Characterization of local polarity and structure of Cys121 domain in beta-lactoglobulin with a new thiol-specific fluorescent probe

The design and synthesis of a new polarity-sensitive fluorescent probe, 3-(4-chloro-6-p-maleimidylphenoxyl-1,3,5-triazinylamino)-7-dimethylamino-2-methylphenazine, is reported for characterizing the local polarity and structure, such as the thiol domain, of a protein. The probe comprises a polarity-sensitive fluorophore (neutral red moiety) and a thiol-specific labeling group (maleimidyl moiety). The probe exhibits a sensitive response of shift of fluorescence maximum emission wavelength to solvent polarity, but not to pH and temperature, which makes the probe suitable for determining the local polarity change of a protein denatured by pH or temperature. The application of this kind has been first demonstrated for the polarity detection of the Cys121 domain of beta-lactoglobulin. It is found that the polarity of the Cys121 domain corresponds to a dielectric constant of 17.3, and this value hardly alters after heat treatment, which may be attributed to the improved thermal reversibility by the Cys121 modification. The simple mixture of the probe and the protein at pH 5.6 is also studied, revealing that the free probe prefers to bind to an outer hydrophobic region. Heat treatment of the mixture causes the modification of Cys121 residues; this modification does not completely destroy the calyx but results in the opening of a channel for the probe to enter the calyx of beta-lactoglobulin. These results show that Cys121 plays an important role not only in the thermal reversibility but also in the accessibility of the calyx to a ligand. The strategy presented here further indicates that the combination of polarity-sensitive fluorescence probe with site-specific labeling may serve as a powerful means for elucidating structures and properties of proteins.     

Probing the Polarity of Sol-Gels and Ormosils via the Absorption of Nile Red

Conventional sol-gels are rather hydrophilic. A more hydrophobic material is obtained preparing organically modified siloxanes (ormosils). The polarity-sensitive probe Nile Red (NR) was doped in various sol-gels to probe their micro-polarity. The experiments show that the NR is an excellent probe and sensitive to the polarity of its microenvironment. Spectroscopic studies reveal remarkable changes in the absorption band positions and intensities as a function of the polarity of the sol-gel, which depends on the different precursors used. Furthermore, sol aging, gelation and temporal stability as a function of different ormosils have been investigated.     

The styrylpyridine dye for the silane sol-gel transition studies by time-dependent fluorescence

A trans-4-(p-N,N-dimethylaminostyryl)-N-vinylbenzylpyridinium chloride (vbDMASP) fluorescence probe was optimized in ground and excited state as a function of change in the microenvironment polarity, using the Amsol HyperChem program package. In the calculations, protic and aprotic solvents were used. On this basis a change in the molecule geometry after excitation, depending on the surrounding solvent, was determined. Absorption and steady-state fluorescence spectra of vbDMASP in the solvent of different polarity and in the model water-glycerol solutions were also recorded. On the basis of Stokes' shift change with the Onsager polarity scale a change in the dipole moment of the probe during transition from ground to excited state, in protic and aprotic solvents was determined. Since during the sol-gel transition of tetraethylorthosilane in the acidic environment both polarity and viscosity of the microenvironment change the vbDMASP probe was applied and fluorescence time-resolved measurements were done. On this basis the correlations between the results of time-resolved measurements for the multichromophoric probe applied in the gelation process and molecular optimization data are discussed.     

The styrylpyridine dye for the silane sol–gel transition studies by time-dependent fluorescence

A trans-4-(p-N,N-dimethylaminostyryl)-N-vinylbenzylpyridinium chloride (vbDMASP) fluorescence probe was optimized in ground and excited state as a function of change in the microenvironment polarity, using the Amsol HyperChem program package. In the calculations, protic and aprotic solvents were used. On this basis a change in the molecule geometry after excitation, depending on the surrounding solvent, was determined.Absorption and steady-state fluorescence spectra of vbDMASP in the solvent of different polarity and in the model water–glycerol solutions were also recorded. On the basis of Stokes’ shift change with the Onsager polarity scale a change in the dipole moment of the probe during transition from ground to excited state, in protic and aprotic solvents was determined.Since during the sol–gel transition of tetraethylorthosilane in the acidic environment both polarity and viscosity of the microenvironment change the vbDMASP probe was applied and fluorescence time-resolved measurements were done. On this basis the correlations between the results of time-resolved measurements for the multichromophoric probe applied in the gelation process and molecular optimization data are discussed.     

Unusual fluorescence spectral response of 1-(4-N,N-dimethylaminophenylethynyl)pyrene towards the thermotropic phase change in lipid bilayer membranes

The applicability of 1-(4-N,N-dimethylaminophenylethynyl)pyrene (DMAPEPy), a pyrene derivative showing intramolecular charge transfer, as a prospective probe for lipid bilayer membranes has been evaluated. High sensitivity of DMAPEPy to solvent polarity and viscosity makes it to act both as a polarity-sensitive probe and as a fluorescence anisotropy probe. The molecule shows high partition efficiency towards bilayer membranes in both solid gel as well as in the liquid crystalline phases. The emission spectrum, quenching experiment and lifetime data suggest bimodal distribution of DMAPEPy in the bilayer. Using the solvent polarity scales the polarity parameters of the two locations in lipid bilayer have been estimated. In the bilayer environment it exhibits remarkable spectral changes with temperature. The thermotropic phase change of the bilayer is sensitively monitored by fluorescence intensity as well as fluorescence anisotropy parameters. DMAPEPy is also capable of sensing the changes induced by membrane modifiers like cholesterol.     

Ratiometric Fluorescence Imaging of Cellular Polarity: Decrease in Mitochondrial Polarity in Cancer Cells

Mitochondrial polarity strongly influences the intracellular transportation of proteins and interactions between biomacromolecules. The first fluorescent probe capable of the ratiometric imaging of mitochondrial polarity is reported. The probe, termed BOB, has two absorption maxima (λ_abs=426 and 561 nm) and two emission maxima—a strong green emission (λ_em=467 nm) and a weak red emission (642 nm in methanol)—when excited at 405 nm. However, only the green emission is markedly sensitive to polarity changes, thus providing a ratiometric fluorescence response with a good linear relationship in both extensive and narrow ranges of solution polarity. BOB possesses high specificity to mitochondria (R_r=0.96) that is independent of the mitochondrial membrane potential. The mitochondrial polarity in cancer cells was found to be lower than that of normal cells by ratiometric fluorescence imaging with BOB. The difference in mitochondrial polarity might be used to distinguish cancer cells from normal cells.     

Electron spin resonance investigation of microscopic viscosity, ordering, and polarity in Nafion membranes containing methanol-water mixtures

Electron spin resonance (ESR) was used to monitor the local environment of 2,2,6,6-tetramethyl-4-piperidone N-oxide (Tempone) spin probe in water and methanol mixtures in solution and in Li(+) ion exchanged Nafion 117 membranes. Solution spectra were analyzed using the standard fast-motion line width parameters, while membrane spectra were fitted using the microscopic order macroscopic disorder (MOMD) slow-motional line shape program of Freed and co-workers. The (14)N hyperfine splitting, aN, which reflects the local polarity of the nitroxide probe, decreases with increasing methanol concentration, consistent with the decrease in solvent polarity. The polarity depended only weakly on composition in the Nafion membrane, but was noticeably more temperature-dependent. The microviscosity of the membrane aqueous phase as reflected by the rotational correlation time (tauc) of the probe, was nearly 2 orders of magnitude longer in the membrane than in solution and varied by an order of magnitude over the composition range studied. The probe exhibits significant local ordering in the aqueous phase of Nafion membranes that is diminished with increasing methanol concentration.     

Markov-type evolution of materials into a polar state

Assembling polar building blocks into a solid material by a Markov-chain process of unidirectional growth principally results in a metastable state that shows effects of macroscopic polarity. Stochastic polarity formation can be described by probabilities for the attachment of building blocks to a surface. Because of the polar symmetry of the building blocks, there is a fundamental difference in the probabilities for attaching them "tip-first" or "back-first" to growth sites at a surface. A difference in the corresponding probabilities drives the evolution of a vectorial property through a gain in configurational entropy. Examples from the mechanical, the crystalline and the biological world demonstrate growth-induced macroscopic polarity. In crystals, growth upon centrosymmetric seeds can produce twinned crystals with a "sectorwise" pyroelectric effect. Polarity formation in connective tissues is explained by a Markov-chain mechanism, which drives the self-assembly of collagen fibril segments. An unified stochastic growth model brings up a general concept for the formation of materials with polar properties.     

Diverse Role of Solvents in Controlling Supramolecular Chirality

Supramolecular self-assembly stands for the spontaneous aggregation of small organic compounds or polymers into ordered structures at any scale. When being induced by inherent molecular chiral centers or ambient asymmetric factors, asymmetric spatial arrangement between building units shall occur, which is defined as supramolecular chirality. Except for molecular design, utilizing external stimulus factors to tune supramolecular chirality is a promising approach. In this Concept article, we particularly discuss the important role of solvents in manipulating the chirality of self-assembled systems. The impact of solvents on the chirality is generally based on three properties of solvents, i.e., chirality, polarity, and active coassembly with building blocks. Molecular self-assembly in chiral solvents could undergo the chirality transfer, exhibiting a chiral induction effect. Solvent polarity often determines intermolecular orientation. As a consequence, those building blocks with both polar and apolar segments might change their chirality depending on the solvent polarity. We elaborate the active participation of solvent molecules into ordered structures together with building blocks, where solvents and building blocks exhibit a coassembly manner. By specific treatments such as heating and cooling, solvents could be released or re-entrapped, allowing a smart control over supramolecular chirality. The solvent effect in manipulating two-dimensional chiral self-assemblies is then discussed. The perspective and future development in this research field are presented at last.     

A lysosomal polarity-specific two-photon fluorescent probe for visualization of autophagy

Autophagy plays a vital role in maintaining the balance of normal physiological state of living cells. In this paper, a polarity-specific two-photon fluorescent probe Lyso-NA based on naphthalimide was synthesized for the purpose of monitoring autophagy during biological research. The results of photophysical properties and theoretical calculation confirmed that different polarities of solvents mainly effected fluorescent intensities of probe. Fluorescent intensity, quantum yield and fluorescence lifetime of probe kept a good linear relationship with polarity respectively. In addition, due to its low toxicity and selective accumulation in lysosomes, Lyso-NA is suitable for detecting changes in lysosomal polarity of living cells. Compare with the imaging results of plasmid transfection, a better performed real-time long-term fluorescent visualization of autophagy in living cells was achieved. Probe Lyso-NA can work as an efficient and cost effective imaging tool for visualizing autophagy in living cells.     

Polarity of layer-by-layer deposited polyelectrolyte films as determined by pyrene fluorescence.

The polarity of polyelectrolyte (PE) multilayer films is investigated with pyrene as a polarity-sensitive probe. Multilayer films of poly(styrene sulfonate) (PSS) and various polycations were prepared by the layer-by-layer self-assembly technique. Pyrene (PY) molecules were inserted into the films by exposing the multilayers to pyrene solutions. By this method a homogeneous distribution of pyrene molecules at low concentration within the film was obtained. The ratio of the fluorescence intensities of the first (I) to the third (III) vibronic band (Py-value) of the pyrene emission spectrum is employed here to determine the polarity of the PE films. PSS and poly(allylamine hydrochloride) (PAH) multilayer films yielded a pyrene value close to the solvent polarity of acetone, while multilayers of PSS and poly(diallyldimethylammonium chloride) (PDADMAC) displayed a value higher than the one corresponding to water. The pyrene values of the polyelectrolyte films were independent from the solvent employed for probe dissolving. Although no direct relationship between solvent polarity and dielectric constant (epsilon) is available, an estimate of the static dielectric constant of the films can be provided by comparing the Py-values of the films with those of various solvents. Changes in the humidity conditions of the film environment in a closed cell did not affect the film polarity. However, a drastic and irreversible reduction of polarity could be induced by actively drying the samples by a nitrogen flow.     

Determination of solvatochromic solvent polarity parameters for the characterisation of some nematic liquid crystal in anisotropic and isotropic phases

Characterisation of liquid crystals (LCs) as solvents is needed, to obtain the polarity and solvatochromic polarity parameters of these media. Polarity parameters demonstrate the effects of LC media on the photo-physical behaviour of solute molecules in an anisotropic medium. The practical limitations in determining solvent polarity scale parameters for LCs can overcome the overlapping absorption band of LCs and solvent-sensitive standard compounds or their insolubility in LCs. In this work, we report Kamlet–Abboud–Taft polarity functions of some nematic LCs in different temperatures and phases, isotropic and anisotropic, with the solvatochromic method, using the Reichardt's dye and 2,6-diphenyl-4-(2,4,6-triphenyl-1-pyridinio)-phenolate standard probe. In addition, a new azo and coumarin dye were used as probes to obtain some solvatochromic polarity parameters. Finally, a new polarity parameter, the LC anisotropic matrix, is introduced.     

光诱导电子转移与电荷分离——N-[(1-芘基)-亚甲基]-N-甲基苯胺与N-[(1-芘基)-亚乙基]-N-甲基苯胺光物理性质的比较

设计合成了二个含芘和苯胺基的二元化合物,研究了溶剂极性和粘度对它们分子内激基复合物的影响。此外,CTAB胶束中的光物理性质表明它们可以作为微环境的粘度探针和极性探针。     

结构受阻的取代的苯乙烯基吡嗪衍生物作为荧光探针的研究

本工作合成了两种旋转受阻取代的苯乙烯基吡嗪衍生物。详细研究了环境温度、极性和粘度对其光物理及发光行为的影响。结果表明两化合物在基态时可能存在着"平面构象与扭曲构象"间的平衡, 环境的温度、极性和粘度对于化合物存在的构象有很大影响。扭曲构象为较好的发光构象。分子受激后, 平面构象可经旋转松弛到扭曲构象。两化合物的荧光量子产率在室温条件下基本不随溶剂极性变化而变, 仅与溶剂粘度有关, 利用该性质可建议其作为一种有效的粘度荧光探针。     

Determination of polarity parameters for liquid crystals using solvatochromic method in anisotropic and isotropic phases

The use of liquid crystals (LCs) as anisotropic solvents is desired for various potential applications and usually for other organic and inorganic compounds. In this work, solvent polarity parameters are obtained using a spectroscopic method for four LCs with a range of high and low dielectric anisotropy (?ε). Solvatochromic polarity parameters for these LCs were defined via Kamlet–Abboud–Taft polarity functions characterizing different temperatures and phases, isotropic and anisotropic, and using the Reichardt’s dye and 2,6-diphenyl-4-(2,4,6-triphenyl-1-pyridinio) phenolate standard probe. The investigated polarity parameters reveal the effects of LC media on the photo-physical behaviour of solute molecules in isotropic and anisotropic media. Subsequently, a new LC polarity parameter (Z_o) is introduced as an overall matrix anisotropy polarity parameter to characterize variation between isotropic and anisotropic phases. The values of Z_o are sorted from higher to lower dielectric anisotropies (?ε).     

Recent progress in studies on polarity of ionic liquids

Although many ionic liquids have been reported, their polarity is not completely understood. Different empirical polarity scales for molecular solvents always lead to different polarity orders when they are applied on ionic liquids. Based on a literature survey, this review summarizes the recent polarity scales of ionic liquids according to the following 4 classes: (1) equilibrium and kinetic rate constants of chemical reactions; (2) empirical polar parameters of ionic liquids; (3) spectral properties of probe molecules; (4) multiparameter approaches. In addition, their interrelations are presented. A systematic understanding of the relationship between different polarity parameters of ionic liquids is of great importance for finding a universal set of parameters that can be used to predict the polarities of ionic liquids quantitatively. The potential utilization of the electron paramagnetic resonance in this field is also addressed.     

荧光探针法研究新型Gemini表面活性剂在水溶液中的聚集行为

以芘为荧光探针、二苯酮为猝灭剂,用稳态荧光探针法测定了新型Gemini表面活性剂的临界胶团浓度(CMC)、胶团聚集数(N_agg)及胶团微极性.研究了Gemini表面活性剂结构和氯化钠浓度对CMC、N_agg、胶团微极性的影响.结果表明,新型Gemini表面活性剂的CMC比常规表面活性剂的CMC低1—2个数量级.当疏水基碳原子数增加时,CMC依次降低,N_agg增大,胶团微极性减小.当氯化钠浓度增大时,N_agg增大,胶团微极性减小.     

利用荧光探针溶致变色效应研究有机改性蒙脱土的层间环境性质

利用荧光探针的溶致变色行为,研究了以不同阳离子表面活性剂处理过的化学修饰蒙脱土内腔的极性.清楚地观察到随所用表面活性剂长链碳原子数目的增多,荧光探针N,N-二甲氨基查尔酮在蒙脱土内的荧光光谱峰值波长移向短波,表明蒙脱土内腔的极性随活性剂疏水链长的增长而不断减小.本工作所得结果将对具体的插层材料(包括不同极性的聚合单体或聚合物分子)在插层时选择合适的表面处理剂有所帮助.     

A ratiometric fluorescent probe based on carbon dots assembly for intracellular lysosomal polarity imaging with wide range response

Lysosomal polarity is considered a key indicator of lysosomal function due to its significant impact on membrane fluidity and enzymatic reactions in lysosomes. Monitoring lysosomal polarity can gain insight into the related physiological and pathological processes and develop new diagnostic methods. However, current fluorescent probes with lysosomal polarity response suffer from narrow linear range, photobleaching and complicated preparation. Herein, a ratiometric fluorescent probe(r-b CDs) for ...     

A bright two-photon fluorescent probe for real-time monitoring autophagy in living cells

A novel donor-acceptor(D-A) type of two-photon(TP) fluorescent probe,i.e.Lyso-OSC,based on the lysosome-targeting morpholine group was developed.The polarity sensing coumarin group was functionalized as the acceptor and the 1-vinyl-4-methoxybenzene group was engineered as the donor.The fluorescence intensity and emission maximum wavelength of Lyso-OSC are highly sensitive to the polarity changes of solvent.The two-photon absorption cross-section and tissue penetration depth are up to 254 GM and 150 μm,respectively.The strong fluorescence,high sensitivity to polarity,low cytotoxicity,and accurate lysosome-targeting ability entail Lyso-OSC the excellent performance in detecting the polarity changes ofcellular environment.To this end,a bright,real-time imaging autophagy of living cells has been achieved.