Oxazine Dyes Research Articles

The role of microenvironments on computed vibrationally-resolved emission spectra: The case of oxazines.

Oxazine dyes act as reporters of their near environment by the response of their fluorescence spectra. At the same time, their fluorescence spectra exhibit a pronounced vibrational progression. In this work, we computationally investigate the impact of near-environment models consisting of aggregated water as well as betaine molecules on the vibrational profile of fluorescence spectra of different oxazine derivatives. For aggregated betaine and a water molecule located above the plane of the dyes, we observe a distinct modification of the vibrational profile, which is more pronounced than the effect of a continuum description of a solvent environment. Our analysis shows that this effect cannot be explained by a pure change in the electronic structure, but that also vibrational degrees of freedom of the environment can be decisive for the vibrational profile and should, hence, not generally be neglected.

TD‐DFT analysis of the excitation of H‐dimers of cationic dyes in an aqueous solution using functionals without additional dispersion correction

AbstractThis work is a development and extension of the previous one (DOI: 10.1039/d3cp00882g). Here, H‐dimers of acridine (acridine orange—AO and proflavine—PF), thiazine (methylene blue—MB and thionine—TH), and oxazine (brilliant cresyl blue—BCB and Nile blue—NB) dyes were modeled using hybrid functionals with a large proportion of exact Hartree–Fock exchange and long‐range correction. It turned out that nine functionals (LC‐ωHPBE, M06HF, M052X, M062X, M08HX, M11, MN15, SOGGA11X, and ωB97XD) reliably stabilize these molecular aggregates in both the ground and excited states. In addition, these functionals ensure that the conditions for transition moments (M(dimer) ≈ M(monomer) from strong coupling theory for H‐aggregates) and absorption maxima (λmax(dimer) < λmax(monomer) from Kasha exciton theory) are met. The S2 excited state stabilizes the H‐dimers more strongly than the ground state, while the S1 state stabilizes even more than S2. This is due to the large overlap between the corresponding molecular orbitals (LUMO > HOMO−1 > HOMO). When calculating the vibronic absorption spectra, the best agreement with the experiment for AO2, PF2, and NB2 showed the M08HX functional, and M11—for MB2 and BCB2. For dye monomers, these functionals gave the worst agreement, and MN15 demonstrated the closest similarity to the experiment. Vibronic absorption spectra for AO2, MB2, BCB2, and NB2 were calculated for the first time. The exciton splitting is calculated, which for MB2 is in good agreement with the experimental value.

Half-broadband two-dimensional electronic spectroscopy with active noise reduction

Two-dimensional electronic spectroscopy (2DES) provides detailed insight into coherent ultrafast molecular dynamics in the condensed phase. Here we report a referenced broadband pump-compressed continuum probe half-broadband (HB) 2DES spectrometer in a partially collinear geometry. To optimize signal-to-noise ratio (SNR) we implement active noise reduction referencing, which has not previously been applied in 2DES. The method is calibrated against the well characterized 2DES response of the oxazine dye cresyl violet and demonstrated at visible wavelengths on the photochromic photoswitch 1,2-Bis(2-methyl-5-phenyl-3-thienyl) perfluorocyclopentene (DAE). The SNR is improved by a factor of ∼2 through active referencing. This is illustrated in an application to resolve a low frequency mode in the excited electronic state of DAE, yielding new data on the reaction coordinate. We show that the active noise reduction referencing, coupled with the rapid data collection, allows the extraction of weak vibronic features, most notably a low frequency mode in the excited electronic state of DAE.

Cryogenic fluorescence spectroscopy of oxazine ions isolated in vacuo.

Recent developments in fluorescence spectroscopy have made it possible to measure both absorption and dispersed fluorescence spectra of isolated molecular ions at liquid-nitrogen temperatures. Absorption is here obtained from fluorescence-excitation experiments and does not rely on ion dissociation. One large advantage of reduced temperature compared to room-temperature spectroscopy is that spectra are narrow, and they provide information on vibronic features that can better be assigned from theoretical simulations. We report on the intrinsic spectroscopic properties of oxazine dyes cooled to about 100 K. They include six cations (crystal violet, darrow red, oxazine-1, oxazine-4, oxazine-170 and nile blue) and one anion (resorufin). Experiments were done with a home-built setup (LUNA2) where ions are stored, mass-selected, cooled, and photoexcited in a cylindrical ion trap. We find that the Stokes shifts are small (14-50 cm-1), which is ascribed to rigid geometries, that is, there are only small geometrical changes between the electronic ground and excited states. However, both the absorption and the emission spectra of darrow-red cations are broader than those of the other ionic dyes, which is likely associated with a less symmetric electronic structure and more non-zero Franck-Condon factors for the vibrational progressions. In the case of resorufin, the smallest ion under study, vibrational features are assigned based on calculated spectra.

Photoexcitation of Nile blue dye in aqueous solution: TD-DFT study

The vibronic absorption spectra of Nile blue (NB) oxazine dye in an aqueous solution using 13 hybrid functionals, the 6-31++G(d,p) basis set, and the IEFPCM solvent model were calculated. It turned out that the O3LYP functional provided the best agreement with the experiment. Various parameters of the NB cation in the ground and excited states (IR spectra, atomic charges, dipole moments, and transition moment) were obtained. Maps of the distribution of electron density and electrostatic potential have been built. The influence of four strong hydrogen bonds of the dye with water molecules on the absorption spectrum was analyzed. It has been shown that two from these bonds were strengthened upon NB excitation and two ones were weakened. It was found that explicit assignment of water molecules strongly bound to the dye leads to a redshift of the spectrum as a whole and worsened its shape.

Titrimetric Determination of Ascorbic Acid with Chloramine-T using Oxazine Dyes as lndicators

Titrimetric Determination of Ascorbic Acid with Chloramine-T using Oxazine Dyes as lndicators

In Situ Synthesis of Phenoxazine Dyes in Water: Application for “Turn‐On” Fluorogenic and Chromogenic Detection of Nitric Oxide**

AbstractThe synthesis of phenoxazine dyes was revisited in order to access these fluorescent N,O‐heterocycles under mild conditions. The combined sequential use of nitrosonium tetrafluoroborate (NOBF4) and triphenylphosphine enables the facile conversion of bis(3‐dimethylaminophenyl) ether into the methyl analogue of popular laser dye oxazine 1. The ability of nitrosonium cation (NO+) to initiate the domino reaction resulting in π‐conjugated phenoxazine molecules under neutral conditions, then led us to explore the feasibility of expanding it in aqueous media. Thus, we explored the use of reactive signaling molecule nitric oxide (NO) as a biological trigger of phenoxazine synthesis in water. The implementation of a robust analytical methodology based on fluorescence assays and HPLC‐fluorescence/‐MS analyses, have enabled us to demonstrate the viability of this novel fluorogenic reaction‐based process to selectively yield an intense “OFF‐ON” response in the near‐infrared (NIR‐I) spectral region. This study is an important step towards the popularization of the concept of “covalent assembly” in the fields of optical sensing, bioimaging and molecular theranostics.

Photoexcitation of brilliant cresyl blue dye in aqueous solution: TD-DFT study

The vibronic absorption spectra of brilliant cresyl blue (BCB) oxazine dye in an aqueous solution using 40 hybrid functionals, the 6-31++G(d,p) basis set, and the SMD solvent model were calculated. It turned out that the X3LYP functional provided the best agreement with the experiment in the positions of the main maximum and the short-wavelength shoulder. Our calculations showed that this shoulder is vibronic and is not caused by a separate electronic transition. At the same time, the shoulder intensity in the calculated spectrum turned out to be lower than in the experimental one. Various parameters of the BCB cation in the ground and excited states (IR spectra, atomic charges, dipole moments, and transition moment) were calculated. Maps of the distribution of electron density and electrostatic potential have been built. The influence of four strong hydrogen bonds of the dye with water molecules on the absorption spectrum was analyzed. It has been shown that these bonds are strengthened upon BCB excitation. It was found that explicit assignment of water molecules strongly bound to the dye leads to a redshift of the spectrum. Photoexcitation of the dye leads to a noticeable polarization of only one of the four considered water molecules.

Excited states of six oxazine 1 conformers in aqueous solution: TD-DFT/DFT study

The paper analyzes the electronic-vibrational states of six possible conformers of the dye oxazine 1 (also known as oxazine 725) in an aqueous solution using the SMD solvent model and the 6-31++G(d,p) basis set. Vibronic absorption spectra are calculated using a wide range of hybrid functionals. The X3LYP functional gives the best agreement with the experiment for the positions of the main maximum and shoulder. According to the calculations, the shoulder in the experimental spectrum is of vibronic origin. Various characteristics of the ground and excited states (Duschinsky matrices, IR spectra, atomic charges, dipole moments, maps of the distribution of the electrostatic potential) were obtained. All six investigated conformers have similar vibronic absorption spectra and frontier molecular orbitals. Significant differences between them are observed only for the set of vibronic transitions and relaxation from nonequilibrium to an equilibrium excited state. The effect of two strongly bound water molecules on the excitation of the dye was analyzed.

Gallocyanine as a Fluorogen for the Identification of NADPH-Dependent Production of Superoxide Anion Radical by Blood Cells

The interaction of the oxazine dye gallocyanine with reactive oxygen ( $$^{\centerdot }{\text{O}}_{2}^{ - },$$ Н2О2) and halogen (HOCl) species has been studied by spectrophotometry, mass spectrometry, and spectrofluorimetry. It has been shown that gallocyanine reacts with HOCl and $$^{\centerdot }{\text{O}}_{2}^{ - }$$ (but not with Н2О2) to form fluorescent products. It has been found using the inhibition assay that, both in the xanthine/xanthine oxidase system and in activated human blood neutrophils, the main contribution to the conversion of gallocyanine to a fluorophore comes from its reaction with $$^{\centerdot }{\text{O}}_{2}^{ - }{\text{.}}$$ The results obtained suggest that gallocyanine can act as a fluorogenic chemosensor and be used for estimating the activation of neutrophils and the NADPH-dependent production of superoxide anion radical by neutrophils and other blood cells, as well as for testing antioxidant drugs designed with the aim to correct diseases associated with oxidative stress.

Application of Celestine Blue B and Gallocyanine for Studying the Effect of Drugs on the Production of Reactive Oxygen and Halogen Species by Neutrophils

We investigated the effect of drugs (dapsone, paracetamol, and isoniazid) on the production of reactive oxygen and halogen species by neutrophils. The standard fluorescent method using scopoletin as well as recently developed fluorescent methods using oxazine dyes, celestine blue B, and gallocyanine were employed for this purpose. Celestine blue B reacts selectively with hypochlorous acid, while gallocyanine reacts mainly with the superoxide radical-anion, which reveals the regulatory effect of anti-inflammatory drugs on neutrophil NADPH-oxidase and myeloperoxidase responsible for the production of reactive oxygen and halogen species, respectively. These results indicate that gallocyanine and celestine blue B hold promise as chemosensors for studying the effect of drugs used in anti-inflammatory therapy for the neutrophil respiratory burst.

Ultrafast degradation of brilliant cresyl blue under hydrodynamic cavitation based advanced oxidation processes (AOPs)

The study highlights effectiveness of hydrodynamic cavitation (HC) in the degradation of effluents polluted by Brilliant Cresyl Blue (BCB) dye. Optimal process parameters were cavitation number 0.27, inlet pressure 1.70 bar, temperature 20 °C and pH 7. The efficiency of HC was investigated in combination with other advanced oxidation processes, including the addition of external oxidants (hydrogen peroxide, ozone, and sodium persulfate) and photooxidation. A detailed investigation of reactive radical species present in the system is also presented. Hybrid processes based on HC revealed the highest synergism. Finally, ultrafast degradation of target oxazine dye (decolorization efficiency of 100%) within the short treatment time of 1 min was developed. The most effective process was a combination of hydrodynamic cavitation and ozonation with a synergistic coefficient of 1.84 and electrical energy per order EEO of 0.03 kWh m−3 order−1. The presented solution is ready to be implemented as technology for industrial practice.

Probing of local polarity in poly(methyl methacrylate) with the charge transfer transition in Nile red.

The permittivity of polymers and its spatial distribution play a crucial role in the behavior of thin films, such as those used, e.g., as sensor coatings. In an attempt to develop a conclusive approach to determine these quantities, the polarity of the model polymer poly(methyl methacrylate) (PMMA) in 600 nm thin films on a glass support was probed by the energy of the charge transfer transition in the oxazine dye Nile red (NR) at 25 °C. The absorption and fluorescence spectra of NR were observed to shift to the red with increasing solvent polarity, because of the intramolecular charge transfer character of the optical transition. New types of solvatochromic plots of emission frequency against absorption frequency and vice versa afforded the Onsager radius-free estimation of the ground and excited states dipole moment ratio. With this approach the values of these dipole moments of 11.97 D and 18.30–19.16 D, respectively, were obtained for NR. An effective local dielectric constant of 5.9–8.3 for PMMA thin films was calculated from the solvatochromic plot and the fluorescence maximum of NR observed in the PMMA films. The fluorescence band of NR in the rigid PMMA films shifted to the red by 130 cm−1 with increasing excitation wavelength from 470 to 540 nm, while in a series of liquids the position of the emission maximum of NR remained constant within same range of the excitation wavelength. It is concluded that the fluorescence spectrum of NR in PMMA undergoes inhomogeneous broadening due to different surroundings of NR molecules in the ground state and slow sub-glass transition (Tg) relaxations in PMMA.

Plasmon-enhanced Förster energy transfer in Langmuir-Blodgett films based on organic dyes

AbstractThe effect of plasmon resonance of silver island films (SIF) on the interlayer Förster resonance energy transfer (FRET) between xanthene and oxazine dye molecules was studied. It has been shown that the enhancement of FRET can be controlled by changing in the distance between the donor-acceptor system and the SIF. The maximum increase in energy transfer efficiency (EET) by a factor of 2.6 was recorded at a distance of 6 nm from the SIF. The assumption was made that an increase in EET can be associated with both the direct appearance of a plasmon-enhanced rate constant of energy transfer and an increase in the quantum yield of the energy donor in direct contact with the SIF. The results can serve as a basis for studying of photoinduced processes in hybrid materials such as “organic dye-plasmon nanoparticles”, to increase the photosensitivity of solar cells in the visible region of the spectrum, and for the studying of photobiological processes, as well as to create materials with desired properties, sensors and light energy converters.

Resolving the Fluorescence Quenching Mechanism of an Oxazine Dye Using Ultrabroadband Two-Dimensional Electronic Spectroscopy.

The design and optimization of fluorescent labels and fluorogenic probes rely heavily on their ability to distinguish among multiple competing fluorescence quenching mechanisms. Cresyl violet, a member of the 1,4-oxazine family of dyes, has generally been regarded as an exemplary fluorescent probe; however, recent ultrafast experiments revealed an excited-state decay kinetic of 1.2 ps, suggesting the presence of a transient photochemical state. Here, we present ultrabroadband two-dimensional electronic spectroscopy (2D ES) measurements of cresyl violet in the presence of the fluorescence quenching agent 3,6-di(2-hydroxyethyl)-1,2,4,5-tetrazine. The broad spectral bandwidth allows for the evaluation of multiple fluorescence quenching mechanisms such as exciton formation, photoinduced electron transfer, resonance energy transfer, and excited-state proton transfer. The 2D electronic spectra in the presence and absence of the quencher suggest that excited-state proton transfer drives the system's excited-state dynamics and leads to a cresyl violet tautomer involved in fluorescence quenching. The invocation of the tautomeric form of cresyl violet neatly resolves longstanding inconsistencies in the photophysics of oxazine dyes more generally. Although still under development, the application of ultrabroadband 2D ES to a molecular system represents a compelling demonstration of the technique's future role in the study of photochemical reaction mechanisms.

Deuterium isotope effect in fluorescence of gaseous oxazine dyes.

The increased utility of fluorescence-based methods in recent years has highlighted the need for brighter, more efficient fluorophores. In order to design these fluorophores, an improved fundamental understanding is necessary of the structural components that intrinsically effect fluorescence efficiency. Here, we characterize the intrinsic effects of deuteration on fluorescence from gaseous oxazine dyes, without the influence of dye-solvent interactions, by making use of an ion trap mass spectrometer that has been altered to enable optical measurements. Comparison of emission spectra of four oxazine dyes: cresyl violet, oxazine 4, oxazine 170, and darrow red, show little change in profile upon deuteration of amine groups. However, deuteration significantly increases the efficiency of fluorescence with an increase in fluorescence lifetime and brightness by 10-23% for the gaseous dyes. This increase is less than half that of the quantum yield increase observed in deuterated solution. This indicates the large fluorescence efficiency changes for the oxazine dyes in deuterated solution result from a combination of both intrinsic effects as well as substantial contribution from altered fluorophore-solvent interactions. The intrinsic effects behind increased lifetime upon deuteration are explored using time-dependent density functional theory (TD-DFT) calculations of potential energy surfaces (PESs) for ground and low lying excited electronic states. In accord with experimental observations, calculated S1-S0 emission spectra show only minor differences between deuterated and non-deuterated forms indicating that the deuteration does not affect the radiative channel appreciably. Relaxed PES scans along the torsional motions of the amino groups reveal that the increase in lifetimes upon deuteration is likely due to quenching of different radiationless changes channels in different oxazine dyes. Calculations suggest that tunneling to access twisted intramolecular charge transfer states in S1 is critical in several of the oxazines. However, in at least one of the dyes examined, the large isotope effect is more likely due to differences in intersystem crossing rates. Overall, this combined experimental and computational investigation elucidates the photophysics of a well-known fluorescent scaffold and provides insight into how small differences can dramatically affect fluorescence outcomes.

Correlation between Photoelectrochemical and Spectrophotometric Study of Dye-Surfactant Combination in Photogalvanic Cell

The photogalvanic cells are rechargeable device with the sun light in which surfactant solutions are potentially important for efficient energy conversion and storage. Here, effect of cationic cetyltrimethyl ammonium bromide (CTAB), anionic sodium dodecyl sulphate (SDS) and non ionic tween 80 surfactants on the electrical output of the cationic oxazine dye brilliant cresyl blue (BCB)-fructose (redox couple) system in photogalvanic cells have been studied. For this, the photopotential and photocurrent for different cells having BCB-fructose system without and with surfactant (CTAB, SDS and tween 80) in the alkaline medium have been measured. The total 30 different cells have been prepared for optimizing the concentration of electrolytes corresponding to the higher electrical out put. Generally, the electrical output increases in presence of a particular surfactant, due to increase in solubilization and stabilization properties of dye molecules in the water. The amount of enhancement in electrical output of BCB-fructose system was highest with SDS and lowest with tween 80, i.e. the order for BCB-fructose system with different surfactants in photogalvanic cells is: BCB-fructose-SDS > BCB-fructose-CTAB > BCB-fructose-tween 80. This order for electrical output was good agreement with the stability order of BCB-SDS/CTAB/tween 80 system, on the basis of spectrophotometric study. Hence, among these surfactants, SDS has stronger capacity, (due to opposite charge on BCB and SDS) to stabilize the BCB-fructose system leads to enhancement in electrical output of photogalvanic cells. Therefore, dye and surfactant, having chemical structure, like BCB and SDS, has a great importance of improvement of electrical performance to photogalvanic cells in the future.

Luminescence spectroscopy of oxazine dye cations isolated in vacuo.

Here we report gas-phase action and luminescence spectra of cationic dyes derived from oxazine: cresyl violet (CV+), oxazine 170 (Ox-170+), nile blue (NB+), darrow red (DR+), oxazine 1 (Ox-1+), oxazine 4 (Ox-4+), and brilliant cresyl blue (BCB+). The first four have a benzofused structure, which results in asymmetric charge distributions along the long axis. The positive charge is also asymmetrically distributed in BCB+ while Ox-1+ and Ox-4+ are symmetric. As the ions are isolated in vacuo, there are no interactions with solvent molecules or counter ions, and the effect of chemical modifications is therefore more easily revealed than from solution-phase experiments. The transition energy decreases in the order: DR+ > CV+ > Ox-4+ > Ox-170+ > BCB+ > Ox-1+ > NB+, and the fluorescence from BCB+ is less than from the others. We discuss the results based on electron delocalisation, degree of charge-transfer character, rigidity of the chromophore structure, and substituents.

Synthesis and Application of Water-Soluble Oxazine Dyes for Detection of PHAs-Producing Bacteria.

Derivatives of oxazine dyes were synthesized on mulitigram scales via efficient synthetic strategies. One practical route was selected to prepare compounds 6, 9 and 10, especially water-soluble compound 6 was obtained in better yield than reported, and compound 10 was insoluble in aqueous media in absence of phenolic-OH. Compounds 3 and 9 were found to be clear pH-dependent between pH = 4.0 and 10.0, and could be used as acid-base indicators to measure intracellular pH. Compounds 6, 9, 10 all have carboxylic acid functionalities, which could be activated and used to conjugate the dyes to biomolecules. In addition, compounds 6 and 9 with good solubility in aqueous media were used to develop a simple, quick, safe, highly sensitive staining method to detect PHAs-producing bacteria on heat-fixed smears, which was confirmed by fluorescence images of PHAs granules of bacteria.

Silicon Substitution in Oxazine Dyes Yields Near-Infrared Azasiline Fluorophores That Absorb and Emit beyond 700 nm.

Exchanging the bridging oxygen atom in rhodamine dyes with a dimethylsilyl group red-shifts their excitation and emission spectra, transforming orange fluorescent rhodamines into far-red Si-rhodamines. To study the effect of this substitution in other dye scaffolds, synthetic approaches to incorporate silicon into the bridging position of oxazine dyes were developed. The fluorescence of the compact azasiline dyes ASiFluor710 and ASiFluor730 is red-shifted by 57-83 nm from that of Oxazine 1.