BODIPY-based red fluorescent molecular rotors for microviscosity sensing Karolina Maleckaitė 1 , Jelena Dodonova-Vaitkūnienė 2 , Stepas Toliautas 2 , Džiugas Jurgutis 4 , Rugilė Žilėnaitė 1 , Vitalijus Karabanovas 3,4 , Sigitas Tumkevičius 2 and Aurimas Vyšniauskas 1 1 State Research Institute Center for Physical Sciences and Technology, Lithuania, 2 Vilnius University, Lithuania, 3 National Cancer Institute, Lithuania, 4 Vilnius Gediminas Technical University, Lithuania Microscopic viscosity (microviscosity) - an essential biological property that can provide information about diffusion-controlled mechanisms inside the live cell. Changes in microviscosity may indicate the development of serious illnesses, such as atherosclerosis and Alzheimer's disease [1]. In addition, microviscosity imaging in vivo can reveal cell death of cancer cells affected by photodynamic therapy [2]. Fluorescent molecular rotors (FMRs) provide a simplified way to image microviscosity in biological environments. The operating principle of fluorescence lifetime-based FMRs relies on competition between intramolecular rotation and non-radiative relaxation to the electronic ground state [1]. The viscous environment restricts the rotation of the rotor while increasing the radiative relaxation. Therefore, the longer fluorescence lifetime is observed in a more viscous medium. FMRs are used in Fluorescence Lifetime Imaging Microscopy (FLIM), which allows simultaneous imaging of microviscosity throughout the sample. FMRs based on the BODIPY group are widely used because of their ease of functionalization and monoexponential fluorescence decay, the latter being very important for simplified FLIM data analysis. Unfortunately, viscosity-sensitive BODIPY derivatives are usually green fluorescent, which complicates their use in live cell imaging. Red fluorophores are preferred in cellular imaging because they avoid autofluorescence and provide a deeper tissue penetration [3]. However, it is challenging to develop a red fluorescent probe that retains viscosity sensitivity. In this work [4,5], we present β -phenyl- and β -vinyl-substituted viscosity-sensitive red fluorescent molecular rotors based on a meso -phenyl-substituted BODIPY core (Fig. 1). We show how quantum calculations can help determine the viscosity sensitivity prior to chemical synthesis. Additionally, we analyse photophysical properties of the new derivatives and their sensitivity to viscosity, temperature, and polarity of the solvent. Moreover, we show how the BP-PH-2M-NO 2 probe can be used for imaging lipid droplets in live cells. In addition, BP-Vinyl-NO 2 demonstrates the ability to measure viscosity in highly viscous environments and extend the range of viscosity sensitivity by up to seven orders of magnitude.
Figure 1. Molecular structures of BODIPY-based fluorescent molecular rotors presented in this work. References 1. M. K. Kuimova; Physical Chemistry Chemical Physics 14 (2012) pp. 12671–12686. 2. L. E. Shimolina; Scientific Reports 7 (2017), art.n. 41097 3. D. Zhang et al; Physical Chemistry Chemical Physics 13 (2011) pp. 13026–13033. 4. K. Maleckaite et al; Methods and Applications in Fluorescence 10 , 3 (2022), pp. 034008 5. K. Maleckaite et al; Chemistry – A European Journal 27 , 67 (2021), pp. 16768-16775
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