Darcy Lichlyter

Dyes with Segmental Mobility: Molecular Rotors

Molecular rotors are fluorescent molecules that are characterized by the ability to form twisted states through the rotation of one segment of the structure with respect to the rest of the molecule. Intramolecular rotation changes the ground- state and excited-state energies, and molecular rotors deexcite from the twisted state either without photon emission or with a different wavelength than from the LE state. Intramolecular rotation is strongly dependent on the solvent. Solvent polarity, hydrogen bond formation, isomerization, excimer formation, and steric hindrance are predominant forms of solvent-fluorophore interaction. Of highest importance is sterichindrance,becauseitlinksthesolventsmicroviscositytotheformationrateof TICT states, which, in turn, determines the spectral emission. For this reason, molecular rotors have found a wide range of applications as fluorescent sensors of microviscosity and solvent free volume. Application examples include bulk viscos- ity measurement, probing dynamics of polymer formation, protein sensing and probing of protein aggregation, and microviscosity probing in living cells.

Self-calibrating viscosity probes: Design and subcellular localization

We describe the design, synthesis and fluorescence profiles of new self-calibrating viscosity dyes in which a coumarin (reference fluorophore) has been covalently linked with a molecular rotor (viscosity sensor). Characterization of their fluorescence properties was made with separate excitation of the units and through resonance energy transfer from the reference to the sensor dye. We have modified the linker and the substitution of the rotor in order to change the hydrophilicity of these probes thereby altering their subcellular localization. For instance, hydrophilic dye 12 shows a homogeneous distribution inside the cell and represents a suitable probe for viscosity measurements in the cytoplasm.

Synthesis and evaluation of self-calibrating ratiometric viscosity sensors

We describe the design, synthesis and fluorescent profile of a family of self-calibrating dyes that provide ratiometric measurements of fluid viscosity. The design is based on covalently linking a primary fluorophore (reference) that displays a viscosity-independent fluorescence emission with a secondary fluorophore (sensor) that exhibits a viscosity-sensitive fluorescence emission. Characterization of fluorescent properties was made with separate excitation of the units and through Resonance Energy Transfer from the reference to the sensor dye. The chemical structures of both fluorophores and the linker length have been evaluated in order to optimize the overall brightness and sensitivity of the viscosity measurements. We also present an application of such ratiometric dyes for the detection of membrane viscosity changes in a liposome model.