular FA-amino acid and FA-albumin irradiations resulted in the formation of photoproducts arising from coupling between both moieties, as indicated by mass spectrometric analysis. Mechanistic studies using model drug-amino acid linked systems indicated that the key photochemical step involved in photoallergy is formal hydrogen atom transfer from an amino acid residue to the excited benzophenone chromophore of FA or (more likely) its photoproducts. This results in the formation of caged radical pairs followed by C-C coupling to give covalent photoaducts., Financial support from the Spanish Government (CTQ2013-47872-C2-1-P, JCI-2011-09926, and Miguel Servet CP11/00154), EU (PCIG12GA-2012-334257) and Generalitat Valenciana (PROMETEOII/2013/005) is gratefully acknowledged.

[EN] Three stereoisomeric salts of naproxen (NPX) with arginine (Arg), namely (S)-NPX/(S)-Arg, (R)-NPX/(S)-Arg and (S)NPX/(R)-Arg, have been prepared, and their fluorescence spectra recorded in solution and in the solid state. While the emission properties in solution did not show significant differences with lambda(max) = 355 nm, tau(F) (MeOH) ca. 11.5 ns and tau(F) (H2O) ca. 9 ns (as NPX/Na), the (R)-NPX/(S)-Arg and (S)-NPX/(R)-Arg solid salts displayed red-shifted fluorescence spectra With maxima at 375 mn and tau(F) = 1.1 ns. By contrast, the behaviour of solid (S)-NPX/(S)-Arg was similar to that of NPX/Na With; ax = 355 nm and tau(F) ca. 5.5 ns. These results are explained based on the X-ray crystal structures and attributed to formation of NPX excimers emitting at longer wavelengths. Accordingly, such excimer emission was also observed in the fluorescence spectrum of a model NPX dyad in solution., The UPV (PI 2003-0522 and predoctoral fellowship to I.V.), the MYCT (Grant CTQ2004-03811) the Generalitat Valenciana (Grupos03/082 and GV04B-468) are gratefully acknowledged for financial support. We also thank our colleague Dr. M. L. Marin for providing a gift of (2-methoxynaphthalen-6-yl)acetic acid and Dr. A. Llamas (Unidade de Raios X de la Universidade de Santiago de Compostela) for the X-ray measurements.

The photophysical properties of S-flurbiprofen [S-2-fluoro-alpha-methyl-4-biphenylacetic acid], a nonsteroidal anti-inflammatory drug, have been examined using steady-state and time-resolved spectroscopic techniques. The energy of its first singlet excited state is 99 kcal mol(-1). The fluorescence quantum yields and lifetimes (at 300 nm) have been determined in acetonitrile, methanol, hexane and PBS; they are in the range 0.15phi(F)0.33 and 0.7tau(F)2.0 ns. The intersystem crossing quantum yields are between 0.45 and 0.71; the lambda(max) of the T-T absorption is 360 nm, and the triplets live from 15 to 106 micros. Steady state photolysis in aqueous medium leads to S-2-hydroxy-alpha-methyl-4-biphenylacetic acid via photonucleophilic aromatic substitution, in addition to the photodecarboxylation products observed in organic solvents.

The properties of singlet and triplet excited states are strongly medium-dependent. Hence, these species constitute valuable tools as reporters to probe compartmentalised microenvironments, including drug@protein supramolecular systems. In the present review, the attention is focused on the photophysical properties of the probe drugs (rather than those of the protein chromophores) using transport proteins (serum albumins and α1-acid glycoproteins) as hosts. Specifically, fluorescence measurements allow investigation of the structural and dynamic properties of biomolecules or their complexes. Thus, the emission quantum yields and the decay kinetics of the drug singlet excited states provide key information to determine important parameters such as the stoichiometry of the complex, the binding constant, the relative degrees of occupancy of the different compartments, etc. Application of the FRET concept allows determination of donor–acceptor interchromophoric distances. In addition, anisotropy measurements can be related to the orientation of the drug within the binding sites, where the degrees of freedom for conformational relaxation are restricted. Transient absorption spectroscopy is also a potentially powerful tool to investigate the binding of drugs to proteins, where formation of encapsulated triplet excited states is favoured over other possible processes leading to ionic species (i.e. radical ions), and their photophysical properties are markedly sensitive to the microenvironment experienced within the protein binding sites. Even under aerobic conditions, the triplet lifetimes of protein-complexed drugs are remarkably long, which provides a broad dynamic range for identification of distinct triplet populations or for chiral discrimination. Specific applications of the laser flash photolysis technique include the determination of drug distribution among the bulk solution and the protein binding sites, competition of two types of proteins to bind a drug, occurrence of drug–drug interactions within protein binding sites, enzymatic-like activity of the protein or determination of enantiomeric compositions. The use of proteins as supramolecular hosts modifies the photoreactivity of encapsulated substrates by providing protection against oxygen or other external reagents, by imposing conformational restrictions in the binding pockets, or by influencing the stereochemical outcome. In this review, a selected group of examples is presented including decarboxylation, dehalogenation, nucleophilic addition, dimerisation, oxidation, Norrish type II reaction, photo-Fries rearrangement and 6π electrocyclisation.

International audience; We report here on a recent time-resolved fluorescence study [1] of the interaction between flurbiprofen (FBP), a chiral non-steroidal anti-inflammatory drug, and human serum albumin (HSA), the main transport protein in the human body. We compare the results obtained for the drug-protein complex with those of various covalently linked flurbiprofentryptophan dyads having well-defined geometries. In all cases stereoselective dynamic fluorescence quenching is observed, varying greatly from one system to another. In addition, the fluorescence anisotropy decays also display a clear stereoselectivity. For the drug-protein complexes, this can be interpreted in terms of the protein microenvironment playing a significant role in the conformational relaxation of FBP, which is more restricted in the case of the (R)- enantiomer.

The photooxygenation of model compounds containing the two enantiomers of naproxen (NPX) covalently linked to histidine (His), tryptophan (Trp) and tyrosine (Tyr) has been investigated by steady state irradiation, fluorescence spectroscopy and laser flash photolysis. The NPX–His systems presented the highest oxygen-mediated photoreactivity. Their fluorescence spectra matched that of isolated NPX and showed a clear quenching by oxygen, leading to a diminished production of the NPX triplet excited state ( 3 NPX*–His). Analysis of the NPX–His and NPX–Trp photolysates by UPLC-MS–MS revealed in both cases the formation of two photoproducts, arising from the reaction of singlet oxygen (1 O2) with the amino acid moiety. The most remarkable feature of NPX–Trp systems was a fast and stereoselective intramolecular fluorescence quenching, which prevented the efficient formation of 3 NPX*–Trp, thus explaining their lower reactivity towards photooxygenation. Finally, the NPX–Tyr systems were nearly unreactive and exhibited photophysical properties essentially coincident with those of the parent NPX. Overall, these results point to a type II photooxygenation mechanism, triggered by generation of 1 O2 from the 3 NPX* chromophore, Financial support from the Spanish Government (CTQ2010-14882, JCI-2011-09926, Miguel Servet CP11/00154), from the EU (PCIG12-GA-2012-334257), from the Universitat Politecnica de Valencia (SP20120757) and from the Conselleria de Educacio, cultura i Esport (PROMETEOII/2013/005, GV/2013/051) is gratefully acknowledged.

The excited states of deoxyadenosine (dA) and 9-methyladenine (9Me-Ade) were studied in water and acetonitrile by a combination of steady-state and time-resolved spectroscopy and quantum chemical calculations. Femtosecond fluorescence upconversion experiments show that the decays of dA and 9Me-Ade after excitation at 267 nm are very similar, confirming that 9Me-Ade is a valid model for the calculations. The fluorescence decays can be described by an ultrafast component (, R.I. thanks MIUR (FIRB 2008 Futuro in Ricerca and PRIN 2010-2011) for financial support. The French Agency for Research (ANR-10-BLAN-0809-01, "DNAExciton") is acknowledged for financial support. Financial support from the Spanish Government (JCI-2011-09926 and Salvador Madariaga Program (grant to M.C.J.)) is gratefully acknowledged.

[EN] Guanine-cytosine hairpins, containing a hexaethylene glycol bridge, are studied by steady-state fluorescence spectroscopy and time-correlated single photon counting; their properties are compared to those of duplexes with the same sequence. It is shown that, both in hairpins and in duplexes, base pairing induces quenching of the pi pi(star) fluorescence, the quantum yield decreasing by at least two orders of magnitude. When the size of the systems increases from two to ten base pairs, a fluorescent component decaying on the nanosecond time-scale appears at energy higher than that stemming from the bright states of non-interacting mono-nucleotides (ca. 330 nm). For ten base pairs, this new fluorescence forms a well-defined band peaking at 305 nm. Its intensity is about 20% higher for the hairpin compared to the duplex. Its position (red-shifted by 1600 cm(-1)) and width (broader by 1800 cm(-1) FWHM) differ from those observed for large duplexes containing 1000 base pairs, suggesting the involvement of electronic coupling. Fluorescence anisotropy reveals that the excited states responsible for high energy emission are not populated directly upon photon absorption but are reached during a relaxation process. They are assigned to charge transfer states. According to the emerging picture, the amplitude of conformational motions determines whether instantaneous deactivation to the ground state or emission from charge transfer states will take place, while pi pi(star) fluorescence is associated to imperfect base-pairing., The French Agency for Research (ANR-10-BLAN-0809-01), the Conselleria de Educacion-Generalitat Valenciana (VALi+D program to I.V., N° 2010033) and the US Department of Energy, Office of Basic Energy Sciences (grant no. DE-FG02-96ER14604 to F.D.L.) are acknowledged for financial support.

[EN] DNA double-stranded oligomers are studied by steady-state and time-resolved fluorescence spectroscopy from the femtosecond to the nanosecond time-scale, following excitation at 267 nm. It is shown that emission arises from three types of excited states. (i) Bright pi pi* states emitting around 330 nm and decaying on the sub-picosecond time-scale with an average lifetime of ca. 0.4 ps and a quantum yield lower than 4 x 10(-6). (ii) Excimers/exciplexes emitting around 430 nm and decaying on the sub-nanosecond time-scale. (iii) Excited states emitting mainly at short wavelengths (lambda < 330 nm) and decaying on the nanosecond time-scale, possibly correlated to GC pairs. The properties of the examined duplexes, exhibiting significant disorder with respect to the nearest neighbour base sequence, are radically different than those of the much longer and disordered calf thymus DNA. Such behaviour suggests that long range and/or sequence effects play a key role in the fate of excitation energy., The French Agency for Research (ANR-10-BLAN-0809-01) and the Conselleria de Educacion-Generalitat Valenciana (VALi+D program to I.V., no. 2010033) are acknowledged for financial support.

Excited-state interactions between (S)- or (R) fluorescence (S)- or (R)-FBP) and thymidine (dThd) covalently linked in dyads 1 or 2 have been investigated. In both dyads, the only emitting species is (FBP)-F-1*, but with a lower fluorescence quantum yield (phi(F)) and a shorter fluorescene lifetime (tau(F)) than when free in solution. These an results indicate that dynamic quenching occurs either by electron transfer or via exciplex formation, with FBP as the charge-donating species. In acetonitrile, both mechanisms are favored, while in dioxane exciplex formation is predominating. Isomer 1 displays lower values of OF and rF than its analogue 2, indicating that the relative spatial arrangement of the chromophores plays a significant role. The triplet quantum yields (0-F) of 1 and 2 are significantly higher than the expectations based solely on 'FBP* dThd intersystem crossing quantum yields (Oisc), with OT (1) > OT (2). This can be explained in terms of intramolecular charge recombination at the radical ion pairs and/or the exciplexes, which would be again dependent on geometrical factors. The triplet lifetimes (IT) of 3FBP* dThd and free 3FBP* are similar, indicating the lack of excited-state interactions at this stage. The FBP dThd dyads could, in principle, constitute appropriate model systems for the elucidation of the excited-state interactions in noncovalent DNA ligand complexes., Financial support from the Spanish Government (Grants CTQ2010-14882 and CTQ2009-13699), Salvador Madariaga Program (Grant to M.C.J.), Juan de la Cierva Program (contract to I.V.), Generalitat Valenciana (Prometeo 2008/090), and the Universitat Politecnica de Valencia (PAID 05-11, ref 2766) is gratefully acknowledged.

[EN] Transfer of the electronic excitation energy in calf thymus DNA is studied by time-resolved fluorescence spectroscopy. The fluorescence anisotropy, after an initial decay starting on the femtosecond time scale, dwindles down to ca. 0.1. The in-plane depolarized fluorescence decays are described by a stretched exponential law. Our observations are consistent with one-dimensional transfer mediated by charge-transfer excited states., The French Agency for Research (ANR-10-BLAN-0809-01), the Conselleria de Educacion-Generalitat Valenciana (VALi+D program to I.V., No. 2010033), and ONR/MURI (Y.B.) are acknowledged for financial support.

International audience; The excited state properties of two amino substituted uracils, 5-aminouracil (5AU) and 5-dimethylaminouracil (5DMAU) have been studied by steady-state and time-resolved fluorescence spectroscopy. In addition, the electronic transitions corresponding to photon absorption and emission were calculated using the TD-DFT/PCM method (with PBEO and CAM-B3LYP functionals), explicitly including water molecules of the first solvent shell. The fluorescence decays, recorded by fluorescence upconversion, reveal that the emitting states of both molecules are substantially longer-lived than that of uracil. However, while the 5AU fluorescence decays on a picosecond timescale, the 5DMAU fluorescence contains a weak, long-lived component, continuing beyond 100 ps. Calculations show that the excited state deactivation of 5AU is governed by the existence of a minimum on the pi pi* excited state potential energy surface. For 5DMAU, on the other hand, a very long-lived but weakly fluorescent intramolecular charge transfer (ICT) state, involving a significant charge transfer between the dimethylamino group and the pyrimidine ring, is found. This stable ICT state is characterized by a rotation of the dimethylamino group towards a quasi-perpendicular geometry

[EN] The study addresses interconnected issues related to two major types of cycloadditions between adjacent thymines in DNA leading to cyclobutane dimers (T <> Ts) and (6-4) adducts. Experimental results are obtained for the single strand (dT)(20) by steady-state and time-resolved optical spectroscopy, as well as by HPLC coupled to mass spectrometry. Calculations are carried out for the dinucleoside monophosphate in water using the TD-M052X method and including the polarizable continuum model; the reliability of TD-M052X is checked against CASPT2 calculations regarding the behavior of two stacked thymines in the gas phase. It is shown that irradiation at the main absorption band leads to cyclobutane dimers (T <> Ts) and (6-4) adducts via different electronic excited states. T <> Ts are formed via (1)pi pi* excitons; [2+2] dimerization proceeds along a barrierless path, in line with the constant quantum yield (0.05) with the irradiation wavelength, the contribution of the (3)pi pi* state to this reaction being less than 10%. The formation of oxetane, the reaction intermediate leading to (6-4) adducts, occurs via charge transfer excited states involving two stacked thymines, whose fingerprint is detected in the fluorescence spectra; it involves an energy barrier explaining the important decrease in the quantum yield of (6-4) adducts with the irradiation wavelength., The French Agency for Research (ANR-10-BLAN-0809-01) and the Conselleria de Educacion-Generalitat Valenciana (VALi+D program to I.V., no. 2010033). R.I. thanks MIUR (PRIN 2008 and FIRB Futuro in Ricerca 2008) for financial support.

The two enantiomers of 2-(2-hydroxybiphenyl-4-yl)propanoic acid ((S)- and (R)-BPOH) have been selected as probes for human serum albumin (HSA). Photophysical characterization in the absence of protein led to emission maxima, singlet energies, quantum yields, and fluorescence lifetime values of 332 nm, 91 kcal mol(-1), and 0.28 and 1.8 ns for BPOH or 414 nm, 79 kcal mol(-1), and 0.31 and 3.3 ns for the corresponding phenolate BPO-; the pK(a)* was found to be 1.17. In the presence of HSA, a light-absorbing ground-state complex (S)-BPOH@HSA was detected (maximum at ca. 300 nm) whose intensity increased with increasing protein concentration. The fluorescence spectra of (S)-BPOH in PBS, after addition of HSA, revealed a progressive diminution of the phenolate band, indicating that excited-state deprotonation is disfavored within the hydrophobic protein cavities. A similar trend was observed for (R)-BPOH, but the extent of deprotonation was significantly lower for this enantiomer. Addition of increasing amounts of the site II displacement probe (S)-ibuprofen ((S)-IBP) to BPOH@HSA led to a significant decrease of the absorption maximum at ca. 300 nm and to a recovery of the phenolate emission band at ca. 410 nm, which were again configuration dependent. The transient absorption spectrum of (S)-BPOH consisted on a broad band centered at 380 nm, attributed to the first triplet excited state. A dramatic enhancement of the triplet lifetimes within HSA was observed (19.0 mu s within protein versus 1.3 mu s in bulk PBS), although no stereodifferentiation was noticed in this case., Financial support from the Spanish Government (CTQ2010-14882, CTQ2009-13699, BES-2008-003314, JCI-2011-09926), from the Generalitat Valenciana (Prometeo 2008/090) and from the Universitat Politecnica de Valencia (PAID 05-11, 2766) is gratefully acknowledged.

[EN] The photochemical properties of the DNA duplex (dA)(20) center dot (dT)(20) are compared with those of the parent single strands. It is shown that base pairing increases the probability of absorbing UVA photons, probably due to the formation of charge-transfer states. UVA excitation induces fluorescence peaking at similar to 420 nm and decaying on the nanosecond time scale. The fluorescence quantum yield, the fluorescence lifetime, and the quantum yield for cyclobutane dimer formation increase upon base pairing. Such behavior contrasts with that of the UVC-induced processes., We thank Mrs. Si. Karpati and M. Perron for their help, Dr. R. lmprota for helpful discussions, and the French Agency for Research (ANR PCV07_ 194999) for financial support. I.V. acknowledges the Conselleria de Educacion-Generalitat Valenciana (VALi+D program, No. 20100331).

International audience; Dark DNA light: Dark excited states of alternating GC double strands emit fluorescence at 4000 cm-1 higher energy (see spectrum) and with four orders of magnitude longer lifetime compared to the bright * states. Such high-energy long-lived excited states are expected to play a key role in the DNA photoreactivity associated with the appearance of carcinogenic mutations.

International audience; The fluorescence of calf thymus DNA is studied by steady-state and time-resolved spectroscopy combining fluorescence upconversion and time-correlated single photon counting. The fluorescence spectrum is very similar to that of a stoichiometric mixture of monomeric chromophores, arising from bright ππ* states, and contrasts with the existing picture of exciplex emission in natural DNA. Yet, the DNA fluorescence decays span over five decades of time, with 98% of the photons being emitted at times longer than 10 ps. These findings, in association with recent studies on model duplexes, are explained by the involvement of dark states, possibly related to charge separation, serving as a reservoir for the repopulation of the bright ππ* states.

International audience; Long nanowires formed by ca. 800 guanine tetrads (G4-wires) are studied in phosphate buffer containing sodium cations. Their room temperature optical properties are compared to those of the monomeric chromophore 2-deoxyguanine monophosphate (dGMP). When going from dGMP to G4-wires, both the absorption and the fluorescence spectra change. Moreover, the fluorescence quantum yield increases by a factor of 7.3 whereas the average fluorescence lifetime increases by more than 2 orders of magnitude, indicating emission associated with weakly allowed transitions. The behavior of G4-wires is interpreted in the light of a theoretical study performed in the frame of the exciton theory combining data from molecular dynamics and quantum chemistry. These calculations, carried out for a quadruplex composed of three tetrads, reveal the existence of various exciton states having different energies and oscillator strengths. The degree of delocalization of the quadruplex Franck−Condon excited states is larger than those found for longer duplexes following the same methodology. The slower excited-state relaxation in G4-wires compared to dGMP is explained by emission from exciton states, possibly limited on individual tetrads, whose coherence is reserved by the reduced mobility of guanines due to multiple Hoogsteen hydrogen bonds.

International audience; The fluorescence spectrum of the homopolymeric double helix poly(dG)·poly(dC) is dominated by emission decaying on the nanosecond time-scale, as previously reported for the alternating homologue poly(dGdC)·poly(dGdC). Thus, energy trapping over long periods of time is a common feature of GC duplexes which contrast with AT duplexes. The impact of such behaviour on DNA photodamage needs to be evaluated.

International audience; Recent fluorescence studies of DNA duplexes with a simple repetitive base sequence have revealed important collective effects which are very sensitive to conformational disorder. In contrast to the monomeric chromophores, whose fluorescence lifetimes are shorter than 1 ps, the fluorescence decays of duplexes span several decades of time. The occurrence of excitation energy transfer, evidenced by the decay of the fluorescence anisotropy on the femtosecond time scale, is a common feature of all of the examined systems. It is explained by the existence of Franck-Condon exciton states, in line with the Vabsorption spectra. Understanding the nature of the long-lived excited states, whose emission dominates the steady-state fluorescence spectrum of natural DNA, will be one of the challenges for the years to come.

[EN] Laser flash photolysis (LFP) has been used to determine the degree of binding of (S)- or (R)-flurbiprofen methyl ester (FBPMe) to human and bovine serum albumins (HSA and BSA, respectively). Regression analysis of the triplet decay of the drug (lambda = 360 nm) in the presence of the proteins led to a satisfactory fitting when considering a set of three lifetimes; the corresponding A(free), A(I) and A(II) preexponential coefficients can be correlated with the presence of FBPMe in the bulk solution and within the two known binding sites. The most remarkable differences between HSA and BSA were found under nonsaturating conditions; thus, when the [FBPMe]/[SA] ratio was 1: 1, all the drug was bound to HSA, whereas 20-30% of it remained free in the bulk solution in the presence of BSA. The UP approach was also applicable to the study of more complex FBPMe/HSA/BSA mixtures; the obtained results were in good agreement with the previous findings in FBPMe/HSA and FBPMe/BSA systems. This suggests the possibility of making use of the transient triplet-triplet absorption for investigating the distribution of a drug between several compartments in different host biomolecules., Financial support from the MCYT (CTQ2004-03811 and CTQ2007-67010) and the Generalitat Valenciana (GV06/099) is gratefully acknowledged. I.V. thanks MEC for a fellowship.

Financial support from the MEC (Grant CTQ2007-67010 and predoctoral fellowship to C.J.B. and I. V.), from the Carlos III Institute of Health (Grant RIRAAF, RETICS program) and from the Generalitat Valenciana (Prometeo Program) is gratefully acknowledged.

[EN] Fluorescence and laser-flash photolysis measurements have been performed on two pairs of diastereomeric dyads that contain the nonsteroidal anti-inflammatory drug (S)- or (R)-flurbiprofen (FBP) and (S)-tryptophan (Trp), which is a relevant amino acid present in site I of human serum albumin. The fluorescence spectra were obtained when subjected to excitation at 266 nm, where similar to 60% of the light is absorbed by FBP and similar to 40% is absorbed by Trp; the most remarkable feature observed in all dyads was a dramatic fluorescence quenching, and the residual emission was assigned to the Trp chromophore. In addition, an exciplex emission was observed as a broad band between 380 and 500 nm, especially in the case of the (R,S) diastereomers. The fluorescence lifetimes (tau(F)) at lambda(em) = 340 nm were clearly shorter in the dyads than in Trp-derived model compounds; in contrast, the values of tau(F) at lambda(em) = 440 nm (exciplex) were much longer. On the other hand, the typical FBP triplet-triplet transient absorption spectrum was obtained when subjected to laser-flash photolysis, although the signals were less intense than when FBP was directly excited under the same conditions. The main photophysical events in FBP-Trp dyads can be summarized as follows: (1) most of the energy provided by the incident radiation at 266 nm reaches the excited singlet state of Trp ((1)Trp*), either via direct absorption by this chromophore or by singlet singlet energy transfer from excited FBP ((FBP)-F-1*); (2) a minor, yet stereoselective deactivation of (FBP)-F-1* leads to detectable exciplexes and/or radical ion pairs; (3) the main process observed is intramolecular (1)Trp* quenching; and (4) the first triplet excited-state of FBP can be populated by triplet-triplet energy transfer from excited Trp or by back-electron transfer within the charge-separated states., Financial support from the MCYT (CTQ2004-03811) and the Generalitat Valenciana (GV06/099) is gratefully acknowledged. Author I.V. thanks MEC for a fellowship.

[EN] Four new bichromophoric naphthalene dyads in form of diastereomeric pairs are synthesized and photophysically characterized. For a closely related group of compounds (NAP-NAP MNAP-NAP, MNAP-MNAP), systematic variation of the urea linker led to the observation of intramolecular fluorescence quenching for tertiary urea, but not for secondary urea. Chiral information contained in the dyads has an impact on the efficiency of this process. Furthermore, for the case of (RR)-MNAP-MNAP excimer formation in the pre-organized dyad was noted, while being absent in the corresponding (R,S) diastereomer. These differences in photophysical behavior are ascribed to the geometry of the linker and to the chiral information contained in the diastereomeric compounds. Other dyads, such as NPX-NPX, included naproxen as the chromophore. For these compounds strong excimer emission is obtained, however, only very small diastereodifferentiation is observed for the two investigated stereoisomers., We acknowledge the financial support by the Spanish Ministry of Education and Science (Grant No. CTQ 2004-03811, doctoral fellowships for S.A. and I.V., Ramón y Cajal grant for U.P.).

[EN] The triplet excited states of (S)- and (R)-flurbiprofen (FBP) have been used as reporters for the microenvironments experienced within the binding sites of human and bovine serum albumins. Regression analysis of triplet decay provides valuable information on the degree of protection that these excited states are afforded from attack by a second FBP molecule, oxygen, or other reagents. The multiexponential fitting of these decays can be satisfactorily correlated with the distribution of the drug among the two binding sites and its presence as the noncomplexed form in the bulk solution. This assignment has been confirmed by using (S)-ibuprofen or capric acid as selective site II replacement probes. Triplet lifetimes and site occupancy are sensitive to the type of serum albumin employed (human versus bovine). Finally, the binding behaviour of (S)- and (R)-FBP exhibits little stereoselectivity., Financial support from the MCYT (CTQ2004-03811) and the Generalitat Valenciana (GV06/099) isgratefully acknowledged. I.V. and C.J.B. thank “Ministerio de Educación y Ciencia” for a fellowship.

[EN] The triplet excited state of flurbiprofen methyl ester (FBPMe) has been used as a chiral reporter for the two binding sites of human serum albumin (HSA). The occupation level of the binding sites has been estimated from regression analysis of the triplet decays at several [FBPMe]/[HSA] ratios. The data agree with two high affinity binding sites (I and II) that are populated to a different extent. A remarkable stereodifferentiation has been found in the drug triplet lifetimes within the protein microenvironment., The UPV (Grant PI 2003-0522 and fellowship to I.V.), the MEC (Grant CTQ2004-03811), and Generalitat Valenciana (Grant GV2004-0536 and Grupos03/082) are gratefully acknowledged for financial support.

The photochemical processes occurring in diastereomeric dyads (S, S)-1 and (S, R)-1, prepared by conjugation of (S)-2-(2-hydroxy-1,1'-biphenyl-4-yl) propanoic acid ((S)-BPOH) with (S)- and (R)-Trp, have been investigated. In acetonitrile, the fluorescence spectra of (S, S)-1 and (S, R)-1 were coincident in shape and position with that of (S)-BPOH, although they revealed a markedly stereoselective quenching. Since singlet energy transfer from BPOH to Trp is forbidden (5 kcal mol(-1) uphill), the quenching was attributed to thermodynamically favoured (according to Rehm-Weller) electron transfer or exciplex formation. Upon addition of 20% water, the fluorescence quantum yield of (S)-BPOH decreased, while only minor changes were observed for the dyads. This can be explained by an enhancement of the excited state acidity of (S)-BPOH, associated with bridging of the carboxy and hydroxy groups by water, in agreement with the presence of water molecules in the X-ray structure of (S)-BPOH. When the carboxy group was not available for coordination with water, as in the methyl ester (S)-BPOHMe or in the dyads, this effect was prevented; accordingly, the fluorescence quantum yields did not depend on the presence or absence of water. The fluorescence lifetimes in dry acetonitrile were 1.67, 0.95 and 0.46 ns for (S)-BPOH, (S, S)-1 and (S, R)-1, respectively, indicating that the observed quenching is indeed dynamic. In line with the steady-state and time-resolved observations, molecular modelling pointed to a more favourable geometric arrangement of the two interacting chromophores in (S, R)-1. Interestingly, this dyad exhibited a folded conformation in the solid state., Financial support from the Spanish Government (CTQ2010-14882, BES-2008-003314, JCI-2011-09926, PR2011-0581), from the Generalitat Valenciana (Prometeo 2008/090) and from the Universitat Politecnica de Valencia (PAID 05-11, 2766) is gratefully acknowledged.