Issue 10, 2020

One- and two-photon solvatochromism of the fluorescent dye Nile Red and its CF3, F and Br-substituted analogues

Abstract

The solvatochromic fluorophore Nile Red, 9-diethylamino-5H-benzo[a]phenoxazine-5-one, is one of the most commonly used stains to enhance contrast of lipid-rich areas of microscopic biosamples. Quite surprisingly, relatively little is known about the spectrally-resolved two-photon absorption (2PA) properties of this dye despite its promising features for two-photon microscopy of biological matter. For this reason, the two-photon solvatochromism of Nile Red still remains an uncharted territory as well. Also, no study has yet reported on how electron-withdrawing substituents attached to the Nile Red backbone affect its solvatochromic properties and two-photon brightness. In this paper, we demonstrate how solvent polarity influences the one- and two-photon absorption spectra of Nile Red as well as its fluorescence parameters, and we present new analogues that contain –CF3, –F and –Br substituents on its eastern side. Two-photon excited fluorescence experiments in a broad spectral range (780–1240 nm) and electronic structure calculations show that both the nature and location of the substituent have particular influence on the strength of 2PA, peaking in all cases at approx. 860 and 1050 nm. 2PA cross sections are higher at 1050 nm than at 860 nm, which suggests that Nile Red and its analogues are best suited for two-photon imaging employing excitation in the NIR-II optical transparency window of biological tissues.

Graphical abstract: One- and two-photon solvatochromism of the fluorescent dye Nile Red and its CF3, F and Br-substituted analogues

Supplementary files

Article information

Article type
Paper
Submitted
05 Mar 2020
Accepted
25 Aug 2020
First published
25 Aug 2020

Photochem. Photobiol. Sci., 2020,19, 1382-1391

One- and two-photon solvatochromism of the fluorescent dye Nile Red and its CF3, F and Br-substituted analogues

M. Hornum, P. Reinholdt, J. K. Zaręba, B. B. Jensen, D. Wüstner, M. Samoć, P. Nielsen and J. Kongsted, Photochem. Photobiol. Sci., 2020, 19, 1382 DOI: 10.1039/D0PP00076K

To request permission to reproduce material from this article, please go to the Copyright Clearance Center request page.

If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given.

If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page.

Read more about how to correctly acknowledge RSC content.

Spotlight

Advertisements