Antileishmanial Anthracene Endoperoxides: Efficacy In Vitro, Mechanisms and Structure-Activity Relationships.

Leishmaniasis is a vector-borne disease caused by protozoal Leishmania parasites. Previous studies have shown that endoperoxides (EP) can selectively kill Leishmania in host cells. Therefore, we studied in this work a set of new anthracene-derived EP (AcEP) together with their non-endoperoxidic analogs in model systems of Leishmania tarentolae promastigotes (LtP) and J774 macrophages for their antileishmanial activity and selectivity. The mechanism of effective compounds was explored by studying their reaction with iron (II) in chemical systems and in Leishmania. The correlation of structural parameters with activity demonstrated that in this compound set, active compounds had a LogPOW larger than 3.5 and a polar surface area smaller than 100 Å2. The most effective compounds (IC50 in LtP < 2 µM) with the highest selectivity (SI > 30) were pyridyl-/tert-butyl-substituted AcEP. Interestingly, also their analogs demonstrated activity and selectivity. In mechanistic studies, it was shown that EP were activated by iron in chemical systems and in LtP due to their EP group. However, the molecular structure beyond the EP group significantly contributed to their differential mitochondrial inhibition in Leishmania. The identified compound pairs are a good starting point for subsequent experiments in pathogenic Leishmania in vitro and in animal models.

Pyridinium Alkynylanthracenes as Sensitizers for Photodynamic Therapy.

Photodynamic therapy (PDT) is a mild but effective method to treat certain types of cancer upon irradiation with visible light. Here, three isomeric methylpyridinium alkynylanthracenes 1o─p were evaluated as sensitizers for PDT. Upon irradiation with blue or green light, all three compounds show the ability to initiate strand breaks of plasmid DNA. The mayor species responsible for cleavage is singlet oxygen (1 O2 ) as confirmed by scavenging reagents. Only isomers 1m and 1p can be incorporated into HeLa cells, whereas isomer 1o cannot permeate through the membrane. While isomer 1m targets the cell nucleus, isomer 1p assembles in the cellular cytoplasm and impacts the cellular integrity. This is in accordance with a moderate toxicity of 1p in the dark, whereas 1m exhibits no dark toxicity. Both isomers are suitable as PDT reagents, with a CC50 of 3 µm and 75 nm, for 1p and 1m, respectively. Thus, derivative 1m, which can be easily synthesized, becomes an interesting candidate for cancer therapy.

DNA-Triggered Enhancement of Singlet Oxygen Production by Pyridinium Alkynylanthracenes.

There is an ongoing interest in 1 O2 sensitizers, whose activity is selectively controlled by their interaction with DNA. To this end, we synthesized three isomeric pyridinium alkynylanthracenes 2 o-p and a water-soluble trapping reagent for 1 O2 . In water and in the absence of DNA, these dyes show a poor efficiency to sensitize the photooxygenation of the trapping reagent as they decompose due to electron transfer processes. In contrast, in the presence of DNA 1 O2 is generated from the excited DNA-bound ligand. The interactions of 2 o-p with DNA were investigated by thermal DNA melting studies, UV/vis and fluorescence spectroscopy, and linear and circular dichroism spectroscopy. Our studies revealed an intercalative binding with an orientation of the long pyridyl-alkynyl axis parallel to the main axis of the DNA base pairs. In the presence of poly(dA : dT), all three isomers show an enhanced formation of singlet oxygen, as indicated by the reaction of the latter with the trapping reagent. With green light irradiation of isomer 2 o in poly(dA : dT), the conversion rate of the trapping reagent is enhanced by a factor >10. The formation of 1 O2 was confirmed by control experiments under anaerobic conditions, in deuterated solvents, or by addition of 1 O2 quenchers. When bound to poly(dG : dC), the opposite effect was observed only for isomers 2 o and 2 m, namely the trapping reagent reacted significantly slower. Overall, we showed that pyridinium alkynylanthracenes are very useful intercalators, that exhibit an enhanced photochemical 1 O2 generation in the DNA-bound state.

Influence of Functional Groups on the Ene Reaction of Singlet Oxygen with 1,4-Cyclohexadienes.

The photooxygenation of 1,4-cyclohexadienes has been studied with a special focus on regio- and stereoselectivities. In all examples, only the methyl-substituted double bond undergoes an ene reaction with singlet oxygen, to afford hydroperoxides in moderate to good yields. We explain the high regioselectivities by a "large-group effect" of the adjacent quaternary stereocenter. Nitriles decrease the reactivity of singlet oxygen, presumably by quenching, but can stabilize proposed per-epoxide intermediates by polar interactions resulting in different stereoselectivities. Spiro lactams and lactones show an interesting effect on regio- and stereoselectivities of the ene reactions. Thus, singlet oxygen attacks the double bond preferentially anti to the carbonyl group, affording only one regioisomeric hydroperoxide. If the reaction occurs from the opposite face, the other regioisomer is exclusively formed by severe electrostatic repulsion in a perepoxide intermediate. We explain this unusual behavior by the fixed geometry of spiro compounds and call it a "spiro effect" in singlet oxygen ene reactions.

Stereoselective [4+2] Cycloaddition of Singlet Oxygen to Naphthalenes Controlled by Carbohydrates.

Stereoselective reactions of singlet oxygen are of current interest. Since enantioselective photooxygenations have not been realized efficiently, auxiliary control is an attractive alternative. However, the obtained peroxides are often too labile for isolation or further transformations into enantiomerically pure products. Herein, we describe the oxidation of naphthalenes by singlet oxygen, where the face selectivity is controlled by carbohydrates for the first time. The synthesis of the precursors is easily achieved starting from naphthoquinone and a protected glucose derivative in only two steps. Photooxygenations proceed smoothly at low temperature, and we detected the corresponding endoperoxides as sole products by NMR. They are labile and can thermally react back to the parent naphthalenes and singlet oxygen. However, we could isolate and characterize two enantiomerically pure peroxides, which are sufficiently stable at room temperature. An interesting influence of substituents on the stereoselectivities of the photooxygenations has been found, ranging from 51:49 to up to 91:9 dr (diastereomeric ratio). We explain this by a hindered rotation of the carbohydrate substituents, substantiated by a combination of NOESY measurements and theoretical calculations. Finally, we could transfer the chiral information from a pure endoperoxide to an epoxide, which was isolated after cleavage of the sugar chiral auxiliary in enantiomerically pure form.

pH-Sensitive Fluorescence Switching of Pyridylanthracenes: The Effect of the Isomeric Pattern.

9,10-substituted anthracenes are known for their useful optical properties like fluorescence, which makes them frequently used probes in sensing applications. In this article, we investigate the fundamental photophysical properties of three pyridyl-substituted variants. The nitrogen atoms in the pyridinium six-membered rings are located in the ortho-, meta-, and para-positions in relation to the anthracene core. Absorption, fluorescence, and transient absorption measurements were carried out and were complemented by theoretical calculations. We monitored the photophysics of the anthracene derivatives in chloroform and water investigating the protonated as well as their nonprotonated forms. We found that the optical properties of the nonprotonated forms are strongly determined by the anthracene chromophore, with only small differences to other 9,10-substituted anthracenes, for example diphenyl anthracene. In contrast, protonation leads to a strong decrease in fluorescence intensity and lifetime. Transient absorption measurements and theoretical calculations revealed the formation of a charge-transfer state in the protonated chromophores, where electron density is shifted from the anthracene moiety toward the protonated pyridyl substituents. While the para- and ortho-derivatives' charge transfer is still moderately fluorescent, the meta-derivative is affected much stronger and shows nearly no fluorescence. This nitrogen-atom-position-dependent sensitivity to hydronium activity makes a combination of these fluorophores very attractive for pH-sensing applications covering a broadened pH range.

Structural motives controlling the binding affinity of 9,10-bis(methylpyridinium)anthracenes towards DNA.

In the search of new DNA groove binding agents a series of substituted 9,10-methylpyridiniumanthracenes have been synthesized and their interactions with DNA have been studied by UV/vis absorption, CD and fluorescence spectroscopy. A minor groove binding mode is confirmed by DNA melting studies, strong CD effects, the dependence of the binding affinity on ionic strength, and the differentiation between AT and GC base pairs. No binding occurs to GC sequences. Binding constants to calf thymus DNA (ct-DNA) and poly(dA:dT) in the range between 1 × 104 and 3 × 105 M-1 have been determined. The binding strength decreases with the size of substituents attached at the anthracene site. Variation of the substitution pattern of the charged groups shows that methyl groups in meta position cause slightly stronger binding than methyl groups in para position. In contrast, with these groups in ortho position, no binding interaction has been observed. The strongest binding is achieved with an expansion of the peripheral heterocycle from pyridine to quinoline. Molecular modeling reveals the pivotal role of the substitution pattern: Anthracenes with para and meta pyridines align along the minor grooves. On the other hand, the ortho derivative adopts no groove-alignment.

Capture and Release of Singlet Oxygen in Coordination-Driven Self-Assembled Organoplatinum(II) Metallacycles.

Singlet oxygen (1O2), as an important active reagent, has found wide applications in photodynamic therapy (PDT), synthetic chemistry, and materials science. Organic conjugated aromatics serving as hosts to capture and release singlet oxygen have been systematically investigated over the last decades. Herein, we present a [6 + 6] organoplatinum(II) metallacycle by using ∼180° dipyridylanthracene donor and ∼120° Pt(II) acceptor as the building blocks, which enables the capture and release of singlet oxygen with relatively high photooxygenation and thermolysis rate constants. The photooxygenation of the metallacycle to the corresponding endoperoxide was performed by sensitized irradiation, and the resulting endoperoxide is stable at room temperature and can be stored under ambient condition over months. Upon simple heating of the neat endoperoxide under inert atmosphere at 120 °C for 4 h, the resulting endoperoxide can be reconverted to the corresponding parent form and singlet oxygen. The photooxygenation and thermolysis products were characterized by NMR spectroscopy and electrospray ionization time-of-flight mass spectrometric analysis. Density functional theory calculations were conducted in order to reveal the frontier molecular orbital interactions and reactivity. This work provides a new material platform for singlet oxygen related promising applications.

Release of Singlet Oxygen from Aromatic Endoperoxides by Chemical Triggers.

The generation of reactive singlet oxygen under mild conditions is of current interest in chemistry, biology, and medicine. We were able to release oxygen from dipyridylanthracene endoperoxides (EPOs) by using a simple chemical trigger at low temperature. Protonation and methylation of such EPOs strongly accelerated these reactions. Furthermore, the methyl pyridinium derivatives are water soluble and therefore serve as oxygen carriers in aqueous media. Methylation of the EPO of the ortho isomer affords the parent form directly without increasing the temperature under very mild conditions. This exceptional behavior is ascribed to the close contact between the nitrogen atom and the peroxo group. Singlet oxygen is released upon this reaction, and can be used to oxygenate an acceptor such as tetramethylethylene in the dark with no heating. Thus, a new chemical source of singlet oxygen has been found, which is triggered by a simple stimulus.

Activation of Anthracene Endoperoxides in Leishmania and Impairment of Mitochondrial Functions.

Leishmaniasis is a vector-borne disease caused by protozoal Leishmania. Because of resistance development against current drugs, new antileishmanial compounds are urgently needed. Endoperoxides (EPs) are successfully used in malaria therapy, and experimental evidence of their potential against leishmaniasis exists. Anthracene endoperoxides (AcEPs) have so far been only technically used and not explored for their leishmanicidal potential. This study verified the in vitro efficiency and mechanism of AcEPs against both Leishmania promastigotes and axenic amastigotes (L. tarentolae and L. donovani) as well as their toxicity in J774 macrophages. Additionally, the kinetics and radical products of AcEPs' reaction with iron, the formation of radicals by AcEPs in Leishmania, as well as the resulting impairment of parasite mitochondrial functions were studied. Using electron paramagnetic resonance combined with spin trapping, photometry, and fluorescence-based oximetry, AcEPs were demonstrated to (i) show antileishmanial activity in vitro at IC50 values in a low micromolar range, (ii) exhibit host cell toxicity in J774 macrophages, (iii) react rapidly with iron (II) resulting in the formation of oxygen- and carbon-centered radicals, (iv) produce carbon-centered radicals which could secondarily trigger superoxide radical formation in Leishmania, and (v) impair mitochondrial functions in Leishmania during parasite killing. Overall, the data of different AcEPs demonstrate that their structures besides the peroxo bridge strongly influence their activity and mechanism of their antileishmanial action.

Synthesis of Pyridylanthracenes and Their Reversible Reaction with Singlet Oxygen to Endoperoxides.

The ortho, meta, and para isomers of 9,10-dipyridylanthracene 1 have been synthesized and converted into their endoperoxides 1-O2 upon oxidation with singlet oxygen. The kinetics of this reaction can be controlled by the substitution pattern and the solvent: in highly polar solvents, the meta isomer is the most reactive, whereas the ortho isomer is oxidized fastest in nonpolar solvents. Heating of the endoperoxides affords the parent anthracenes by release of singlet oxygen.

Thiocarbohydrates on Gold Nanoparticles: Strong Influence of Stereocenters on Binding Affinity and Interparticle Forces.

Carbohydrates carrying thiol groups at the C-2 position have been attached to gold nanoparticles (AuNPs) with stereocenters in close proximity to the surface for the first time. Their configurations can be clearly distinguished by the tendency of particle aggregation. AuNP surface plasmon resonance (SPR), X-ray photoelectron spectroscopy (XPS), and IR spectroscopy indicate that the thiocarbohydrates replace citrate molecules at different rates, causing aggregation and eventually precipitation. A quantitative formulation of this aggregation process shows that reactivities can vary by several magnitudes. Adsorption isotherms and kinetics also demonstrate that the number of thiocarbohydrates varies by a factor of two. Molecular mechanics force field (MMFF) calculations reveal their relative orientations. Based on these models, the different binding behavior can be ascribed to attractive van der Waals forces and hydrogen bonds. Such interactions occur either between the carbohydrate and AuNPs, by lateral intermolecular forces at the surface, or by interparticle attraction, in analogy to cell-surface carbohydrates of biological recognition systems. Aggregation of NPs therefore act as an indicator to differentiate between various carbohydrates with defined configurations.

Role of Distance in Singlet Oxygen Applications: A Model System.

Herein, we present a model system that allows the investigation of a directed intramolecular singlet oxygen ((1)O2) transfer. Furthermore, we show the influence of singlet oxygen lifetime and diffusion coefficient (D) on the preference of the intramolecular reaction over the intermolecular one in competition experiments. Finally, we demonstrate the distance dependence in quenching experiments, which enables us to draw conclusions about the role of singlet oxygen and (1)O2 carriers in photodynamic therapy.

Why triple bonds protect acenes from oxidation and decomposition.

An experimental and computational study on the impact of functional groups on the oxidation stability of higher acenes is presented. We synthesized anthracenes, tetracenes, and pentacenes with various substituents at the periphery, identified their photooxygenation products, and measured the kinetics. Furthermore, the products obtained from thermolysis and the kinetics of the thermolysis are investigated. Density functional theory is applied in order to predict reaction energies, frontier molecular orbital interactions, and radical stabilization energies. The combined results allow us to describe the mechanisms of the oxidations and the subsequent thermolysis. We found that the alkynyl group not only enhances the oxidation stability of acenes but also protects the resulting endoperoxides from thermal decomposition. Additionally, such substituents increase the regioselectivity of the photooxygenation of tetracenes and pentacenes. For the first time, we oxidized alkynylpentacenes by using chemically generated singlet oxygen ((1)O(2)) without irradiation and identified a 6,13-endoperoxide as the sole regioisomer. The bimolecular rate constant of this oxidation amounts to only 1 × 10(5) s(-1) M(-1). This unexpectedly slow reaction is a result of a physical deactivation of (1)O(2). In contrast to unsubstituted or aryl-substituted acenes, photooxygenation of alkynyl-substituted acenes proceeds most likely by a concerted mechanism, while the thermolysis is well explained by the formation of radical intermediates. Our results should be important for the future design of oxidation stable acene-based semiconductors.

Novel anthracene materials for applications in lithography and reversible photoswitching by light and air.

Herein we demonstrate how the photoreaction between anthracenes and singlet oxygen ((1)O(2)) is employed for applications either as photoswitch or as photoresist. Thin films of the diaryl-alkyl anthracene 1 and the analogous oligomeric species 2 were irradiated under photomasks to generate pattern structures composed of 1/1-O(2) and 2/2-O(2). Kelvin probe force microscopy (KPFM) provided a powerful and nondestructive method to image the pattern information. The following studies based on AFM, KPFM and contact angle measurements unfold that the two species 1 and 2 underwent different progressions after the imaging step. Degrading is observed for the monomeric compound 1 and the pattern eventually becomes recognizable in topography. In the oxidized state (1-O(2)) the monomeric species remains physically stable. In consequence, the unreacted portion is removable and the remaining oxygenated form 1-O(2) is sufficiently stable to protect an underlying substrate (e.g., silver) from etching. Thus, the system 1/1-O(2) operates as photoresist. On the other hand, both states of the oligomer 2 remain stable. The film is stable up to temperatures >120 degrees C required to erase the pattern within acceptable time by cycloreversion. Anthracene 2 therefore acts as erasable and rewritable photochromic switch. The different behavior between 1 and 2 is explained by phase transitions which cause crystallization and finally ablation. Such transitions affect only the monomeric system 1/1-O(2) and not the oligomeric system 2/2-O(2). In conclusion, we designed two very similar materials based on diarylanthracenes, which can act either as a photoresist or as a rewritable photochromic switch.

Photoimaging with singlet oxygen at the solid-air interface.

Films of anthracene carboxylic acids were irradiated through photomasks and oxidized at the exposed regions by singlet oxygen upon sensitization. The efficiency of a photomask to protect the material underneath was investigated by optical and infrared spectroscopy. As the thickness of the film is reduced, the efficiency of the mask drops. This is explained by the migration of singlet oxygen at the solid-air interface, which in turn reacts at the masked area. For films with a thickness of <15 nm, the efficiency of the mask approaches zero; sufficient efficiency is achieved at thicknesses > 100 nm. From the investigations, it will become clear that the contrast between the irradiated and masked area of an image is affected by reduction of the film thickness. On the other hand, the resolution of an image, which relates to the minimum feature size of an image, is not dependent on the thickness of the film. The contributions of "inside" and "outside" reactions are examined separately, and a quantitative approximation of the spatial range of both modes of the oxygenation is given. We set up an approximate relation between mask efficiency and experimental conditions comprising internal and external oxygen diffusion, film thickness, and mask dimensions. These results give a deeper insight into the limits of resolution and contrast in singlet oxygen lithography.

9,10-diarylanthracenes as molecular switches: syntheses, properties, isomerisations and their reactions with singlet oxygen.

A series of 9,10-diarylanthracenes with various substituents at the ortho positions have been synthesised by palladium-catalysed cross-coupling reactions. Such compounds exhibit interesting physical properties and can be applied as molecular switches. Despite the high steric demand of the substituents, products were formed in moderate-to-good yields. In some cases, microwave conditions further improved yields. Bis-coupling afforded two isomers (syn and anti) that do not interconvert at room temperature. These products were easily separated and their relative stereochemistries were unequivocally assigned by NMR spectroscopy and X-ray analysis. The syn and anti isomers exhibit different physical properties (e.g., melting points and solubilities) and interconversion by rotation around the aryl-aryl axis commences at <100 degrees C for fluoro-substituted diarylanthracenes and at >300 degrees C for alkyl- or alkoxy-substituted diarylanthracenes. The reactions with singlet oxygen were studied separately and revealed different reactivities and reaction pathways. The yields and reactivities depend on the size and electronic nature of the substituents. The anti isomers form the same 9,10-endoperoxides as the syn species, occasionally accompanied by unexpected 1,4-endoperoxides as byproducts. Thermolysis of the endoperoxides exclusively yielded the syn isomers. The interesting rotation around the aryl-aryl axis allows the application of 9,10-diarylanthracenes as molecular switches, which are triggered by light and air under mild conditions. Finally, the oxygenation and thermolysis sequence provides a simple, synthetic access to a single stereoisomer (syn) from an unselective coupling step.

Remarkable oxidation stability of glycals: excellent substrates for cerium(IV)-mediated radical reactions.

The remarkable stability of glycals under oxidative conditions becomes apparent by their redox data in solution, computed HOMO energies, and behavior on the addition of electrophilic radicals generated in the presence of cerium(IV) ammonium nitrate. Oxidation potentials up to 2.03 V vs ferrocene were obtained, which are exceptionally high for cyclic enol ethers but correlate nicely with the reaction times of the radical reactions. Protecting groups have a strong influence on the oxidation stability and HOMO energies of glycals as E(ox) is shifted from O-silyl over O-benzyl to O-acetyl by more than 500 mV. Interestingly, this effect must be transmitted through sigma-bonds, even up to the para-position of a benzoate group, as verified by a wide variation of remote substituents in the carbohydrate. Favorable interactions of the sigma*-orbital of the adjacent C-O bond with the HOMO of the double bond are proposed as a mechanistic rationale, which might be important for the redox behavior of other allylic systems. Finally, donors and acceptors in the 1-position exert the strongest influence on the oxidation stability, shifting the potentials by almost 1 V and resulting in different follow-up reactions of the cerium(IV)-mediated additions of malonates. It is the remarkable oxidation stability of glycals that makes them valuable building blocks in carbohydrate chemistry.

Remote substituent effects on the photooxygenation of 9,10-diarylanthracenes: strong evidence for polar intermediates.

Two different reaction pathways in the photooxygenation of 9,10-diarylanthracenes are identified, with strong evidence for polar (forward, singlet oxygen addition) and radical (backward, thermolysis) intermediates.