Structure-Photoreactivity Studies of BODIPY Photocages: Limitations of the Activation Barrier for Optimizing Photoreactions.

BODIPY photocages are photoreactive chromophores that release covalently linked cargo upon absorption of visible light. Here, we used computations of the T1 photoheterolysis barrier to ascertain whether a computational approach could assist in a priori structure design by identifying new structures with higher quantum yields of photorelease. The electronic structure-photoreactivity relationships were elucidated for boron-substituted and core-functionalized 2-substituted BODIPY photocages as well as aryl substitutions at the meso-methyl position. Although there is a clear trend for the 2-substituted derivatives, with donor-substituted derivatives featuring both lower computed barriers and higher experimental quantum yields, no trend in the quantum yield with the computed activation barrier is found for the meso-methyl-substituted or boron-substituted derivatives. The lack of a correlation between the experimental quantum yield with the computed barrier in the latter two substitution cases is attributed to the substituents having larger effects on the rates of competing channels (internal conversion and competitive photoreactions) than on the rate of the photoheterolysis channel. Thus, although in some cases computed photoreaction barriers can aid in identifying structures with higher quantum yields, the ignored impacts of how changing the structure affects the rates of competing photophysical/photochemical channels limit the effectiveness of this single-parameter approach.

Optical and EPR Detection of a Triplet Ground State Phenyl Nitrenium Ion.

Nitrenium ions are important reactive intermediates participating in the synthetic chemistry and biological processes. Little is known about triplet phenyl nitrenium ions regarding their reactivity, lifetimes, spectroscopic features, and electronic configurations, and no ground state triplet nitrenium ion has been directly detected. In this work, m-pyrrolidinyl-phenyl hydrazine hydrochloride (1) is synthesized as the photoprecursor to photochemically generate the corresponding m-pyrrolidinyl-phenyl nitrenium ion (2), which is computed to adopt a π, π* triplet ground state. A combination of femtosecond (fs) and nanosecond (ns) transient absorption (TA) spectroscopy, cryogenic continuous-wave electronic paramagnetic resonance (CW-EPR) spectroscopy, computational analysis, and photoproduct studies was performed to elucidate the photolysis pathway of 1 and offers the first direct experimental detection of a ground state triplet phenyl nitrenium ion. Upon photoexcitation, 1 forms S1, where bond heterolysis occurs and the NH3 leaving group is extruded in 1.8 ps, generating a vibrationally hot, spin-conserving closed-shell singlet phenyl nitrenium ion (12) that undergoes vibrational cooling in 19 ps. Subsequent intersystem crossing takes place in 0.5 ns, yielding the ground state triplet phenyl nitrenium ion (32), with a lifetime of 0.8 µs. Unlike electrophilic singlet phenyl nitrenium ions, which react rapidly with nucleophiles, this triplet phenyl nitrenium reacts through sequential H atom abstractions, resulting in the eventual formation of the reduced m-pyrrolidinyl-aniline as the predominant stable photoproduct. Supporting the triplet ground state, continuous irradiation of 1 in a glassy matrix at 80 K in an EPR spectrometer forms a paramagnetic triplet species, consistent with a triplet nitrenium ion.

Simple Air-Stable [3]Radialene Anion Radicals as Environmentally Switchable Catholytes in Water.

The hexacyano[3]radialene radical anion (1) is an attractive catholyte material for use in redox flow battery (RFB) applications. The substitution of cyano groups with ester moieties enhances solubility while maintaining redox reversibility and favorable redox potentials. Here we show that these ester-functionalized, hexasubstituted [3]radialene radical anions dimerize reversibly in water. The dimerization mode is dependent on the substitution pattern and can be switched in solution. Stimuli-responsive behavior is achieved by exploiting an unprecedented tristate switching mechanism, wherein the radical can be toggled between the free radical, a π-dimer, and a σ-dimer-each with dramatically different optical, magnetic, and redox properties-by changing the solvent environment, temperature, or salinity. The symmetric, triester-tricyano[3]radialene (3) forms a solvent-responsive, σ-dimer in water that converts to the radical anion with the addition of organic solvents or to a π-dimer in brine solutions. Diester-tetracyano[3]radialene (2) exists primarily as a π-dimer in aqueous solutions and a radical anion in organic solvents. The dimerization behavior of both 2 and 3 is temperature dependent in methanol solutions. Dimerization equilibrium has a direct impact on catholyte stability during galvanostatic charge-discharge cycling in static H-cells. Specifically, conditions that favor the free radical anion or π-dimer exhibit significantly enhanced cycling profiles.

meso-Methyl BODIPY Photocages: Mechanisms, Photochemical Properties, and Applications.

meso-methyl BODIPY photocages have recently emerged as an exciting new class of photoremovable protecting groups (PPGs) that release leaving groups upon absorption of visible to near-infrared light. In this Perspective, we summarize the development of these PPGs and highlight their critical photochemical properties and applications. We discuss the absorption properties of the BODIPY PPGs, structure-photoreactivity studies, insights into the photoreaction mechanism, the scope of functional groups that can be caged, the chemical synthesis of these structures, and how substituents can alter the water solubility of the PPG and direct the PPG into specific subcellular compartments. Applications that exploit the unique optical and photochemical properties of BODIPY PPGs are also discussed, from wavelength-selective photoactivation to biological studies to photoresponsive organic materials and photomedicine.

Photo-Uncaging by C(sp3)-C(sp3) Bond Cleavage Restores ß-Lapachone Activity.

ß-Lapachone is an ortho-naphthoquinone natural product with significant antiproliferative activity but suffers from adverse systemic toxicity. The use of photoremovable protecting groups to covalently inactivate a substrate and then enable controllable release with light in a spatiotemporal manner is an attractive prodrug strategy to limit toxicity. However, visible light-activatable photocages are nearly exclusively enabled by linkages to nucleophilic functional sites such as alcohols, amines, thiols, phosphates, and sulfonates. Herein, we report covalent inactivation of the electrophilic quinone moiety of ß-lapachone via a C(sp3)-C(sp3) bond to a coumarin photocage. In contrast to ß-lapachone, the designed prodrug remained intact in human whole blood and did not induce methemoglobinemia in the dark. Under light activation, the C-C bond cleaves to release the active quinone, recovering its biological activity when evaluated against the enzyme NQO1 and human cancer cells. Investigations into this report of a C(sp3)-C(sp3) photoinduced bond cleavage suggest a nontraditional, radical-based mechanism of release beginning with an initial charge-transfer excited state. Additionally, caging and release of the isomeric para-quinone, α-lapachone, are demonstrated. As such, we describe a photocaging strategy for the pair of quinones and report a unique light-induced cleavage of a C-C bond. We envision that this photocage strategy can be extended to quinones beyond ß- and α-lapachone, thus expanding the chemical toolbox of photocaged compounds.

Photocaged DNA-Binding Photosensitizer Enables Photocontrol of Nuclear Entry for Dual-Targeted Photodynamic Therapy.

Photodynamic therapy (PDT) is a clinically approved cancer treatment that requires a photosensitizer (PS), light, and molecular oxygen─a combination which produces reactive oxygen species (ROS) that can induce cancer cell death. To enhance the efficacy of PDT, dual-targeted strategies have been explored where two photosensitizers are administered and localize to different subcellular organelles. To date, a single small-molecule conjugate for dual-targeted PDT with light-controlled nuclear localization has not been achieved. We designed a probe composed of a DNA-binding PS (Br-DAPI) and a photosensitizing photocage (WinterGreen). Illumination with 480 nm light removes WinterGreen from the conjugate and produces singlet oxygen mainly in the cytosol, while Br-DAPI localizes to nuclei, binds DNA, and produces ROS using one- or two-photon illumination. We observe synergistic photocytotoxicity in MCF7 breast cancer cells, and a reduction in size of three-dimensional (3D) tumor spheroids, demonstrating that nuclear/cytosolic photosensitization using a single agent can enhance PDT efficacy.

Efficiency of Functional Group Caging with Second-Generation Green- and Red-Light-Labile BODIPY Photoremovable Protecting Groups.

BODIPY-based photocages release substrates by excitation with wavelengths in the visible to near-IR regions. The recent development of more efficient BODIPY photocages spurred us to evaluate the scope and efficiency of these second-generation boron-methylated green-light and red-light-absorbing BODIPY photocages. Here, we show that these more photosensitive photocages release amine, alcohol, phenol, phosphate, halides, and carboxylic acid derivatives with much higher quantum yields than first-generation BODIPY photocages and excellent chemical yields. Chemical yields are near-quantitative for the release of all functional groups except the photorelease of amines, which react with concomitantly photogenerated singlet oxygen. In these cases, high chemical yields for photoreleased amines are restored by irradiation under an inert atmosphere. The photorelease quantum yield has a weak relationship with the leaving group pKa of the green-absorbing BODIPY photocages but little relationship with the red-absorbing derivatives, suggesting that factors other than leaving group quality impact the quantum yield. For the photorelease of alcohols, in all cases a carbonate linker (that loses CO2 upon photorelease) significantly increases both the quantum yield and the chemical yield compared to those for direct photorelease via the ether.

Mechanistic Insight into Phenol Dearomatization by Hypervalent Iodine: Direct Detection of a Phenoxenium Cation.

Phenol dearomatization is one of several oxidation reactions enabled by hypervalent iodine reagents. However, the presence of a proposed free phenoxenium intermediate in phenol dearomatization is a matter of debate in the literature. Here, we report the unambiguous detection of a free phenoxenium intermediate in the reaction of an electron-rich phenol, 2,4,6-trimethoxyphenol, and (diacetoxyiodo)benzene using UV-vis and resonance Raman spectroscopies. In contrast, we predominantly detect single electron oxidation products of less electron-rich phenols or alkoxy-substituted aromatics in their reaction with (diacetoxyiodo)benzene using UV-vis and electron paramagnetic resonance (EPR) spectroscopies. We conclude that the often-postulated free phenoxenium intermediate, while possible with highly stabilizing substituents, is unlikely to be a general mechanistic pathway in the reaction of typical phenols with hypervalent iodine reagents. The polar solvent 1,1,1,3,3,3-hexafluoroisopropanol (HFIP) or the use of more strongly oxidizing hypervalent iodine reagents, such as [bis(trifluoroacetoxy)iodo]benzene (PIFA) or [hydroxy(tosyloxy)iodo]benzene (HTIB), can help reduce the formation of radical byproducts and favors the formation of phenoxenium intermediates.

Photo-labile BODIPY protecting groups for glycan synthesis.

Protective groups that can be selectively removed under mild conditions are an essential aspect of carbohydrate chemistry. Groups that can be selectively removed by visible light are particularly attractive because carbohydrates are transparent to visible light. Here, different BODIPY protecting groups were explored for their utility during glycan synthesis. A BODIPY group bearing a boron difluoride unit is stable during glycosylations but can be cleaved with green light as illustrated by the assembly of a trisaccharide.

Generation and direct observation of a triplet arylnitrenium ion.

Nitrenium ions are important reactive intermediates in both chemistry and biology. Although singlet nitrenium ions are well-characterized by direct methods, the triplet states of nitrenium ions have never been directly detected. Here, we find that the excited state of the photoprecursor partitions between heterolysis to generate the singlet nitrenium ion and intersystem crossing (ISC) followed by a spontaneous heterolysis process to generate the triplet p-iodophenylnitrenium ion (np). The triplet nitrenium ion undergoes ISC to generate the ground singlet state, which ultimately undergoes proton and electron transfer to generate a long-lived radical cation that further generates the reduced p-iodoaniline. Ab Initio calculations were performed to map out the potential energy surfaces to better understand the excited state reactivity channels show that an energetically-accessible singlet-triplet crossing lies along the N-N stretch coordinate and that the excited triplet state is unbound and spontaneously eliminates ammonia to generate the triplet nitrenium ion. These results give a clearer picture of the photophysical properties and reactivity of two different spin states of a phenylnitrenium ion and provide the first direct glimpse of a triplet nitrenium ion.

Controlling Antimicrobial Activity of Quinolones Using Visible/NIR Light-Activated BODIPY Photocages.

Controlling the activity of a pharmaceutical agent using light offers improved selectivity, reduction of adverse effects, and decreased environmental build-up. These benefits are especially attractive for antibiotics. Herein, we report a series of photoreleasable quinolones, which can be activated using visible/NIR light (520-800 nm). We have used BODIPY photocages with strong absorption in the visible to protect two different quinolone-based compounds and deactivate their antimicrobial properties. This activity could be recovered upon green or red light irradiation. A comprehensive computational study provides new insight into the reaction mechanism, revealing the relevance of considering explicit solvent molecules. The triplet excited state is populated and the photodissociation is assisted by the solvent. The light-controlled activity of these compounds has been assessed on a quinolone-susceptible E. coli strain. Up to a 32-fold change in the antimicrobial activity was measured.

Steric Hindrance Favors σ Dimerization over π Dimerization for Julolidine Dicyanomethyl Radicals.

Metastable radicals exist in a steady-state equilibrium in solution with dimers, which can be either σ dimers or π dimers. Here, we show that steric hindrance at the para position causes julolidine-derived dicyanomethyl radicals to form σ dimers rather than π dimers, the opposite behavior as seen in other carbon-centered radicals, where steric hindrance typically favors pimerization. The change in dimerization mode can be attributed to weaker London dispersion forces and a decreased orbital overlap in the sterically hindered dicyanomethyl radical π dimers, while the bulky groups exert relatively little effect on the energy of the σ dimer.

Anti-Aromaticity Relief as an Approach to Stabilize Free Radicals.

A new strategy to stabilize free radicals electronically is described by conjugating formally antiaromatic substituents to the free radical. With an antiaromatic substituent, the radical acts as an electron sink to allow configuration mixing of a low-energy zwitterionic state that provides antiaromaticity relief to the substituent. A combination of X-ray crystallography, VT-EPR and VT-UV/Vis spectroscopy, as well as computational analysis, was used to investigate this phenomenon. We find that this strategy of antiaromaticity relief is successful at stabilizing radicals, but only if the antiaromatic substituent is constrained to be planar by synthetically imposed conformational restraints that enable state mixing. This work leads to the counterintuitive finding that increasing the antiaromaticity of the radical substituent leads to greater radical stability, providing proof of concept for a new stereoelectronic approach for stabilizing free radicals.

Multiwavelength Control of Mixtures Using Visible Light-Absorbing Photocages.

Selective deprotection of functional groups using different wavelengths of light is attractive for materials synthesis as well as for achieving independent photocontrol over substrates in biological systems. Here, we show that mixtures of recently developed visible light-absorbing BODIPY-derived photoremovable protecting groups (PRPGs) and a coumarin-derived PRPG can undergo wavelength-selective activation, giving independent optical control over a mixture of photocaged substrates using biologically benign long-wavelength light.

Efficient Far-Red/Near-IR Absorbing BODIPY Photocages by Blocking Unproductive Conical Intersections.

Photocages are light-sensitive chemical protecting groups that give investigators control over activation of biomolecules using targeted light irradiation. A compelling application of far-red/near-IR absorbing photocages is their potential for deep tissue activation of biomolecules and phototherapeutics. Toward this goal, we recently reported BODIPY photocages that absorb near-IR light. However, these photocages have reduced photorelease efficiencies compared to shorter-wavelength absorbing photocages, which has hindered their application. Because photochemistry is a zero-sum competition of rates, improvement of the quantum yield of a photoreaction can be achieved either by making the desired photoreaction more efficient or by hobbling competitive decay channels. This latter strategy of inhibiting unproductive decay channels was pursued to improve the release efficiency of long-wavelength absorbing BODIPY photocages by synthesizing structures that block access to unproductive singlet internal conversion conical intersections, which have recently been located for simple BODIPY structures from excited state dynamic simulations. This strategy led to the synthesis of new conformationally restrained boron-methylated BODIPY photocages that absorb light strongly around 700 nm. In the best case, a photocage was identified with an extinction coefficient of 124000 M-1 cm-1, a quantum yield of photorelease of 3.8%, and an overall quantum efficiency of 4650 M-1 cm-1 at 680 nm. This derivative has a quantum efficiency that is 50-fold higher than the best known BODIPY photocages absorbing >600 nm, validating the effectiveness of a strategy for designing efficient photoreactions by thwarting competitive excited state decay channels. Furthermore, 1,7-diaryl substitutions were found to improve the quantum yields of photorelease by excited state participation and blocking ion pair recombination by internal nucleophilic trapping. No cellular toxicity (trypan blue exclusion) was observed at 20 µM, and photoactivation was demonstrated in HeLa cells using red light.

Solvent-Responsive Radical Dimers.

An air- and thermally stable aryl dicyanomethyl radical is reported that switches between two dimeric forms-a σ dimer and a π dimer-by changing the solvent. The two dimer forms exhibit unique optical properties leading to solvatochromic behavior. The solvent-responsive behavior of these radicals can be explained by the higher polarizability of the pimer than the σ dimer that leads to pimer stabilization in polar solvents.

Direct Photorelease of Alcohols from Boron-Alkylated BODIPY Photocages.

BODIPY photocages allow the release of substrates using visible light irradiation. They have the drawback of requiring reasonably good leaving groups for photorelease. Photorelease of alcohols is often accomplished by attachment with carbonate linkages, which upon photorelease liberate CO2 and generate the alcohol. Here, we show that boron-alkylated BODIPY photocages are capable of directly photoreleasing both aliphatic alcohols and phenols upon irradiation via photocleavage of ether linkages. Direct photorelease of a hydroxycoumarin dye was demonstrated in living HeLa cells.

Water-Soluble BODIPY Photocages with Tunable Cellular Localization.

Photoactivation of bioactive molecules allows manipulation of cellular processes with high spatiotemporal precision. The recent emergence of visible-light excitable photoprotecting groups has the potential to further expand the established utility of the photoactivation strategy in biological applications by offering higher tissue penetration, diminished phototoxicity, and compatibility with other light-dependent techniques. Nevertheless, a critical barrier to such applications remains the significant hydrophobicity of most visible-light excitable photocaging groups. Here, we find that applying the conventional 2,6-sulfonation to meso-methyl BODIPY photocages is incompatible with their photoreaction due to an increase in the excited state barrier for photorelease. We present a simple, remote sulfonation solution to BODIPY photocages that imparts water solubility and provides control over cellular permeability while retaining their favorable spectroscopic and photoreaction properties. Peripherally disulfonated BODIPY photocages are cell impermeable, making them useful for modulation of cell-surface receptors, while monosulfonated BODIPY retains the ability to cross the cellular membrane and can modulate intracellular targets. This new approach is generalizable for controlling BODIPY localization and was validated by sensitization of mammalian cells and neurons by visible-light photoactivation of signaling molecules.

Anomalous Electronic Properties of Iodous Materials: Application to High-Spin Reactive Intermediates and Conjugated Polymers.

Manipulating frontier orbital energies of aromatic molecules with substituents is key to a variety of chemical and material applications. Here, we investigate a simple strategy for achieving high-energy in-plane orbitals for aromatics simply by positioning iodine atoms next to each other. The lone pair orbitals on the iodines mix to give a high-energy in-plane σ-antibonding orbital as the highest occupied molecular orbital (HOMO). We show that this effect can be used to manipulate orbital gaps, the symmetry of the highest occupied orbital, and the adopted electronic state for reactive intermediates. This electronic effect is not limited to reactive intermediates, and we demonstrate that this iodine buttressing strategy also can be used to achieve small HOMO-lowest unoccupied molecular orbital (HOMO-LUMO) gaps in organic electronic materials. Iodinated oligomers of several of the most popular conducting polymers are computed to have smaller HOMO-LUMO gaps than the unsubstituted materials. This iodine buttressing approach for generating high-energy in-plane HOMOs is anticipated to be highly general. While the unusual properties of fluorous materials are well established, at the other extreme on the periodic table novel properties of iodous materials may await discovery.

Spin Delocalization, Polarization, and London Dispersion Forces Govern the Formation of Diradical Pimers.

Some free radicals are stable enough to be isolated, but most are either unstable transient species or exist as metastable species in equilibrium with a dimeric form, usually a spin-paired sigma dimer or a pi dimer (pimer). To gain insight into the different modes of dimerization, we synthesized and evaluated a library of 15 aryl dicyanomethyl radicals in order to probe what structural and molecular parameters lead to σ- versus π-dimerization. We evaluated the divergent dimerization behavior by measuring the strength of each radical association by variable-temperature electron paramagnetic resonance spectroscopy, determining the mode of dimerization (σ- or π-dimer) by UV-vis spectroscopy and X-ray crystallography, and performing computational analysis. We evaluated three different hypotheses to explain the difference in the dimerization behavior: (1) that the dimerization behavior is dictated by radical spin densities; (2) that it is dictated by radical polarizability; (3) that it is dictated by London dispersion stabilization of the pimer. However, no single parameter model in itself was predictive. Two-parameter models incorporating either the computed degree of spin delocalization or the radical polarizability as well as computed estimates for the attractive London dispersion forces in the π-dimers lead to improved forecasts of σ- vs π-dimerization mode, and suggest that a balance of spin delocalization of the isolated radical as well as attractive forces between the stacked radicals, govern the formation of diradical pimers.