Naphthyridyl‐Substituted 4,4‐Difluoro‐4‐bora‐3a,4a‐diaza‐s‐indacene (BODIPY) Luminophores: Photophysics and Application as Molecular Imaging Probes in Live Cells

Abstract

The synthesis, optical properties, and cellular uptake of a family of 4,4‐difluoro‐4‐bora‐3a,4a‐diaza‐s‐indacene (BODIPY) derivatives that incorporate a naphthyridine substituent at the 8‐position of the BODIPY core and either alkyl, bromo, or dimethylamino aryl groups at the 2‐ and 6‐positions are described. The family of compounds is classified, based on their optical properties, into two types; Class I, compounds with alkyl substituents at the 2‐ and 6‐positions show intense solvent‐independent emission, which arises from the BODIPY core with no involvement of the naphthyridine. Class II, the dimethylaminoaryl‐containing compounds exhibit charge transfer transitions that lead to NIR fluorescence and remarkably large Stokes shifts, which make them potentially attractive in bioimaging. Class II compounds exhibit complex solvent dependence. There is evidence for dual fluorescence from multiple rotamers; this is rationalized with the aid of time‐dependant density functional theory (TDDFT) calculations. A representative compound from each class was PEGylated, which rendered them water soluble, but their photophysical properties were maintained in aqueous buffered media. The uptake of the PEGylated BODIPY compounds and the nonPEGylated parent compounds by live mammalian cells was compared by using confocal fluorescence microscopy and resazurin blue cytotoxicity assays were performed. All compounds tested exhibited good cell uptake, were largely nuclear excluding, and had low toxicity at 10 μM. This is to our knowledge the first demonstration of such a mega‐Stokes‐shifted BODIPY probe in cell imaging.

The application of water soluble, mega‐Stokes‐shifted BODIPY fluorophores to cell and tissue imaging

“…The detailed photophysical properties of the parent compounds were reported previously (Martin et al. , ) and absorbance and emission spectra of each are shown in Figure S1. The visible emitting BODIPY parent (VIS) exhibits an absorbance maximum, λ abs, at 532 nm and emission maximum, λ em 547 nm (15 nm Stokes shift), the PEGylated analogue (VIS‐PEG) has an absorbance maximum λ abs at 528 nm and λ em 552 nm (Stokes shift of 24 nm).…”

“…, ; Martin et al. , ). Both compounds were rendered water‐soluble by conjugation to PEG, a well established approach to improving cellular transport of otherwise cell impermeable material (Greenwald, ; Uppal et al.…”

“…Self‐quenching was not observed up to 100 μM of the NIR dye, a concentration far exceeding that needed for cellular imaging (Martin et al. , ). Uptake and distribution between the dyes in five cell lines were compared.…”

“…Conjugation to PEG rendered these compounds water soluble, with retention of their attractive photophysical characteristics in aqueous media. Preliminary data was also reported, which showed the ability of this NIR emitting dye to cross the cell membrane when conjugated to PEG (Martin et al, 2013).…”

“…Preliminary data was also reported, which showed the ability of this NIR emitting dye to cross the cell membrane when conjugated to PEG (Martin et al. , ).…”