Fluorescence Emission Anisotropy Research Articles

Influence of archaeal lipids isolated from Aeropyrum pernix K1 on physicochemical properties of sphingomyelin-cholesterol liposomes

We investigated the influence of archaeal lipids (C25,25) isolated from thermophilic archaeon Aeropyrum pernix K1 on physicochemical properties of liposomes comprised of egg sphingomyelin (SM) and cholesterol (CH) using fluorescence emission anisotropy, calcein release studies, dynamic light scattering, transmissive electron microscopy and phase analysis light scattering. The 2 mol% addition of archaeal lipids enabled formation of small unilamellar vesicles by sonication while also having significant effect on reducing mean size, polydispersity index and zeta potential of C25,25/SM/CH vesicles. Increasing the ratio of C25,25 lipids in mixture of C25,25/SM/CH decreased lipid ordering parameter in dose dependent manner at different temperatures. We also demonstrated that adding 15 mol% C25,25 to SM/CH mixture will cause it to notably interact with fetal bovine serum which could make them a viable alternative adjuvant to synthetic ether-linked lipids in development of advanced liposomal vaccine delivery systems. The prospect of combining the proven strengths of SM/CH mixtures with the unique properties of C25,25 opens exciting possibilities for advancing drug delivery technologies, promising to yield formulations that are both highly effective and adaptable to a range of therapeutic applications.

Quantitative fluorescence emission anisotropy microscopy for implementing homo-fluorescence resonance energy transfer measurements in living cells.

Quantitative fluorescence emission anisotropy microscopy reveals the organization of fluorescently labeled cellular components and allows their characterization in terms of changes in either rotational diffusion or homo-Förster's energy transfer characteristics in living cells. These properties provide insights into molecular organization, such as orientation, confinement, and oligomerization in situ. Here we elucidate how quantitative measurements of anisotropy using multiple microscope systems may be made by bringing out the main parameters that influence the quantification of fluorescence emission anisotropy. We focus on a variety of parameters that contribute to errors associated with the measurement of emission anisotropy in a microscope. These include the requirement for adequate photon counts for the necessary discrimination of anisotropy values, the influence of extinction ratios of the illumination source, the detector system, the role of numerical aperture, and excitation wavelength. All these parameters also affect the ability to capture the dynamic range of emission anisotropy necessary for quantifying its reduction due to homo-FRET and other processes. Finally, we provide easily implementable tests to assess whether homo-FRET is a cause for the observed emission depolarization.

Nature of Linear Spectral Properties and Fast Relaxations in the Excited States and Two-Photon Absorption Efficiency of 3-Thiazolyl and 3-Phenyltiazolyl Coumarin Derivatives.

Coumarin-based fluorescent agents play an important role in the manifold fundamental scientific and technological areas and need to be carefully studied. In this research, linear photophysics, photochemistry, fast vibronic relaxations, and two-photon absorption (2PA) of the coumarin derivatives, methyl 4-[2-(7-methoxy-2-oxo-chromen-3-yl)thiazol-4-yl]butanoate (1) and methyl 4-[4-[2-(7-methoxy-2-oxo-chromen-3-yl)thiazol-4-yl]phenoxy]butanoate (2), were comprehensively analyzed using stationary and time-resolved spectroscopic techniques, along with quantum-chemical calculations. The steady-state one-photon absorption, fluorescence emission, and excitation anisotropy spectra, as well as 3D fluorescence maps of 3-hetarylcoumarins 1 and 2 were obtained at room temperature in solvents of different polarities. The nature of relatively large Stokes shifts (∼4000-6000 cm-1), specific solvatochromic behavior, weak electronic π → π* transitions, and adherence to Kasha's rule were revealed. The photochemical stability of 1 and 2 was explored quantitatively, and values of photodecomposition quantum yields, on the order of ∼10-4, were determined. A femtosecond transient absorption pump-probe technique was used for the investigation of fast vibronic relaxation and excited-state absorption processes in 1 and 2, while the possibility of efficient optical gain was shown for 1 in acetonitrile. The degenerate 2PA spectra of 1 and 2 were measured by an open aperture z-scan method, and the maximum 2PA cross-sections of ∼300 GM were obtained. The electronic nature of the hetaryl coumarins was analyzed by quantum-chemical calculations using DFT/TD-DFT level of theory and was found to be in good agreement with experimental data.

Beta-Lactam Antibiotic Discrimination Using a Macromolecular Sensor in Water at Neutral pH.

Penicillins and cephalosporins belong to the β-lactam antibiotic family, which accounts for more than half of the world market for antibiotics. Misuse of antibiotics harms human health and the environment. Here, we describe an easy, fast, and sensitive optical method for the sensing and discrimination of two penicillin and five cephalosporin antibiotics in buffered water at pH 7.4, using fifth-generation poly (amidoamine) (PAMAM) dendrimers and calcein, a commercially available macromolecular polyelectrolyte and a fluorescent dye, respectively. In aqueous solution at pH 7.4, the dendrimer and dye self-assemble to form a sensor that interacts with carboxylate-containing antibiotics through electrostatic interaction, monitored through changes in the dye’s spectroscopic properties. This response was captured through absorbance, fluorescence emission, and fluorescence anisotropy. The resulting data set was processed through linear discriminant analysis (LDA), a common pattern-base recognition method, for the differentiation of cephalosporins and penicillins. By pre-hydrolysis of the β-lactam rings under basic conditions, we were able to increase the charge density of the analytes, allowing us to discriminate the seven analytes at a concentration of 5 mM, with a limit of discrimination of 1 mM.

Interactions and Dynamics in Aqueous Solutions of pH-Responsive Polymers: A Combined Fluorescence and Dielectric Relaxation Study.

Interaction and dynamics of aqueous solutions of pH-responsive smart polymers are investigated via steady-state, time-resolved fluorescence emission spectroscopy with the help of external local reporter coumarin 153 (C153), while MHz to GHz dielectric relaxation spectroscopic (DRS) measurement reports the intrinsic medium relaxation features. A series of pH-responsive random copolymers (DPL-DP60) comprising of a pH-responsive moiety 2-((leucinyl)oxy)ethyl methacrylate (l-Leu-HEMA) and hydrophobic methyl methacrylate (MMA) are synthesized and characterized. A balance between the pH-responsive (l-Leu-HEMA) and the hydrophobic (MMA) content dictates the phase transition pH, which is found to be ∼5-7 for these aqueous copolymer solutions (1 mg/mL). Dynamic light scattering measurements in aqueous solutions of these polymers reflect a small particle size (∼2-8 nm) at solution pH below their individual phase transition pH, while a large particle size (∼140-340 nm) forms beyond their phase transition pH. No signature of a phase transition pH-driven abrupt change in static and dynamic properties of aqueous polymer solutions has been registered from pH-dependent dielectric relaxation as well as solute (C153)-centric fluorescence measurements. A significant impact of varying the l-Leu-HEMA/MMA segment ratio on steady-state fluorescence emission and rotational anisotropy decay of the fluorophore solute (C153) has been observed. MHz to GHz DRS in aqueous solutions of these pH-responsive polymers reflects bulk water-like dielectric features.

Synthesis and spectral characterization of the first fluorescein-tagged iron(ii) clathrochelates, their supramolecular interactions with globular proteins, and cellular uptake.

A fluorescein-tagged iron(ii) cage complex was obtained in a moderate total yield using a two-step synthetic procedure starting from its propargylamine-containing clathrochelate precursor. An 11-fold decrease in fluorescence quantum yield is observed in passing from the given fluorescein-based dye to its clathrochelate derivative. An excitation energy transfer from the terminal fluorescent group of the macrobicyclic molecule to its quasiaromatic highly π-conjugated clathrochelate framework can explain this effect. The kinetics of the hydrolysis of the acetyl groups of acetylated fluorescein azide and its clathrochelate derivative in the presence of one equivalent of BSA evidenced no strong supramolecular host–guest interactions between BSA and the tested compounds. Study of a chemical stability of the deacetylated iron(ii) clathrochelate suggested the formation of a supramolecular 1 : 1 BSA–clathrochelate assembly. Moreover, an addition of BSA or HSA to its solution caused the appearance of strong clathrochelate-based ICD outputs. The fluorescence emission anisotropy studies also evidenced the supramolecular binding of the fluorescein-tagged iron(ii) clathrochelate to the BSA macromolecule, leading to a high increase in this type of anisotropy. Subcellular uptake of the fluorescein-tagged molecules was visualized using fluorescence microscopy and showed its distribution to be mainly in the cytosol without entering the nucleus or accumulating in any other organelle. An X-rayed crystal of the above propargylamide macrobicyclic precursor with a reactive terminal C Created by potrace 1.16, written by Peter Selinger 2001-2019 ]]> C bond contains the clathrochelate molecules of two types, A and B. The encapsulated iron(ii) ion in these molecules is situated in the center of its FeN6-coordination polyhedron, the geometry of which is intermediate between a trigonal prism (TP) and a trigonal antiprism (TAP). The Fe–N distances vary from 1.8754(6) to 1.9286(4) Å and the heights h of their distorted TP–TAP polyhedra are very similar (2.30 and 2.31 Å); their values of φ are equal to 25.3 and 26.6°. In this crystal, the molecules of types A and B participate in different types of hydrogen bonding, giving H-bonded clathrochelate tetramers through their carboxylic and amide groups, respectively; these tetramers are connected to H-bonded chains.

Location, dynamics and solvent relaxation of a nile red-based phase-sensitive fluorescent membrane probe

Fluorescent membrane probes offer the advantage of high sensitivity, suitable time resolution, and multiplicity of measurable parameters, and provide useful information on model and cell membranes. In this paper, we have explored the location, dynamics, and solvent relaxation characteristics of a novel Nile Red-based phase-sensitive probe (NR12S). Unlike Nile Red, NR12S enjoys unique orientation and location in the membrane, and is localized exclusively in the outer leaflet of the membrane bilayer. By analysis of membrane depth using the parallax approach, we show that the fluorescent group in NR12S is localized at the membrane interface, a region characterized by slow solvent relaxation. Our results show that NR12S exhibits REES (red edge excitation shift), consistent with its interfacial localization. More interestingly, REES of NR12S displays sensitivity to the membrane phase. In addition, fluorescence emission maximum, anisotropy, and lifetime of NR12S are dependent on the membrane phase. We envision that NR12S may prove to be a useful probe in future studies of complex natural membranes.

Tuning the Nature of the Fluorescent State: A Substituted Polycondensed Dye as a Case Study

An extensive spectroscopic analysis is presented of an elongated polycondensed dye with a donor-acceptor substitution. The charge-transfer (CT) state, polarized along the long molecular axis, is close in energy to a local excitation (LE) of the polycondensed system, roughly polarized along the short molecular axis, which makes this system particularly suitable to investigate the subtle LE/CT interplay. An essential-state model is presented that quantitatively reproduces absorption and fluorescence spectra, as well as fluorescence emission and excitation anisotropy spectra collected in solvents of different polarity and viscosity, which sets a sound basis for the understanding of how solvent polarity and solvent relaxation affect the nature of low-lying excitations. The markedly different fluorescence emission and excitation anisotropy spectra measured in glassy and liquid polar solvents unambiguously demonstrate the major role played by solvent relaxation in the definition of fluorescence properties of the dye.

Probing interaction of human serum albumin with the tautomers of 3,4′,7,8-tetrahydroxyflavone by spectroscopic and mass spectrometric approaches

Abstract 3,4′,7,8-Tetrahydroxyflavone (3,4′,7,8-tetraHF) is a kind of plant flavonol, and possesses anti-inflammatory, anti-nociceptive and anti-platelet effects. For understanding its pharmacology, the binding mechanism of 3,4′,7,8-tetraHF to a model protein, human serum albumin (HSA), was probed by fluorescence and UV absorption and mass spectrometric approaches. The binding affinities of the drug with the native HSA and its isomer were about 1.64 ± 0.54 × 10 5 L mol −1 and 0.68 ± 0.53 × 10 5 L mol −1 , respectively. The UV absorption results showed that the hydroxyl groups for both the keto and enol forms of 3,4′,7,8-tetraHF were dissociated and the drug bound with the protein in anion. The fluorescence emission intensities and anisotropy of 3,4′,7,8-tetraHF were dramatically enhanced after interacting with the protein. The binding site of the drug on HSA was located on the motional restricted hydrophobic pocket of subdomain IIA, namely site I, and the drug–protein complex was mainly stabilized by electrostatic and ionic forces.

Spectroscopic and Photophysical Properties of dUTP and Internally DNA Bound Fluorophores for Optimized Signal Detection in Biological Formats

Efficient signal generation in DNA-based assays requires understanding of the influence of fluorophore's interactions on the spectroscopic properties. The resulting changes in fluorescence intensity, quantum yield, emission anisotropy, and fluorescence lifetime provide straightforward tools for the study of molecular dynamics and interaction between labels and nucleic acids. Searching for bright fluorescent reporters for rolling circle amplification (RCA) as efficient signal enhancement strategy for biological formats, we investigated the spectroscopic properties of seven dyes: cyanines, rhodamines, and BODIPYs. They spectrally resemble Cy3, the most frequently used fluorophore in biodetection formats, and are measured in six samples (free dye, dye-dUTP, internally labeled ssDNA and dsDNA-single- and triple-labeled) using steady-state and time-resolved fluorometry. Special emphasis was dedicated to characterizing the nature of the interaction of these fluorophores differing in dye class, charge, and rigidity. Our results suggest dye charge and structure as main factors governing the dye's interactions, with DY-555 and Cy3B presenting the best candidates for our envisaged signal amplification strategy. This label comparison underlines the importance of a proper understanding of structure-property relations and dye-biomolecule interactions for reporter choice and presents a road map towards the design and interpretation of experiments using these labels on DNA of known sequence.

Reversible energy transfer between monomers and fluorescent dimers of rhodamine S in polyvinyl alcohol films

Nonradiative energy transfer and excitation trapping are studied for rhodamine S in polyvinyl alcohol (PVA) films. It occurs that fluorescent dimers of rhodamine S can play a role of imperfect traps for the excitation energy. At highest dye concentrations experimental data of fluorescence quantum yield and emission anisotropy cannot be described by the model of energy transfer neglecting the possibility of excitation return to the monomers. However, the agreement between experimental data and the results of computations can be much improved, if the reverse energy transfer is taken into account. Based on the quantitative analysis it is possible to estimate selected characteristics of rhodamine S dimer in polyvinyl alcohol matrix. The experiments and corresponding analysis are made both at room and elevated temperature.

Codon-Dependent tRNA Fluctuations Monitored with Fluorescence Polarization

During protein synthesis dictated by the codon sequence of messenger RNA, the ribosome selects aminoacyl-tRNA (aa-tRNA) with high accuracy, the exact mechanism of which remains elusive. By using a single-molecule fluorescence resonance energy transfer method coupled with fluorescence emission anisotropy, we provide evidence of random thermal motion of tRNAs within the ribosome in nanosecond timescale that we refer to as fluctuations. Our results indicate that cognate aa-tRNA fluctuates less frequently than near-cognate. This is counterintuitive because cognate aa-tRNA is expected to fluctuate more frequently to reach the ribosomal A-site faster than near-cognate. In addition, cognate aa-tRNA occupies the same position in the ribosome as near-cognate. These results argue for a mechanism which guides cognate aa-tRNA more accurately toward the A-site as compared to near-cognate. We suggest that a basis for this mechanism is the induced fit of the 30S subunit upon cognate aa-tRNA binding. Our single-molecule fluorescence resonance energy transfer time traces also point to a mechanistic model for GTP hydrolysis on elongation factor Tu mediated by aa-tRNA.

Acyl-Chain Mismatch Driven Superlattice Arrangements in DPPC/DLPC/Cholesterol Bilayers

Fluorescence and infrared spectroscopy and cholesterol oxidase activity were employed to investigate the effect of phosphatidylcholine (PC) acyl chain length mismatch on the lateral organizations of lipids in liquid-ordered dipalmitoyl-PC/dilauroyl-PC/cholesterol (DPPC/DLPC/CHOL) bilayers. Plots of steady-state fluorescence emission anisotropy of diphenylhexatriene (DPH) labeled PC (DPH-PC) embedded in the DPPC/DLPC/CHOL bilayers revealed significant peaks at several DPPC mole fractions (Y(DPPC)) when the cholesterol mole fraction (X(CHOL)) was fixed to particular values. Analogously, the DPH-PC anisotropy peaked at several critical X(CHOL)'s when Y(DPPC) was fixed. Acyl chain C-H and C horizontal lineO vibrational peak frequencies of native PC as well as the activity of cholesterol oxidase also revealed dips and peaks at similar Y(DPPC)'s. Importantly, most of the observed peaks/dips coincide with the critical mole fractions predicted by the Superlattice (SL) model. A three-dimensional map of DPH-PC anisotropy versus composition in the range 0.32 <or= X(CHOL) <or= 0.50; 0.54 <or= Y(DPPC) <or= 0.72 revealed a prominent peak at (X(CHOL), Y(DPPC)) approximately (0.42, 0.64). This suggests a simultaneous presence of two different types of superlattices, one where cholesterol is the quest molecule in a PC host lattice and another where DPPC is the guest in the DLPC host lattice. Time-resolved measurements of DPH-PC fluorescence indicated the existence of an ordered, rotationally hindered environment of acyl chains at that "critical" composition consistent with the existence of SL arrangements. We propose that beside CHOL/PC superlattices, DPPC, and DLPC as well tend to adopt regular SL-like lateral distributions relative to each other, presumably because the less hydrophobic DLPC molecule is slightly displaced toward the aqueous phase, thus allowing more room and mobility for the head groups of both DPPC and DLPC as well as for the acyl chain tails of DPPC. The parallel presence of two kinds of superlattices, that is, CHOL/PC-SL and DPPC/DLPC-SL as demonstrated here, has intriguing implications regarding lipid homeostasis of eukaryote membranes.

Influence of MT7 toxin on the oligomerization state of the M1 muscarinic receptor1

The idea that GPCRs (G-protein-coupled receptors) may exist as homo- or hetero-oligomers, although still controversial, is now widely accepted. Nevertheless, the functional roles of oligomerization are still unclear and gaining greater insight into the mechanisms underlying the dynamics of GPCR assembly and, in particular, assessing the effect of ligands on this process seems important. We chose to focus our present study on the effect of MT7 (muscarinic toxin 7), a highly selective allosteric peptide ligand, on the oligomerization state of the hM1 (human M1 muscarinic acetylcholine receptor subtype). We analysed the hM1 oligomerization state in membrane preparations or in live cells and observed the effect of MT7 via four complementary techniques: native-PAGE electrophoresis analysed by both Western blotting and autoradiography on solubilized membrane preparations of CHO-M1 cells (Chinese-hamster ovary cells expressing muscarinic M1 receptors); FRET (fluorescence resonance energy transfer) experiments on cells expressing differently tagged M1 receptors using either an acceptor photobleaching approach or a novel fluorescence emission anisotropy technique; and, finally, by BRET (bioluminescence resonance energy transfer) assays. Our results reveal that MT7 seems to protect the M1 receptor from the dissociating effect of the detergent and induces an increase in the FRET and BRET signals, highlighting its ability to affect the dimeric form of the receptor. Our results suggest that MT7 binds to a dimeric form of hM1 receptor, favouring the stability of this receptor state at the cellular level, probably by inducing some conformational rearrangements of the pre-existing muscarinic receptor homodimers.

Encapsulation of serotonin in β-cyclodextrin nano-cavities: Fluorescence spectroscopic and molecular modeling studies

Serotonin is a physiologically important biogenic amine, deficiency of which leads to mental disorders such as Alzheimer’s disease, schizophrenia, infantile autism, and depression. Both β-cyclodextrin (β-CD) and its chemically substituted synthetic varieties (often possessing enhanced aqueous solubility and improved drug complexing abilities) are finding wide applications as drug delivery vehicles. Here we have studied the encapsulation of serotonin in β-CD and succinyl-2-hydroxypropyl β-cyclodextrin (SHP-β-CD) by exploiting the intrinsic serotonin fluorescence. Enhanced fluorescence emission intensity (which increases by ∼18% and 34% in β-CD and SHPβ-CD respectively) and anisotropy ( r) ( r = 0.075 and 0.1 in β-CD and SHPβ-CD respectively) are observed in presence of the cyclodextrins. From the fluorescence data host–guest interaction with 1:1 stoichiometry is evident, the association constants ( K) being 126.06 M −1 and 461.62 M −1 for β-CD and SHPβ-CD respectively. Additionally, molecular docking and semiempirical calculations have been carried out which provide, for the first time, detailed insights regarding the encapsulation process. In particular, it is evident that the indole ring is inserted within the β-CD cavity with the aliphatic amine side chain protruding towards the primary rim of the β-CD cavity. Docking calculations reveal that hydrogen bonding interactions are involved in the formation of the inclusion complex. Semiempirical calculations indicate that formation of the 1:1 inclusion complex is energetically favorable which is consistent with the fluorescence data.

Polarization Portraits of Single Multichromophoric Systems: Visualizing Conformation and Energy Transfer

A novel technique, two-dimensional (2D) polarization single-molecule imaging, is presented. It is based on measurements and analysis of fluorescence intensity as a function of excitation and emission polarization angles. The technique allows recording of full information on the steady-state polarization properties of fluorescent objects. It is particularly suitable for application to single multichromophoric systems (molecules or nanoparticles) with energy transfer (ET) between different chromophores (e.g., single fluorescent pi-conjugated polymer chains). The 2D polarization data simultaneously provide information on the conformation of the system and the efficiency of its internal excitation ET. The technique is used to characterize single chains and different kinds of chain aggregates of different conjugated polymers at different temperatures. The 2D polarization measurements reveal a dramatic difference in ET taking place in these systems. Clear temperature dependence of ET is observed for individual aggregates as well as for their statistical ensembles. Also, a dependence on solvent and aggregate size is shown. Additionally, extensive "traditional one-dimensional" polarization results on the polarization anisotropy of fluorescence excitation and emission are presented. These results and findings are discussed in relation to internal organization of the nano-objects under study.

Orientation and Dynamics of a Novel Fluorescent Cholesterol Analogue in Membranes of Varying Phase

Fluorescent analogues of cholesterol offer a powerful approach for monitoring cholesterol behavior in membranes because of their high sensitivity, suitable time resolution, and multiplicity of measurable parameters. In this work, we have monitored the orientation and dynamics of a novel fluorescent cholesterol probe, 6-dansylcholestanol (DChol), in membranes of different phase type utilizing sensitive fluorescence techniques including the red-edge excitation shift (REES) approach. Our results show that fluorescence emission maximum, anisotropy, and lifetime of DChol are dependent on the phase of the membrane. Interestingly, DChol exhibits significant red-edge excitation shift (REES) that appear to depend on the phase of the membrane. Analysis of membrane penetration depth by the parallax method shows that the dansyl group of DChol is localized at the interfacial region of the membrane ( approximately 15.6 A from the center of the bilayer). This is in excellent agreement with the previously reported location of cholesterol in fluid-phase membranes. We propose that DChol could be a potentially useful cholesterol analogue in future studies of model and biological membranes.

Molecular Basis of Calmodulin and Ca2+ interaction with thermoTRP channels

The TRP (transient receptor potential) family of proteins are relatively nonselective cation channels, most of which are highly permeable to Ca2+. They participate in many sensory and physiological processes, and subdivided into seven major subfamilies: TRPV, TRPA, TRPC, TRPM, TRPP, TRPML, and TRPN. They are predicted to have six transmembrane domains per subunit, intracellular N- and C-termini, and form tetrameric assemblies. Several TRP channels have been proposed to be regulated by intracellular Ca2+, and/or Calmodulin (CaM). Immunoprecipitation of TRP channels expressed in cultured cells suggests that CaM forms complexes with some of them, but questions remain about whether full-length TRP channels bind CaM directly or through some intermediary protein(s), and about the affinity and kinetics of complex formation. We have used fluorescence emission anisotropy of Alexa 488-labeled CaM to measure binding of CaM and Ca2+-CaM to full-length TRP channels expressed in yeast and purified to homogeneity in detergent solution. At nanomolar concentrations, CaM bound rapidly to TRPA1 and to purified ryanodine receptor used as a positive control. CaM binding was enhanced by Ca2+, but not strictly dependent on it, and binding of labeled CaM was antagonized by addition of unlabeled CaM. Dissociation of the complex occurred much more slowly than association. Under conditions in which TRPA1 and ryanodine receptor bound Ca2+-CaM with high affinity, no binding of Ca2+-CaM was observed for TRPV1 or TRPV2.

Effects of molecular orientation of fluorescently-labeled duplex DNA on emission anisotropy

The molecular orientation of Cy5-labeled target DNA hybridized on a probe DNA layer was studied on a planar waveguide in the Kretchmann configuration at various target DNA bulk concentration. Optical waveguide fluorescence spectroscopy (OWFS) was employed to evaluate the fluorescence emission anisotropy by analyzing two polarized modes, i.e., TM and TE modes propagating in a waveguide. In this way, the orientational order parameter S could be determined. It was found that the orientation of DNA duplex molecules is disordered at lower concentrations, while relatively ordered at higher concentrations.

Molecular Dynamics Simulations of Perylene and Tetracene Librations: Comparison With Femtosecond Upconversion Data

In a prior manuscript by Xu et al. [Xu, J.; Shen, X.; Knutson, J. R. J. Phys. Chem. A 2003, 107, 8383], time-resolved fluorescence emission anisotropy measurements were performed on perylene and tetracene in hexadecane using an upconversion technique with approximately 100 fs resolution. The anisotropy transients contained previously unseen decay terms of approximately 300 fs. In perylene, their amplitude corresponded to the " r(o) defect" that has gathered interest over decades. We ascribed this term to a predominantly in-plane libration. In this manuscript, we present molecular dynamics simulations for the motions of perylene and tetracene using the CHARMm molecular dynamics program (version c29b2). Both rotational correlation functions contain subpicosecond decay terms that resemble experimental anisotropy decays. It was suggested that the r(o) defect might arise from excited-state distortions of perylene, so we conducted quantum mechanical calculations to show that such distortion does not significantly displace the oscillators. We compare the case of perylene, with a strongly allowed singlet emission transition, to that of the weakly allowed tetracene transition. In perylene, motion alone can explain subpicosecond anisotropy decay, while tetracene decay also contains vibrational coupling terms, as previously reported by Sarkar et al. [Sarkar, N.; Takeuchi, S.; Tahara, T. J. Phys. Chem. A 1999, 103, 4808].