Signaling Recognition Events with Fluorescent Sensors and Switches.

6. Rehybridization of the Acceptor (RICT) 3908 7. Planarization of the Molecule (PICT) 3909 III. Fluorescence Spectroscopy 3909 A. Solvent Effects and the Model Compounds 3909 1. Solvent Effects on the Spectra 3909 2. Steric Effects and Model Compounds 3911 3. Bandwidths 3913 4. Isoemissive Points 3914 B. Dipole Moments 3915 C. Radiative Rates and Transition Moments 3916 1. Quantum Yields and Radiationless Deactivation Processes 3916

Structural Changes Accompanying Intramolecular Electron Transfer: Focus on Twisted Intramolecular Charge-Transfer States and Structures

Rhodamine dyes are widely used as fluorescent probes owing to their high absorption coefficient and broad fluorescence in the visible region of electromagnetic spectrum, high fluorescence quantum yield and photostability. A great interest in the development of new synthetic procedures for preparation of Rhodamine derivatives has arisen in recent years because for most applications the probe must be covalently linked to another (bio)molecule or surface. In this critical review the strategies for modification of Rhodamine dyes and a discussion on the variety of applications of these new derivatives as fluorescent probes are given (108 references).

Synthesis and applications of Rhodamine derivatives as fluorescent probes

The understanding of the dual fluorescence of certain aromatic systems has greatly advanced in recent years. The accompanying large charge separation has been shown to be linked to a twisted (or small overlap) arrangement of the chromophores. Recent theoretical models are able to describe the excited-state twisting of both single bonds (TICT compounds) and double bonds (olefins) in a unified picture. These models can help to elucidate the photophysical behavior of many organic, inorganic, and biologically relevant compounds, and their application to laser dyes and fluorescent probes provides a route to new “tailor-made” fluorescent materials. Applied to the primary processes of vision and photosynthesis, these models can lead to a deeper understanding of basic photobiological processes.

Charge Separation in Excited States of Decoupled Systems—TICT Compounds and Implications Regarding the Development of New Laser Dyes and the Primary Process of Vision and Photosynthesis

Specific labeling of biomolecules with bright fluorophores is the keystone of fluorescence microscopy. Genetically encoded self-labeling tag proteins can be coupled to synthetic dyes inside living cells, resulting in brighter reporters than fluorescent proteins. Intracellular labeling using these techniques requires cell-permeable fluorescent ligands, however, limiting utility to a small number of classic fluorophores. Here we describe a simple structural modification that improves the brightness and photostability of dyes while preserving spectral properties and cell permeability. Inspired by molecular modeling, we replaced the N,N-dimethylamino substituents in tetramethylrhodamine with four-membered azetidine rings. This addition of two carbon atoms doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging. The novel substitution is generalizable, yielding a palette of chemical dyes with improved quantum efficiencies that spans the UV and visible range.

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A general method to improve fluorophores for live-cell and single-molecule microscopy

Structural relaxation of rhodamine dyes with different N-substitution patterns: A study of fluorescence decay times and quantum yields

The high fluorescence quantum yields ϕf of most Triphenylmethane(TPM)-dyes and carbon-bridged TPM-dyes in highly viscous solutions (ϕf ranging from 03 to 08) decrease with increasing temperature by a factor > 100, due to viscosity dependent nonradiative deactivation The deactivation rates krd of bridged model dyes are significantly smaller (by a factor of up to 170) as compared to krd of the unbridged crystal violet - Application of the Twisted Intramolecular Charge Transfer (TICT)-model to neutral and ionic TPM-dyes yields an explanation for the different behaviour of similarly structured compounds The influence of steric and electronic factors on various TPM-dyes is discussed within this framework This leads to a simple rule involving donor and acceptor properties of molecular subunits, which permits the prediction of fluorescence properties (high quantum efficiency for laser dyes or ultrafast deactivation for saturable absorbers)

Efficient intramolecular fluorescence quenching in triphenylmethane-dyes involving excited states with charge separation and twisted conformations.

The nonexponential fluorescence decay behaviour of triphenylmethane-(TPM)-dyes can be understood as barrierless relaxation, i.e. intramolecular rotational relaxation of dialkylanilino groups under the influence of a driving force. The observed viscosity dependent rotational relaxation rates of TPM-dyes with large and small, rigid and flexible rotor groups do not obey the Debye-Stokes-Einstein relationship but can be fitted within the microviscosity theory of Gierer and Wirtz and show that free volume effects are important. The relative size of free volume effects in n-alcohols, diols and glycerol can be determined by comparing the rates of different TPM-dyes.

Excited state dynamics of triphenylmethane‐dyes used for investigation of microviscosity effects

Evidence for the formation of biradicaloid charge-transfer (BCT) states in xanthene and related dyes

The decay of the dual fluorescence of the para-substituted dialkylanilines (esters and nitriles) as well as the fluorescence decay of several triphenylmethane (TPM) dyes is investigated. The esters relax several times faster than the nitriles, and in the TPM series, relaxation is accelerated by increasing the substituent acceptor strength. In both cases, non-exponential decay behaviour is observed. It is more pronounced for the compounds with stronger driving forces along the reaction pathways. Freezing of the relaxation process transforms the decay into a mono-exponential one. This is discussed in terms of two models, namely by a log-gaussian distribution of relaxation times, and by a non-radiative sink model.

Martin Vogel

Papers

Structural relaxation of rhodamine dyes with different N-substitution patterns: A study of fluorescence decay times and quantum yields

Efficient intramolecular fluorescence quenching in triphenylmethane-dyes involving excited states with charge separation and twisted conformations.

Excited state dynamics of triphenylmethane‐dyes used for investigation of microviscosity effects

Evidence for the formation of biradicaloid charge-transfer (BCT) states in xanthene and related dyes

The dynamics of adiabatic photoreactions as studied by means of the time structure of synchrotron radiation