Synthesis of a fluorinated pyronin that enables blue light to rapidly depolarize mitochondria

Abstract

Fluorinated analogues of the fluorophore pyronin B were synthesized as a new class of amine-reactive drug-like small molecules. In water, 2,7-difluoropyronin B was found to reversibly react with primary amines...

Improving the Brightness of Pyronin Fluorophore Systems through Quantum-Mechanical Predictions

“…Interestingly, isopropylamino substitution lowers molar absorptivity which adversely affects fluorophore brightness, despite relatively high quantum yields. Azetidinyl substitution, as noted in previous studies, 24,58 produced highly fluorescent pyronins in the 550 nm excitation range; nevertheless, both 1 and 8 are less fluorescent than monoalkylamino species 2, 3, and 11, which appear to better mitigate TICT transitions states due to larger energetic barriers of rotation about the amino group in the S 1 -min structures. Overall, fluorophore intensities follow the trend 3 > 2 > 11 > 1 > 8 > 10 > 4 > 12 ≫ PB > PY.…”

“…Xanthene-based structures with 3,6-diamino substitution, more commonly referred to as pyronins (Figure ), have origins in the late 19th century , and became popular as dyes originally in textiles and then later in the scientific community for imaging applications. , Pyronins, like structurally similar rhodamines, typically exhibit fluorescent properties and are commonly employed in microscopy, , cell cycle studies, electrophoresis, , flow cytometry, , paraffin section analysis, , mitochondrial studies, − detecting protein–protein interactions, and chemo/ion sensors, , among other uses. − Most applications utilize commercially available pyronin Y (PY) or pyronin B (PB) and to a lesser extent acridine red (AR) which preferentially intercalate RNA allowing for imaging/detection at an excitation wavelength (λ ex ) of ∼530 nm (AR)/∼550 nm (PY/PB) and an emission wavelength (λ em ) of ∼560 nm (AR)/∼570 nm (PY/PB); however, numerous synthetically tailored pyronins have been generated for specialized uses. ,, While PB, AR, and particularly PY remain popular dyes, the quantum yields (Φ f ) for these fluorophores in aqueous solvents are relatively low in comparison to other xanthene-based fluorophores such as rhodamines and fluoresceins, limiting the sensitivity of these fluorophore systems. As a result, pyronins are often applied in high concentrations (∼100 μM) to stain cells which can lead to cytotoxicity.…”