De Novo Access to BODIPY C-Glycosides as Linker-Free Nonsymmetrical BODIPY-Carbohydrate Conjugates.

Recent years have witnessed an increasing interest in the synthesis and study of BODIPY-glycoconjugates. Most of the described synthetic methods toward these derivatives involve postfunctional modifications of the BODIPY core followed by the covalent attachment of the fluorophore and the carbohydrate through a "connector". Conversely, few de novo synthetic approaches to linker-free carbohydrate-BODIPY hybrids have been described. We have developed a reliable modular, de novo, synthetic strategy to linker-free BODIPY-sugar derivatives using the condensation of pyrrole C-glycosides with a pyrrole-carbaldehyde derivative mediated by POCl3. This methodology allows labeling of carbohydrate biomolecules with fluorescent-enough BODIPYs within the biological window, stable in aqueous media, and able to display singlet oxygen generation.

4,4'-Dicyano- versus 4,4'-Difluoro-BODIPYs in Chemoselective Postfunctionalization Reactions: Synthetic Advantages and Applications.

The presence of F or CN substituents at boron in BODIPYs causes a dramatic effect on their reactivity, which allows their chemoselective postfunctionalization. Thus, whereas 1,3,5,7-tetramethyl B(CN)2-BODIPYs displayed enhanced reactivity in Knoevenagel condensations with aldehydes, the corresponding BF2-BODIPYs can experience selective aromatic electrophilic substitution (SEAr) reactions in the presence of the former. These (selective) reactions have been employed in the preparation of BODIPY dimers and tetramers, with balanced fluorescence and singlet oxygen formation, and all-BODIPY trimers and heptamers, with potential application as light-harvesting systems.

Polar ammoniostyryls easily converting a clickable lipophilic BODIPY in an advanced plasma membrane probe.

A very simple, small and symmetric, but highly bright, photostable and functionalizable molecular probe for plasma membrane (PM) has been developed from an accessible, lipophilic and clickable organic dye based on BODIPY. To this aim, two lateral polar ammoniostyryl groups were easily linked to increase the amphiphilicity of the probe and thus its lipid membrane partitioning. Compared to the BODIPY precursor, the transversal diffusion across lipid bilayers of the ammoniostyryled BODIPY probe was highly reduced, as evidenced by fluorescence confocal microscopy on model membranes built up as giant unilamellar vesicles (GUVs). Moreover, the ammoniostyryl groups endow the new BODIPY probe with the ability to optically work (excitation and emission) in the bioimaging-useful red region, as shown by staining of the plasma membrane of living mouse embryonic fibroblasts (MEFs). Upon incubation, this fluorescent probe rapidly entered the cell through the endosomal pathway. By blocking the endocytic trafficking at 4 °C, the probe was confined within the PM of MEFs. Our experiments show the developed ammoniostyrylated BODIPY as a suitable PM fluorescent probe, and confirm the synthetic approach for advancing PM probes, imaging and science.

Triplet-triplet sensitizing within pyrene-based COO-BODIPY: a breaking molecular platform for annihilating photon upconversion.

We envisioned a new approach for achieving triplet-triplet annihilation-assisted photon upconversion based on the rational design of a heavy-atom-free, all-organic and photoactivatable triplet-triplet synergistic multichromophoric molecular assembly. This single molecular architecture is easily built by covalently anchoring triplet-annihilator units (pyrenes) to a triplet-photosensitizer moiety (BODIPY), to improve the effectiveness and probability of the required triplet-triplet energy transfer and the ulterior triplet-triplet annihilation. This unprecedented design takes advantage of the high synthetic accessibility and chemical versatility of the COO-BODIPY scaffold. The laser-induced photophysical characterization, assisted by computational simulations (quantum mechanics calculations at single molecular level and molecular dynamics in a solvent cage), identifies the key factors to finely control the intersystem crossing and reverse intersystem crossing probability, pivotal to improve energy transfer efficiency between the involved triplet states. Likewise, theoretical simulations highlight the relevance of the new photoactivable chromophoric design to promote intra- and inter-molecular triplet-triplet annihilation towards enhanced photon upconversion, yielding noticeable fluorescence from pyrene units even under unfavorable conditions (aerated solutions of low concentration at room temperature). The understanding of the complex dynamics sustained by this single molecular architecture could approach the next generation of chemically accessible and low-cost materials enabling fluorescence by photon upconversion mediated by triplet-triplet annihilation.

A Computational-Experimental Approach to Unravel the Excited State Landscape in Heavy-Atom Free BODIPY-Related Dyes.

We performed a time-gated laser-spectroscopy study in a set of heavy-atom free single BODIPY fluorophores, supported by accurate, excited-state computational simulations of the key low-lying excited states in these chromophores. Despite the strong fluorescence of these emitters, we observed a significant fraction of time-delayed (microseconds scale) emission associated with processes that involved passage through the triplet manifold. The accuracy of the predictions of the energy arrangement and electronic nature of the low-lying singlet and triplet excited states meant that an unambiguous assignment of the main deactivation pathways, including thermally activated delayed fluorescence and/or room temperature phosphorescence, was possible. The observation of triplet state formation indicates a breakthrough in the "classic" interpretation of the photophysical properties of the renowned BODIPY and its derivatives.

Development of Geometry-Controlled All-Orthogonal BODIPY Trimers for Photodynamic Therapy and Phototheragnosis.

We have established an easy synthetic protocol for selectively developing all-orthogonal BODIPY trimers with unprecedented geometries on the basis of selecting methyl oxidation versus electrophilic formylation of key dimeric precursors. Photophysical characterization together with biological assays unraveled the most suitable BODIPY-BODIPY geometrical arrangements within the trimer, forcing them to serve as molecular platforms for the development of new, advanced heavy-atom-free photosensitizers for photodynamic therapy and phototheragnosis.

BINOL blocks as accessible triplet state modulators in BODIPY dyes.

BINOL moieties of different electronic demand are useful blocks for enabling the photo-production and modulation of triplet excited states in readily-accesible BINOL-based O-BODIPY dyes from standard F-BODIPY precursors. The rapid and rational development of smarter triplet-enabling BODIPY dyes on the basis of this strategy (e.g., TADF biomarker 4a or room temperature phosphor 4g) paves the way for advancing photonic applications based on organic triplet photosensitizers.

Generation of multiple triplet states in an orthogonal bodipy dimer: a breakthrough spectroscopic and theoretical approach.

Generation of triplet states in assemblies of organic chromophores is extremely appealing for their potential use in optoelectronic applications. In this work, we investigate the intricacies of triplet state generation in an orthogonal BODIPY dimer by combining delayed photoemission techniques with electronic structure calculations. Our analysis provides a deep understanding of the electronic states involved, and describes different competing deactivation channels beyond prompt radiative decay. In particular, we identify charge-transfer (CT) mediated intersystem crossing (ISC) as the most likely mechanism for the triplet state generation in this system. The different emission bands at long times can be associated with delayed fluorescence, CT emission and phosphorescence from multiple low-energy triplets. Interestingly, the dependence of the yield of triplet state population and emission profiles with the solvent polarity evidences the decisive role of the CT configuration in the fate of the photoactivated dimer, controlling the relative ISC, reverse ISC, and internal conversion efficiencies. Overall, the present results provide a rather complete description of the delayed photophysics in the BODIPY dimer, but are not able to fully rationalize the unexpected photoluminescence recorded at long wavelengths (≥ 900 nm). We hypothesize that the origin of this emission, not present in BODIPY monomers, emerges from intermonomer interactions triggered by intramolecular distortions opening up a new vision in the controverted mechanism driving the photophysical behavior from orthogonally linked organic monomers.

Access to 2,6-Dipropargylated BODIPYs as "Clickable" Congeners of Pyrromethene-567 Dye: Photostability and Synthetic Versatility.

Hitherto unreported 2,6-dipropargyl-1,3,5,7-tetramethyl BODIPYs can be efficiently prepared by a Nicholas reaction/decomplexation protocol from 1,3,5,7-tetramethyl BODIPYs. The title compounds, which improve the BODIPY photostability by retaining their inherent photophysical and photochemical properties, can be engaged in efficient copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) "click-type" reactions with azido derivatives to provide all-BODIPY-triads or conjugated BODIPYs.

A Concise Route to Water-Soluble 2,6-Disubstituted BODIPY-Carbohydrate Fluorophores by Direct Ferrier-Type C-Glycosylation.

Novel, linker-free, BODIPY-carbohydrate derivatives containing sugar residues at positions C2 and C6 are efficiently obtained by, hitherto unreported, Ferrier-type C-glycosylation of 8-aryl-1,3,5,7-tetramethyl BODIPYs with commercially available tri-O-acetyl-d-glucal followed by saponification. This transformation, which involves the electrophilic aromatic substitution (SEAr) of the dipyrrin framework with an allylic oxocarbenium ion, provides easy access to BODIPY-carbohydrate hybrids with excellent photophysical properties and a weaker tendency to aggregate in concentrated water solutions.

A Concise Synthesis of a BODIPY-Labeled Tetrasaccharide Related to the Antitumor PI-88.

A convergent synthetic route to a tetrasaccharide related to PI-88, which allows the incorporation of a fluorescent BODIPY-label at the reducing-end, has been developed. The strategy, which features the use of 1,2-methyl orthoesters (MeOEs) as glycosyl donors, illustrates the usefulness of suitably-designed BODIPY dyes as glycosyl labels in synthetic strategies towards fluorescently-tagged oligosaccharides.

From photosensitizers to light harvesters adapting the molecular structure in all-BODIPY assemblies.

Herein we detail a protocol to design dyads and triads based solely on BODIPY dyes as halogen-free singlet oxygen photosensitizers or energy transfer molecular cassettes. The conducted photonic characterization reveals the key role of the BODIPY-BODIPY linkage to finely modulate the balance between the triplet state population and fluorescence decay.

Mitochondria selective trackers for long-term imaging based on readily accessible neutral BODIPYs.

We report the design of a new model based on a small neutral 8-aryl-3-formylBODIPY and its suitability to develop privileged highly bright and photostable fluorescent probes for selective and, more importantly, covalent staining of mitochondria.

First Lanthanide Complex for De Novo Phasing in Native Protein Crystallography at 1 Å Radiation.

Phasing agents enabling de novo protein structure determination at ca. 1 Å, the wavelength corresponding to the maximum intensity of the synchrotron facilities applied in biomacromolecular crystallography, have been long sought-after. The first phasing agent designed for solving native protein structures at 0.97934 Å is described herein. The agent consists of a neutral ytterbium(III)-caged complex that exhibits higher anomalous signals at shorter wavelengths when compared to the best, currently applied lanthanide-based phasing agents, all of them based on gadolinium or terbium. As a proof of principle, the complex allows determining the 3D structure of a 36 kDa protein without setting the incident beam wavelength at the metal absorption edge, the strategy followed to date to gain the strongest anomalous signal even at the expense of crystallographic resolution. The agent becomes nondisruptive to the diffraction quality of the marked crystals and allows determining accurate phases, both leading to high-quality electron-density maps that enable the full tracing of the protein structure only with one agent unit bound to the protein. The high phasing power, efficient binding to the protein, low metal-macromolecule ratio, and easy handling support the developed Yb(III) complex as the best phasing agent for X-ray crystallography of a complex biomacromolecule without using modified analogues.

Multichromophoric COO-BODIPYs: an advantageous design for the development of energy transfer and electron transfer systems.

COO-BODIPYs are highlighted as cutting edge scaffolds for easy access to a new generation of multichromophoric architectures with enhanced (photo)chemical stability, showing either boosted capability for excitation energy transfer, glow fluorescence and laser emission, or photoinduced electron transfer. The new finding paves the way for the rapid development of smarter organic dyes for advancing photonics and optoelectronics.

Red/NIR Thermally Activated Delayed Fluorescence from Aza-BODIPYs.

The search for long-lived red and NIR fluorescent dyes is challenging and hitherto scarcely reported. Herein, the viability of aza-BODIPY skeleton as a promising system for achieving thermal activated delayed fluorescent (TADF) probes emitting in this target region is demonstrated for the first time. The synthetic versatility of this scaffold allows the design of energy and charge transfer cassettes modulating the stereoelectronic properties of the energy donors, the spacer moieties and the linkage positions. Delayed emission from these architectures is recorded in the red spectral region (695-735 nm) with lifetimes longer than 100 µs in aerated solutions at room temperature. The computational-aided photophysical study under mild and hard irradiation regimes disclose the interplay between molecular structure and photonic performance to develop long-lived fluorescence red emitters through thermally activated reverse intersystem crossing. The efficient and long-lasting NIR emission of the newly synthesized aza-BODIPY systems provides a basis to develop advanced optical materials with exciting and appealing photonic response.

BCl3-Activated Synthesis of COO-BODIPY Laser Dyes: General Scope and High Yields under Mild Conditions.

A general and straightforward method for the synthesis of COO-BODIPYs from F-BODIPYs and carboxylic acids is established. The method is based on the use of boron trichloride to activate the involved substitution of fluorine, which leads to high yields through rapid reactions under soft conditions. This mild method opens the way to unprecedented laser dyes with outstanding efficiencies and photostabilities, which are difficult to obtain by the current methods.

BODIPYs as Chemically Stable Fluorescent Tags for Synthetic Glycosylation Strategies towards Fluorescently Labeled Saccharides.

A series of fluorescent boron-dipyrromethene (BODIPY, 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) dyes have been designed to participate, as aglycons, in synthetic oligosaccharide protocols. As such, they served a dual purpose: first, by being incorporated at the beginning of the process (at the reducing-end of the growing saccharide moiety), they can function as fluorescent glycosyl tags, facilitating the detection and purification of the desired glycosidic intermediates, and secondly, the presence of these chromophores on the ensuing compounds grants access to fluorescently labeled saccharides. In this context, a sought-after feature of the fluorescent dyes has been their chemical robustness. Accordingly, some BODIPY derivatives described in this work can withstand the reaction conditions commonly employed in the chemical synthesis of saccharides; namely, glycosylation and protecting-group manipulations. Regarding their photophysical properties, the BODIPY-labeled saccharides obtained in this work display remarkable fluorescence efficiency in water, reaching quantum yield values up to 82 %, as well as notable lasing efficiencies and photostabilities.

BODIPYs revealing lipid droplets as valuable targets for photodynamic theragnosis.

Endowing BODIPY PDT agents with the ability to probe lipid droplets is demonstrated to boost their phototoxicity, allowing the efficient use of highly fluorescent dyes (poor ROS sensitizers) as phototoxic agents. Conversely, this fact opens the way to the development of highly bright ROS photosensitizers for performing photodynamic theragnosis (fluorescence bioimaging and photodynamic therapy) from a single simple agent. On the other hand, the noticeable capability of some of the reported dyes to probe lipid droplets in different cell lines under different conditions reveals their use as privileged probes for advancing the study of interesting lipid droplets by fluorescence microscopy.

Tuning the Photonic Behavior of Symmetrical bis-BODIPY Architectures: The Key Role of the Spacer Moiety.

Herein we describe the synthesis, computationally assisted spectroscopy, and lasing properties of a new library of symmetric bridged bis-BODIPYs that differ in the nature of the spacer. Access to a series of BODIPY dimers is straightforward through synthetic modifications of the pending ortho-hydroxymethyl group of readily available C-8 (meso) ortho-hydroxymethyl phenyl BODIPYs. In this way, we have carried out the first systematic study of the photonic behavior of symmetric bridged bis-BODIPYs, which is effectively modulated by the length and/or stereoelectronic properties of the spacer unit. The designed bis-BODIPYs display bright fluorescence and laser emission in non-polar media. The fluorescence response is governed by the induction of a non-emissive intramolecular charge transfer (ICT) process, which is significantly enhanced in polar media. The effectiveness of the fluorescence quenching and also the prevailing charge transfer mechanism (from the spacer itself or between the BODIPY units) rely directly on the electron-releasing ability of the spacer. Moreover, the linker moiety can also promote intramolecular excitonic interactions, leading to excimer-like emission characterized by new spectral bands and the lengthening of lifetimes. The substantial influence of the bridging moiety on the emission behavior of these BODIPY dyads and their solvent-sensitivity highlight the intricate molecular dynamics upon excitation in multichromophoric systems. In this regard, the present work represents a breakthrough in the complex relationship between the molecular structure of the chromophores and their photophysical signatures, thus providing key guidelines for rationalizing the design of tailored bis-BODIPYs with potential advanced applications.