Viscosity imaging at a microscopic scale can provide important information about biosystems, including the development of serious illnesses. Microviscosity imaging is achievable with viscosity-sensitive fluorophores, the most popular of which are based on the BODIPY group. However, most of the BODIPY probes fluoresce green light, whereas the red luminescence is desired for the imaging of biological samples. Designing a new viscosity probe with suitable spectroscopic properties is a challenging task because it is difficult to preserve viscosity sensitivity after modifying the molecular structure. Here we describe how we developed a new redemitting, viscosity-sensitive, BODIPY fluorophore BP-PH-2M-NO 2 that is suitable for reliable intracellular viscosity imaging of lipid droplets in MCF-7 breast cancer cells. The design of BP-PH-2M-NO 2 was aided by DFT calculations that allowed a successful prediction of the viscosity sensitivity of fluorophores before synthesis. In summary, we report a new red viscosity probe possessing monoexponential fluorescence decay that makes it attractive for lifetime-based viscosity imaging.

Mapping microviscosity, temperature, and polarity in biosystems is an important capability that can aid in disease detection. This can be achieved using fluorescent sensors based on a green-emitting BODIPY group. However, red fluorescent sensors are desired for convenient imaging of biological samples. It is known that phenyl substituents in the β position of the BODIPY core can shift the fluorescence spectra to longer wavelengths. In this research, we report how electron-withdrawing (EWG) and -donating (EDG) groups can change the spectral and sensory properties of β-phenyl-substituted BODIPYs. We present a trifluoromethyl-substituted (EWG) conjugate with moderate temperature sensing properties and a methoxy-substituted (EDG) molecule that could be used as a lifetime-based polarity probe. In this study, we utilise experimental results of steady-state and time-resolved fluorescence, as well as quantum chemical calculations using density functional theory (DFT). We also explain how the energy barrier height (Ea) for non-radiative relaxation affects the probe’s sensitivity to temperature and viscosity and provide appropriate Ea ranges for the best possible sensitivity to viscosity and temperature.

Intracellular polarity in lipid droplets as well as other organelles may provide useful knowledge about various processes taking place in live cells. Therefore, small fluorophores capable of visualising polarity are...

Microviscosity has a strong impact for diffusion-controlled processes in biological environments. BODIPY molecular rotors are viscosity-sensitive fluorophores that provide a simple and non-invasive way to visualise microviscosity. Although green-fluorescent probes are already well developed for imaging, thick biological samples require longer wavelengths for investigation. This work focuses on the examination of novel β-substituted meso-phenyl-BODIPYs possessing a red emission. We report a new red-fluorescent BODIPY-based probe BP-Vinyl-NO2 suitable for sensing microviscosity in rigid environments of over 100 000 cP viscosities. Furthermore, we demonstrate that changing the methyl position from ortho to meta on the β-phenyl-substituted conjugate BP-PH-m2M-NO2 redshifts absorbance and fluorescence spectra while maintaining viscosity sensitivity. Finally, we show that nitro-substitution of meso-phenyl is a versatile approach to improve the sensitivity to viscosity while suppressing sensitivity to polarity and temperature of such derivatives. In summary, we present two nitro-substituted red-fluorescent probes that could be used as lifetime-based microviscosity sensors.