Rational Engineering of Bioinspired Anthocyanidin Fluorophores with Excellent Two-Photon Properties for Sensing and Imaging

Abstract

Fluorescent materials are widely employed in biological analysis owing to their biorthogonal chemistries for imaging and sensing purposes. However, it is always a challenge to design fluorophores with desired photophysical and biological properties, due to their complicated molecular and optical nature. Inspired by anthocyanidin, a class of flower pigments, we designed a new fluorescent molecular framework, AC-Fluor. The new fluorescent materials can be rationally engineered to produce a broad range of fluorescent scaffolds with flexibly tunable emission spectra covering the whole visible light range, from 467 to 707 nm. Furthermore, they exhibit unprecedented environment-insensitive two-photon properties with a substantial cross section as large as 1100 GM in aqueous solution. AC-Fluors demonstrate their biological values through two-photon deep tissue imaging, with penetration depths as much as 300 μm, while exhibiting minimal cytotoxicity. These features engender a rational engineering strategy for the design and optimization of new fluorescent materials for biological imaging.

Donor and Ring‐Fusing Engineering for Far‐Red to Near‐Infrared Triphenylpyrylium Fluorophores with Enhanced Fluorescence Performance for Sensing and Imaging

“…Rational molecular engineering of fluorophores, based on a deep understanding of their structure–photophysical property relationship (SPPR), is crucial and imperative to yield novel fluorophores with desired optical and biological characteristics . Despite major efforts, only a few fluorochromes have been subjected to systematic SPPR research .…”

“…Density functional theory (DFT) calculations indicated that the π electrons in the LUMO of TPP are delocalized over the entire π‐conjugated 2,4,6‐triphenylpyrylium framework, whereas the HOMO is mostly localized on the 2,6‐phenyl substituents (Figure b). The 4′‐ and 4′′′‐positions of rings B and D show the highest electron density (Figure b), which indicates that introducing electron‐donating groups at these positions can lead to a significant bathochromic shift in the absorption/emission profiles of TPP dyes . DFT calculations also indicated that C‐2 and C‐6 of the pyrylium moiety are the most electron‐deficient positions (Figure b), implying that nucleophilic attack at these sites is responsible for the instability of TPP .…”

“…However, advanced as they are, these stable, large Stokes shift, and NIR‐emitting TPP dyes are still unsuitable as fluorophores for bioimaging because of their weak fluorescence ( Φ f =0.05 ( TPP1 ), Φ f =0.04 ( TPP2 ) in EtOH) (Table , Figure d). It is well known that rigid and planar molecular structures can endow organic dyes with high fluorescence quantum yields . To confirm our hypothesis, we restricted the rotation between rings B/D and A by linking them with methylene units to afford compounds TPPF1 and TPPF2 (Table ).…”

“…When we appended an electron‐donating NMe 2 group on ring C (578/640 nm, Φ f =0.28 in EtOH, TPPF4 ), an obvious decrease in fluorescence quantum yield accompanied by a slight hypsochromic shift was observed. This may have been due to extension of the molecular orbital and photo‐induced electron transfer (PeT) . On the other hand, when the electron density at R 2 of TPP‐Fluor was increased, as in TPPF6 , a significant bathochromic shift and a slight decrease in fluorescence intensity were observed.…”

“…[23,[28][29][30] Rational molecular engineering of fluorophores, based on a deep understanding of their structure-photophysical property relationship (SPPR), is crucial and imperative to yield novel fluorophores with desired opticala nd biological characteristics. [31,32] Despitem ajor efforts,o nly af ew fluorochromes have been subjected to systematic SPPR research. [31,[33][34][35] This is probablyd ue to the time-consuming synthetic protocols for obtaining structurala nalogues of ac ertain fluorophore.…”