Chinese Chemical Letters
Original article
Highly selective two-photon fluorescent probe for imaging of nitric
oxide in living cells
a,b,
Qing Liu a,c, Lin Xue a, Dong-Jian Zhu a,c, Guo-Ping Li a,c, , Hua Jiang
*
*
a Beijing National Laboratory for Molecular Sciences, CAS Key Laboratory of Photochemistry, Institute of Chemistry, Chinese Academy of Sciences,
Beijing 100190, China
b College of Chemistry, Beijing Normal University, Beijing 100875, China
c University of Chinese Academy of Sciences, Beijing 100049, China
A R T I C L E I N F O
A B S T R A C T
Article history:
A new two-photon fluorescent probe, ADNO, for nitric oxide (NO) based on intramolecular photoinduced
electron transfer (PET) mechanism displays a rapid response to NO with a remarkable fluorescent
enhancement in PBS buffer. The excellent chemoselectivity of ADNO for NO over other ROS/RNS (reactive
oxygen species or nitrogen species) and common metal ions was observed. Moreover, ADNO has been
successfully applied in fluorescence imaging of NO of living cells using both one-photon microscopy
(OPM) and two-photon microscopy (TPM).
Received 12 October 2013
Received in revised form 30 October 2013
Accepted 8 November 2013
Available online 20 November 2013
Keywords:
ß 2013 Guo-Ping Li. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Fluorescent probe
Nitric oxide
Fluorescence imaging
One-photon microscopy
Two-photon microscopy
1. Introduction
central tool for visualizing various ROS/RNS (reactive oxygen
species or nitrogen species) [3]. Compared with standard laser
Nitric oxide (NO), an important and ubiquitous endogenous
second messenger or cytostatic/cytotoxic agent, has attracted
more and more attention in recent years [1]. It is believed that NO
is, at low concentrations, essential for many normal physiological
processes and plays an important role in several physiological
functions, such as regulation of vascular tone, neurotransmission,
anti-viral defense, immune response and memory. Whereas
excessive generation of NO in physiological processes can cause
damage to biomolecules, such as DNA, proteins and lipids, and
consequently lead to cellular necrosis or apoptosis. Therefore,
sensitive and specific detection of NO in living systems is indeed
indispensable for understanding the biological roles of NO. The
development of techniques to analyze NO in organisms has
dramatically increased over the last decades [2], especially,
fluorescence microscopy, which features non-destruction, high
sensitivity and spatiotemporal resolution, has been proved to be a
confocal approaches which employ one photon of higher energy
for the excitation, two-photon microscopy of lower energy as the
excitation source has significant advantages by providing deeper
sectioning, less phototoxicity, and lower background fluorescence
[4]. Although a number of NO-specific fluorescent probes have
been reported to date [5,3b,c,e], two-photon fluorescent probes for
imaging of NO suitable for biological applications are still rare.
In this report, we designed and synthesized a new two-photon
fluorescent probe, ADNO (2-(a-(3,4-diaminophenoxy)acetyl)-6-
(dimethylamino)naphthalene), for monitoring NO based on
photoinduced electron transfer (PET) mechanism. As shown in
Scheme 1, the probe is composed of two moieties: o-phenylene-
diamine as the NO-sensitive fluorescence modulator and 2-acetyl-
6-(dimethylamino)naphthalene (Acedan) as the two-photon
fluorophore. We anticipated that detection of NO can be well
achieved by altering the electron-donating capacity of the o-
phenylenediamine moiety. The electron-rich o-phenylenediamine
group should quench the fluorescence of Acedan due to the
existence of electron transfer from electron-rich diamine moiety to
the excited fluorophore. However, the presence of NO under
aerobic conditions leads to the transformation of o-phenylene-
diamine to benzotriazole, which consequently suppresses the PET
process and revives the strong fluorescence response of the probe.
*
Corresponding authors at: Beijing National Laboratory for Molecular Sciences,
CAS Key Laboratory of Photochemistry, Institute of Chemistry, Chinese Academy of
Sciences, Beijing 100190, China.
(H. Jiang).
1001-8417/$ – see front matter ß 2013 Guo-Ping Li. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.