Reactive oxygen species in photochemistry of the red fluorescent protein "Killer Red"

The fluorescent protein aptly named "Killer Red" (KRed) is capable of killing transfected cells and inactivating fused proteins upon exposure to visible light in the presence of oxygen. We have investigated the source of the bioactive species through a variety of photophysical and photochemical techniques. Our results indicate a Type I (electron transfer mediated) photosensitizing mechanism.     

A general synthetic strategy for the design of new BODIPY fluorophores based on pyrroles with polycondensed aromatic and metallocene substituents

BODIPYrrole: A general strategy for the design of novel BODIPY fluorophores based on pyrroles with polycondensed aromatic and metallocene substituents has been developed. The strategy involves the acylation of the condensed substituent and treatment of the acylated derivative (as oxime) with acetylene in MOH/DMSO (M = alkali metal) to give pyrroles that were then used for assembly of the BODIPY fluorophores (see scheme).     

Application of a modified Pechini method for the synthesis of Ln<Subscript>2</Subscript>MGe<Subscript>4</Subscript>O<Subscript>12</Subscript> (Ln = Y,Eu; M = Ca,Zn, Mn) optical hosts

A modified Pechini method followed by conventional and microwave heating was used to synthesise the new promising Ln₂MGe₄O₁₂ (Ln = Y, Eu; M = Ca, Zn, Mn) optical hosts. Comparison between solid-state and Pechini synthesis methods showed that the latter reduces the temperature required for cyclo-tetragermanate formation. The highest yield of cyclo-tetragermanates for both methods is observed at 1,000–1,100 °C, with significantly shorter time of annealing in the case of the Pechini synthesis. Compositional, structural and morphological characterisations of the samples obtained by both routes were carried out using X-Ray powder diffraction, thermogravimetry, electron spin resonance spectroscopy, energy-dispersive X-Ray spectroscopy and scanning electron microscopy.     

Nanodimer cyclodextrin ligands with high affinity to steroids

Molecular-imprinting by cross-linking of ligands of ??-cyclodextrin (CD) complex with steroids has been developed for the synthesis of tailor-made CD dimer. Steroids of androstane (9??-hydroxy-androst-4-en-3,17-dione, androst-4-en-3,17-dione, androsta-1,4-dien-3,17-dione (ADD)) and pregnane (hydrocortisone, 6-methyl-hydrocortisone, 20-hydroxymethylpregna-1,4-diene-3-one (HMPD)) series were used as template molecules. For imprinting procedure, crystalline ??-CD complexes of exact stoichiometry (??-CD:steroid template = 2:1) were synthesized following by toluene 2,4-diisocyanate (TDI) cross-linking. The attempts to produce CD dimer for steroid without hydrophobic side chain failed, while tailor-made CD dimer has been obtained using HMPD as a template. The dimer was characterized by ¹H NMR and mass-spectrometry. The complex stability constant (K_S) towards HMPD template exceeded 10⁷ M^?1. The K_S of CD dimer with ADD exceeded the corresponded value of TDI-modified CD monomer by more than an order of magnitude. The dimer was applied for quantitative extraction of ADD from aqueous solution using dialysis membranes impermeable for CD. The value of K_S for ADD estimated from balanced concentrations of dialysis data corresponded to that calculated by nonlinear spectrometric method.     

Influence of cholesterol on the phase transition of lipid bilayers: a temperature-controlled force spectroscopy study

Cholesterol (Chol) plays the essential function of regulating the physical properties of the cell membrane by controlling the lipid organization and phase behavior and, thus, managing the membrane fluidity and its mechanical strength. Here, we explore the model system DPPC:Chol by means of temperature-controlled atomic force microscopy (AFM) imaging and AFM-based force spectroscopy (AFM-FS) to assess the influence of Chol on the membrane ordering and stability. We analyze the system in a representative range of compositions up to 50 mol % Chol studying the phase evolution upon temperature increase (from room temperature to temperatures high above the T(m) of the DPPC bilayer) and the corresponding (nano)mechanical stability. By this means, we correlate the mechanical behavior and composition with the lateral order of each phase present in the bilayers. We prove that low Chol contents lead to a phase-segregated system, whereas high contents of Chol can give a homogeneous bilayer. In both cases, Chol enhances the mechanical stability of the membrane, and an extraordinarily stable system is observed for equimolar fractions (50 mol % Chol). In addition, even when no thermal transition is detected by the traditional bulk analysis techniques for liposomes with high Chol content (40 and 50 mol %), we demonstrate that temperature-controlled AFM-FS is capable of identifying a thermal transition for the supported lipid bilayers. Finally, our results validate the AFM-FS technique as an ideal platform to differentiate phase coexistence and transitions in lipid bilayers and bridge the gap between the results obtained by traditional methods for bulk analysis, the theoretical predictions, and the behavior of these systems at the nanoscale.     

Excited-state dynamics of organic dyes at liquid/liquid interfaces

Liquid/liquid interfaces play a crucial role in numerous areas of science. However, direct spectroscopic access to this thin (～1 nm) region is not possible with conventional optical methods. After a brief review of the most used techniques to perform interfacial optical spectroscopy, we will focus on time-resolved surface second harmonic generation, which allows the measurement of the excited-state dynamics of probe molecules at interfaces. By comparing these dynamics with those measured in bulk solutions, precious information on the properties of the interfacial region can be obtained. To illustrate this, several studies performed in our group will be presented.     

From Graftable Biphotonic Chromophores to Water‐Soluble Organic Nanodots for Biophotonics: The Importance of Environmental Effects

The photophysical and two‐photon absorption (TPA) properties of biphotonic chromophores with one or two phenol pendant units were studied and compared with that of a model biphotonic quadrupolar chromophore. A water‐soluble dendritic structure was then synthesized by using the pendant moieties as starting points for the construction of dendritic branches. We show that the polarity of the environment significantly modulates both the fluorescence and the TPA responses of the different chromophoric derivatives. This extends to more subtle effects that involve phenol pendant moieties that were found to act as discrete solvating units and to modify both the photophysics and the TPA response of the chromophore. This demonstrates the high sensitivity of the TPA response of quadrupolar derivatives to minute alterations in the environment. Moreover, the dendritic branches were found to behave as a peculiar cybotactic environment that was able to tune the fluorescence and TPA response of the inner chromophore by creating a polar environment. This reveals a new direction for exploiting such effects by playing on the dendritic architecture (e.g., the nature and shape of the building blocks, the geometry and position of the chromophore) to modulate the TPA responses.     

Reactions of sulfur- and phosphorus-substituted fluoroalkylating silicon reagents with imines and enamines under acidic conditions

Nucleophilic fluoroalkylation reactions of imines and enamines with α-phenylthio, α-phenylsulfonyl, and α-diethylphosphoryl substituted fluorinated silanes have been investigated. The reactions are promoted by hydrofluoric acid generated in situ from potassium hydrodifluoride and trifluoroacetic acid. Sulfur reagents worked well with both imines and enamines, whereas phosphorus reagent efficiently coupled only with enamines.     

Difluoro(trimethylsilyl)acetonitrile: synthesis and fluoroalkylation reactions

A new silicon reagent, difluoro(trimethylsilyl)acetonitrile, was prepared by insertion of difluorocarbene into silyl cyanide. The obtained silane served as a good cyanodifluoromethylating reagent toward aldehydes, N-tosylimines, N-alkylimines, and enamines under basic or acidic conditions.     

What factors control product yield in charge separation reaction from second excited state in zinc-porphyrin derivatives?

Intramolecular charge separation from the second singlet excited state of directly linked Zn-porphyrin-imide dyads and following charge recombination into the first singlet excited state has been investigated in the framework of a model involving three electronic states (the first and the second singlet excited and charge separated states) as well as their vibrational sublevels. Kinetics of the transitions between these states are described in terms of the stochastic point-transition approach. The influence of the model parameters (free energy change of charge separation, magnitude of the reorganization energies of the medium and the high frequency intramolecular vibrations, the rate of relaxation of the medium and the intramolecular high frequency vibrational mode) on the kinetics of population of both the charge separated and the first singlet excited states has been explored. Simulations of the kinetics of the charge separated state population have allowed reproducing the distinctive features of the kinetics observed in the experiment [Wallin, S.; Monnereau, C.; Blart, E.; Gankou, J.-R.; Odobel, F.; Hammarstr?m, L. J. Phys. Chem. A 2010, 114, 1709]: (i) two maxima on short time scale (hundreds of femtoseconds) and long time scale (tens of picoseconds), (ii) the magnitudes of both maxima, and (iii) the depth of the notch between the maxima.