Activatable Near-Infrared Fluorescence Imaging Using PEGylated Bacteriochlorin-Based Chlorin and BODIPY-Dyads as Probes for Detecting Cancer.

Near infrared (NIR) fluorescent probes are attractive tools for biomedical in vivo imaging due to the relatively deeper tissue penetration and lower background autofluorescence. Activatable probes are turned on only after binding to their target, further improving target to background ratios. However, the number of available activatable NIR probes is limited. In this study, we introduce two types of activatable NIR fluorophores derived from bacteriochlorin: chlorin-bacteriochlorin energy-transfer dyads and boron-dipyrromethene (BODIPY)-bacteriochlorin energy-transfer dyads. These fluorophores are characterized by multiple narrow excitation bands with relatively strong emission in the NIR. Targeted bacteriochlorin-based antibody or peptide probes have been previously limited by aggregation after conjugation. Polyethylene glycol (PEG) chains were added to improve the hydrophilicity without altering pharmacokinetics of the targeting moieties. These PEGylated bacteriochlorin-based activatable fluorophores have potential as targeted activatable, multicolor NIR fluorescent probes for in vivo applications.

An Alternate Route of Transforming meso-Tetraarylporphyrins to Porpholactams, and Their Conversion to Amine-Functionalized Imidazoloporphyrins.

A novel and efficient synthetic pathway toward known meso-tetraphenylporpholactams, also applicable to the synthesis of hitherto unknown and inaccessible meso-C6F5-substituted porpholactam, is detailed (dioxochlorin → dioxochlorin urea adduct → porpholactam). meso-Tetraphenylporpholactam was converted to an imidazoloporphyrin-α-triflate derivative that was demonstrated to be of utility for the generation of functionalized imidazoloporphyrins with a substituted amine adjacent to the outside N atom of the imidazole moiety (using pyridine, Et2NH, diethyliminodiacetic acetate, dipicolylamine (DPA), and cyclen). The DPA- and iminodiacetate-imidazoloporphyrin conjugates were structurally characterized. The chemosensing potential of the metal chelate-imidazoloporphyrin conjugates was evaluated, though their constrained metric parameters led to muted chemosensing responses to various divalent metal ions. The accessibility of the meso-arylporpholactams and the meso-tetraphenylimidazoloporphyrin triflate enables the continued exploration of porphyrin-like pyrrole-modified porphyrins that incorporate a nitrogen atom in place of a ß-carbon atom in their macrocycles.

Amphiphilic BODIPY-Hydroporphyrin Energy Transfer Arrays with Broadly Tunable Absorption and Deep Red/Near-Infrared Emission in Aqueous Micelles.

BODIPY-hydroporphyrin energy transfer arrays allow for development of a family of fluorophores featuring a common excitation band at 500 nm, tunable excitation band in the deep red/near-infrared window, and tunable emission. Their biomedical applications are contingent upon retaining their optical properties in an aqueous environment. Amphiphilic arrays containing PEG-substituted BODIPY and chlorins or bacteriochlorins were prepared and their optical and fluorescence properties were determined in organic solvents and aqueous surfactants. The first series of arrays contains BODIPYs with PEG substituents attached to the boron, whereas in the second series, PEG substituents are attached to the aryl at the meso positions of BODIPY. For both series of arrays, excitation of BODIPY at 500 nm results in efficient energy transfer to and bright emission of hydroporphyrin in the deep-red (640-660 nm) or near-infrared (740-760 nm) spectral windows. In aqueous solution of nonionic surfactants (Triton X-100 and Tween 20) arrays from the second series exhibit significant quenching of fluorescence, whereas properties of arrays from the first series are comparable to those observed in polar organic solvents. Reported arrays possess large effective Stokes shift (115-260 nm), multiple excitation wavelengths, and narrow, tunable deep-red/near-IR fluorescence in aqueous surfactants, and are promising candidates for a variety of biomedical-related applications.

Magnetic Circular Dichroism Spectroscopy of meso-Tetraphenylporphyrin-Derived Hydroporphyrins and Pyrrole-Modified Porphyrins.

A large set of free-base and transition-metal 5,10,15,20-tetraphenyl-substituted chlorins, bacteriochlorins, and isobacteriochlorins and their pyrrole-modified analogues were investigated by combined UV-visible spectroscopy, magnetic circular dichroism (MCD), density functional theory (DFT), and time-dependent DFT (TDDFT) approaches and their spectral characteristics were compared to those of the parent compounds, free-base tetraphenylporphyrin 1H2 and chlorin 2H2. It was shown that the nature of the pyrroline substituents in the chlorin derivatives dictates their specific UV-vis and MCD spectroscopic signatures. In all hydroporphyrin-like cases, MCD spectroscopy suggests that the ΔHOMO is smaller than the ΔLUMO for the macrocycle-centered frontier molecular orbitals. DFT and TDDFT calculations were able to explain the large broadening of the UV-vis and MCD spectra of the chlorin diones and their derivatives compared to the other hydroporphyrins and hydroporphyrin analogues. This study contributes to the further understanding of the electronic effects of replacing a pyrrole in porphyrins by pyrrolines or other five-membered heterocycles (oxazoles and imidazoles).

Effects of Strong Electronic Coupling in Chlorin and Bacteriochlorin Dyads.

Achieving tunable, intense near-infrared absorption in molecular architectures with properties suitable for solar light harvesting and biomedical studies is of fundamental interest. Herein, we report the photophysical, redox, and molecular-orbital characteristics of nine hydroporphyrin dyads and associated benchmark monomers that have been designed and synthesized to attain enhanced light harvesting. Each dyad contains two identical hydroporphyrins (chlorin or bacteriochlorin) connected by a linker (ethynyl or butadiynyl) at the macrocycle ß-pyrrole (3- or 13-) or meso (15-) positions. The strong electronic communication between constituent chromophores is indicated by the doubling of prominent absorption features, split redox waves, and paired linear combinations of frontier molecular orbitals. Relative to the benchmarks, the chlorin dyads in toluene show substantial bathochromic shifts of the long-wavelength absorption band (17-31 nm), modestly reduced singlet excited-state lifetimes (τS = 3.6-6.2 ns vs 8.8-12.3 ns), and increased fluorescence quantum yields (Φf = 0.37-0.57 vs 0.34-0.39). The bacteriochlorin dyads in toluene show significant bathochromic shifts (25-57 nm) and modestly reduced τS (1.6-3.4 ns vs 3.5-5.3 ns) and Φf (0.09-0.19 vs 0.17-0.21) values. The τS and Φf values for the bacteriochlorin dyads are reduced substantially (up to ∼20-fold) in benzonitrile. The quenching is due primarily to the increased S1 → S0 internal conversion that is likely induced by increased contribution of charge-resonance configurations to the S1 excited state in the polar medium. The fundamental insights gained into the physicochemical properties of the strongly coupled hydroporphyrin dyads may aid their utilization in solar-energy conversion and photomedicine.

Indachlorins: nonplanar indanone-annulated chlorin analogues with panchromatic absorption spectra between 300 and 900†nm.

Indaphyrins, pyrrole-modified porphyrins containing a cleaved pyrrole ß,ß'-bond and two annulated indanone moieties, possess unusually broadened and redshifted UV/Vis spectra because of their π-expanded chromophores. The parent free base indaphyrin has been crystallographically characterized, highlighting its strongly ruffled conformation incorporating a helimeric twist. It was shown to be susceptible to regiospecific derivatizations at the opposite side of the ring-cleaved pyrrole (dihydroxylation, followed by functional group transformations of the resulting diol functionality), generating indaphyrin-based chlorin analogues, indachlorins, that incorporate a dihydroxypyrroline, pyrrolindione, oxazolone, or a morpholine moiety. Structural modifications resulted in further broadening and hyper- and bathochromic shifts of the optical spectra, some of which possess a nearly panchromatic absorption between 300 to well above 900 nm. The extents to which these modifications affect their solid-state conformations were analyzed.

Mono- and Bisquinoline-Annulated Porphyrins from Porphyrin ß,ß'-Dione Oximes.

An acid-induced reaction of meso-tetraphenyl-2-hydroxyimino-3-oxoporphyrin leads, with concomitant loss of water, to a formal electrophilic aromatic substitution of the ortho-position of the phenyl group adjacent to the oxime, forming a quinoline moiety. Owing in part to the presence of a π-extended chromophore, the resulting meso-triphenylmonoquinoline-annulated porphyrin (λmax = 750 nm) possesses a much altered optical spectrum from that of the starting oxime (λmax = 667 nm). An oxidative DDQ-induced ring-closure process is also possible, generating the corresponding meso-triphenylmonoquinoline-annulated porphyrin quinoline N-oxide, possessing a slightly shifted and sharpened UV-vis spectrum (λmax = 737 nm). The connectivity of the chromophores was conclusively shown by NMR spectroscopy. Both ketone functionalities in meso-tetraphenyl-2,3-dioxoporphyrin can be converted, via the oxime and using the acid- or oxidant-induced reaction pathways, either in one step or in a stepwise fashion, to bisquinoline-annulated porphyrin (λmax = 775 nm) and its N-oxide (λmax = 779 nm), respectively. This process is complementary to a previously established pathway toward bisquinoline-annulated porphyrins. Their zinc(II), nickel(II), and palladium(II) complexes are also described. Several examples of the quinoline-annulated porphyrins were crystallographically characterized, proving their connectivity and showing their conformations that are extremely distorted from planarity. The work presents a full account on the synthesis, structure, and spectroscopic properties of these classes of NIR-absorbing dyes.

The evaluation of NIR-absorbing porphyrin derivatives as contrast agents in photoacoustic imaging.

Six free base tetrapyrrolic chromophores, three quinoline-annulated porphyrins and three morpholinobacteriochlorins, that absorb light in the near-IR range and possess, in comparison to regular porphyrins, unusually low fluorescence emission and (1)O2 quantum yields were tested with respect to their efficacy as novel molecular photo-acoustic imaging contrast agents in a tissue phantom, providing an up to ∼2.5-fold contrast enhancement over that of the benchmark contrast agent ICG. The testing protocol compares the photoacoustic signal output strength upon absorption of approximately the same light energy. Some relationships between photophysical parameters of the dyes and the resulting photoacoustic signal strength could be derived.

Formation, structure, and reactivity of meso-tetraaryl-chlorolactones, -porpholactams, and -chlorolactams, porphyrin and chlorin analogues incorporating oxazolone or imidazolone moieties.

Reaction of known meso-tetraarylporpholactone free bases 3, made from the corresponding porphyrins, with hydrazine produces three products: It converts the lactone functional group into an N-aminolactam moiety, generating porphyrin-like N-aminoporpholactams 8. It also reduces regioselectively the ß,ß'-double bond of the pyrrolic moiety opposite to the imidazolone in both the starting material and the N-aminoporpholactam, thus forming the chlorin-like chlorolactones 7 and N-aminochlorolactams 9. An equivalent set of reaction products is also derived from the reaction of porpholactones 3 with tosylhydrazide. Reductive N-N cleavage of the N-aminoporpholactams 8 generated the parent porpholactams 10. The molecular structures of all key compounds were shown by single crystal X-ray diffraction to be essentially planar. Porpholactam 10a can be converted in two steps (enolization and halogenation α to the imine, followed by reductive removal of the halogen) to known imidazoloporphyrin 5a, thus constituting the third independent pathway to replace a ß-carbon of a tetraphenylporphyrin by a nitrogen. All these transformations show the flexibility of our 'porphyrin breaking and mending' strategy toward the synthesis of novel porphyrin and chlorin analogues incorporating non-pyrrolic heterocycles that carry functionalities at their periphery.

meso-arylporpholactones and their reduction products.

The rational syntheses of meso-tetraaryl-3-oxo-2-oxaporphyrins 5, known as porpholactones, via MnO(4)(-)-mediated oxidations of the corresponding meso-tetraaryl-2,3-dihydroxychlorins (7) is detailed. Since chlorin 7 is prepared from the parent porphyrin 1, this amounts to a 2-step replacement of a pyrrole moiety in 1 by an oxazolone moiety. The stepwise reduction of the porpholactone 5 results in the formation of chlorin analogues, meso-tetraaryl-3-hydroxy-2-oxachlorin (11) and meso-tetraaryl-2-oxachlorins (12). The reactivity of 11 with respect to nucleophilic substitution by O-, N-, and S-nucleophiles is described. The profound photophysical consequences of the formal replacement of a pyrrole with an oxazolone (porphyrin-like chromophore) or (substituted) oxazole moiety (chlorin-like chromophore with, for the parent oxazolochlorin 12, red-shifted Q(x) band with enhanced oscillator strengths) are detailed and rationalized on the basis of SAC-CI and MNDO-PSDCI molecular orbital theory calculations. The single crystal X-ray structures of the porpholactones point at a minor steric interaction between the carbonyl oxygen and the flanking phenyl group. The essentially planar structures of all chromophores in all oxidation states prove that the observed optical properties originate from the intrinsic electronic properties of the chromophores and are not subject to conformational modulation.

Porpholactams and chlorolactams: replacement of a ß,ß'-double bond in meso-tetraphenyl-porphyrin and -chlorin by a lactam moiety.

Reaction of meso-tetraphenylporpholactone with hydrazine converts the lactone moiety to an N-aminolactam. It also reduces the opposite pyrrolic moiety of both the starting material and the N-aminolactam, generating chlorin-like chlorolactone and N-aminochlorolactam, respectively. Reductive N-N cleavage of the N-aminoporpholactam generates the parent porpholactam.

Helimeric porphyrinoids: stereostructure and chiral resolution of meso-tetraarylmorpholinochlorins.

The synthesis and chiral resolution of free-base and Ni(II) complexes of a number of derivatives of meso-tetraphenylmorpholinochlorins, with and without direct ß-carbon-to-o-phenyl linkages to the flanking phenyl groups, is described. The morpholinochlorins, a class of stable chlorin analogues, were synthesized in two to three steps from meso-tetraphenylporphyrin. The conformations and the relative stereostructures of a variety of free-base and Ni(II) complexes of these morpholinochlorins were elucidated by X-ray diffractometry. Steric and stereoelectronic arguments explain the relative stereoarray of the morpholino-substituents, which differ in the free-base and Ni(II) complexes, and in the monoalkoxy, ß-carbon-to-o-phenyl linked morpholinochlorins, and the dialkoxy derivatives. The Ni(II) complexes were all found to be severely ruffled whereas the free-base chromophores are more planar. As a result of the helimeric distortion of their porphyrinoid chromophores, the ruffled macrocycles possess a stable inherent element of chirality. Most significantly, resolution of the racemic mixtures was achieved, both by classical methods via diastereomers and by HPLC on a chiral phase. Full CD spectra were recorded and modeled using quantum-chemical computational methods, permitting, for the first time, an assignment of the absolute configurations of the chromophores. The report expands the range of known pyrrole-modified porphyrins. Beyond this, it introduces large chiral porphyrinoid π-systems that exist in the form of two enantiomeric, stereochemically stable helimers that can be resolved. This forms the basis for possible future applications, for example, in molecular-recognition systems or in materials with chiroptic properties.

Quinoline-annulated porphyrins.

Porphyrin-2,3-dione mono- and dioximes were used as starting materials for the efficient syntheses of mono- and bis-quinoline-annulated porphyrins and their corresponding N-oxides. Owing to an extended π-system, these novel porphyrinoid chromophores show significantly red-shifted UV-vis spectra compared to the parent porphyrins. A crystal structure exemplifies the nonplanar conformation of the macrocycle.

Unexpected hydroxylamine-induced ring-closure reactions of meso-tetraphenylsecochlorin bisaldehyde.

Reaction of meso-tetraphenylsecochlorin bisaldehyde with hydroxylamine results in the formation of the known meso-tetraphenyl-2-nitroporphyrin and the novel meso-tetraphenylimidazolophorphyrin. The products are the result of two diverging pathways of the presumed intermediate monooxime monoaldehyde that are unusual and surprising, but fully rationalized. The structures of both products were confirmed by spectroscopic techniques and single crystal X-ray diffractometry. This reaction represents the first reaction in which a pyrrole in a porphyrin was formally replaced by an imidazole moiety. The optical properties of the free base and metalloimidazoloporphyrin under neutral and acidic conditions are discussed. Further, an alternative synthesis of the imidazoloporphyrin Ni(II) based on meso-tetraphenyl-1-formyl-chlorophin is presented.

Enhanced electrochemical oxygen reduction-based glucose sensing using glucose oxidase on nanodendritic poly[meso-tetrakis(2-thienyl)porphyrinato]cobalt(II)-SWNTs composite electrodes.

The direct electrochemistry of glucose oxidase immobilized on a nanodendritic poly[meso-tetrakis(2-thienyl)porphyrinato]cobalt(II)-single walled carbon nanotube modified glassy carbon electrode (pCoTTP-SWNTs-Nafion-GOD/GCE) is reported. The immobilized GOD retained its activity and exhibited a surface controlled, reversible two-proton and two-electron transfer reaction with a fast heterogeneous electron transfer rate constant (ks) of 1.01 s(-1). The pCoTTP-SWNTs-Nafion matrix also showed an extremely low peak potential of -0.2 V vs. Ag/AgCl and strong response with respect to oxygen reduction. This forms the basis for the use of the pCoTTP-SWNTs-Nafion-GOD composite as a sensing platform for oxygen reduction-based glucose detection. The apparent Michaelis-Menten constant (Km,app) was estimated to be as low as 0.98 mM. A linear range up to 1mM glucose with a low detection limit of 5.33 µM (S/N=3) and a high sensitivity of 16.57 µA mM(-1) cm(-2) was achieved. The biosensor also shows excellent selectivity against 0.2 mM uric acid and ascorbic acid. These results indicate that the pCoTTP-SWNTs-Nafion-GOD/GCE has potential application in sensitive and selective glucose detection.

Oxazolochlorins. 2. Intramolecular cannizzaro reaction of meso-tetraphenylsecochlorin bisaldehyde.

Using mildly basic reaction conditions, the periodate-induced diol cleavage of meso-tetraphenyl-2,3-diolchlorin allows for the generation and isolation of the corresponding hitherto elusive free base secochlorin bisaldehyde. An intramolecular Cannizzaro reaction of this porphyrinoid generates three pyrrole-modified, oxazole-based porphyrins: the known porpholactol (2-oxa-3-hydroxychlorin) as the major product, known porpholactone (2-oxa-3-oxoporphyrin), and a novel porpholactol dimer that is linked through an acetal functionality. The structure of the dimer was confirmed by (1)H NMR spectroscopy, X-ray diffractometry, and ESI(+) collision-induced fragmentation mass spectrometry. The chromophores in the dimer are coupled electronically only to a minor extent. A mechanism to rationalize the formation of all products is advanced.