The near-infrared fluorescent probes based on phenoxazine for the rapid detection of hypochlorous acid
Introduction
Reactive oxygen species (ROS), as one of endogenous metabolites, are of great importance for physiological processes, while excessive ROS are closely correlated with a wide range of various diseases, including inflammation, cancer and neurodegenerative diseases [[1], [2], [3], [4], [5], [6]]. Hypochlorous acid (HOCl), generating from the reaction between hydrogen peroxide and chloride ions via myeloperoxidase (MPO)-mediated in vivo [7,8], is a member of the highly active ROS. It is considered to be an important bactericidal oxidant in the immune system against pathogenic microorganisms including germs, viruses and so on [[9], [10], [11]]. However, excessive production of HOCl can oxidize phospholipids, proteins and deoxyribonucleic acid (DNA), leading to cell and tissue damage [[12], [13], [14], [15]]. Meanwhile, many diseases are closely related to the overproduction of HOCl, such as atherosclerosis, rheumatoid arthritis and so on [[16], [17], [18], [19]]. Therefore, it is greatly urgent to develop highly selective and sensitive fluorescent probes for the rapid detection of HOCl.
Owing to fluorescent probes enabling real-time and in situ detecting HOCl in biological organisms [[20], [21], [22], [23]], a wide range of fluorescent probes, on the basis of a series of fluorophores including BODIPY [[24], [25], [26], [27]], naphthalimide [28,29], rhodamine [30,31], coumarin [32] and so on, have been reported. Although numerous fluorescent probes are capable of detecting HOCl in vitro or in cells [[33], [34], [35], [36], [37], [38], [39], [40], [41], [42], [43], [44], [45]], few probes can be employed in living system mainly owing to the limitation of emission wavelength. Therefore, it is urgently required to develop NIR fluorescent probes to detect HOCl in living organisms because they possess deeper penetration depth and little autofluorescence interference [[46], [47], [48]]. It is well established that the construction of “turn-on” NIR fluorescent probes is an important strategy for detecting HOCl because of significant fluorescence increase after response with high signal-to-noise ratio. Yi developed a novel “turn-on” mechanism for specifically detecting HOCl, taking advantage of remarkable fluorescence change between the reduced form and oxidized form of methylene blue (MB) [49,50]. It is well known that phenoxazine possesses similar photophysical properties to methylene blue (Scheme 1), its maximum fluorescence emission lying in the NIR area (roughly 680 nm) and its reduced form of phenoxazine (RO1) emits little fluorescence. Herein, RO1 and its derivatives are ideal scaffolds for the design of turn-on fluorescent probes.
Taking these into consideration, we designed and synthesized two fluorescent probes BC-2 and BC-3 based on RO1 (Scheme 1) for the specific detection of HOCl both in vitro and in living organisms. The two probes BC-2 and BC-3 released fluorophore phenoxazine (Scheme 1) in the presence of HOCl, accompanying with significant fluorescence intensity increase in NIR emission and remarkable colour change from colourless to blue. It is worth noting that the probe BC-3 exhibited high selectivity, sensitivity and rapid response towards HOCl both in cells and in mouse models of arthritis.
Section snippets
Materials and general methods
The stock solutions (1 mM) of probes was prepared in ethanol (HPLC grade), and further diluted with PBS (sodium phosphate buffer, pH = 7.4) to from a final concentration (10 μM). Materials involved in the preparation of active species, including t-BuOOH (tert-butyl hydroperoxide), 5% NaOCl solution, 30% H2O2 solution, nitroferricyanide (III) dehydrate, KO2, 3-morpholinosydnonimine hydrochloride, 2, 2′-azobis(2-amidinopropane)dihydrochloride and ferrous sulphate (hydroxyl radicals was produced
The response ability of probes towards HOCl
We first evaluated the ability of BC-2 and BC-3 to detect HOCl by spectroscopy in PBS solution (pH = 7.4). The intensity of fluorescence and absorption considerably rose with the increase of HOCl concentrations (Fig. 1 and Fig. S1). Furthermore, the limit of detection (LOD) of probes were tested using fluorescence titration, and the LOD values of BC-2 and BC-3 were as much low as 20.4 nM and 11.1 nM, respectively (Fig. 1b and Fig. S1b). More importantly, the two probes exhibited fast response
Conclusion
In summary, two new fluorescent probes (BC-2 and BC-3) are designed and synthesized based on phenoxazine for the detection of HOCl both in vitro and in vivo. In the presence of HOCl, the probes can rapidly detect HOCl using both fluorescence and absorption changes in vitro. Furthermore, the probes exhibited excellent sensitivity and selectivity for HOCl over other ROS/RNS. More importantly, the probe BC-3 was able to detect the endogenous HOCl level in living cells. Moreover, BC-3 could
CRediT authorship contribution statement
Wubin Zheng: Investigation, Data curation, Writing - original draft. Jiajia Yang: Investigation. Yang Shen: Data curation. Yusi Yao: Validation. Guanglei Lv: Conceptualization, Writing - review & editing, Supervision. Shiyou Hao: Project administration. Chunxia Li: Writing - review & editing, Supervision.
Declaration of competing interest
The authors declare no competing financial interest.
Acknowledgments
This research was funded by the National Natural Science Foundation of China (51572258, 51872263 and 21876158), Zhejiang Provincial Natural Science Foundation of China (LZ19E020001 and LQ19B050003), Zhejiang Provincial Ten Thousands Talents Program (2018R52005), and the Key Construction Project (2017XM022) of Zhejiang Normal University.
References (52)
- et al.
Myeloperoxidase: a new player in autoimmunity
Cell Immunol
(2017) - et al.
A fluorescent probe for the detection of myeloperoxidase activity in atherosclerosis-associated macrophages
Chem Biol
(2007) - et al.
Modeling the reactions of superoxide and myeloperoxidase in the neutrophil phagosome: implications for microbial killing
J Biol Chem
(2006) - et al.
Identification of proteins susceptible to thiol oxidation in endothelial cells exposed to hypochlorous acid and N-chloramines
Biochem Biophys Res Commun
(2012) - et al.
Macrophage myeloperoxidase regulation by granulocyte macrophage colony-stimulating factor in human atherosclerosis and implications in acute coronary syndromes
Am J Pathol
(2001) - et al.
Chlorinative stress: an under appreciated mediator of neurodegeneration
Cell Signal
(2007) - et al.
Bioanalytical methods for hypochlorous acid detection: recent advances and challenges
Trac Trends Anal Chem
(2018) - et al.
A unique iridium(III) complex-based chemosensor for multi-signal detection and multi-channel imaging of hypochlorous acid in liver injury
Biosens Bioelectron
(2017) - et al.
A water-soluble BODIPY-based fluorescent probe for rapid and selective detection of hypochlorous acid in living cells
Spectrochim Acta, Part A
(2019) - et al.
A solvent depend on ratiometric fluorescent probe for hypochlorous acid and its application in living cells
Dyes Pigments
(2017)
Mitochondria-targeted fluorescent probes for oxidative stress imaging
Sens Actuators, B
Toward rapid and selective detection of hypochlorous acid in pure aqueous media and its application to cell imaging: BODIPY-derived water-soluble macromolecular chemosensor with high sensitivity
Dyes Pigments
A dual-channel and fast-response fluorescent probe for selective detection of HClO and its applications in live cells
Sens Actuators, B
Mitochondria-targeted fluorescence probe for endogenous hypochlorite imaging in living cells and zebrafishs
Sens Actuators, B
A lysosome-targetable and ratiometric fluorescent probe for hypochlorous acid in living cells based on a 1,8-naphthalimide derivative
Spectrochim Acta, Part A
A review of NIR dyes in cancer targeting and imaging
Biomaterials
Deformylation reaction-based probe for in vivo imaging of HOCl
Chem Sci
Antioxidant enzynle statusin biliary obstructed rats: efects of N-acetylcysteine
J Hepatol
Oxidative stress, dysfunctional glucose metabolism and Alzheimer disease
Nat Rev Neurosci
Recent progress in the development of fluorescent, luminescent and colorimetric probes for detection of reactive oxygen and nitrogen species
Chem Soc Rev
Specificity of a third kind: reactive oxygen and nitrogen intermediates in cell signalling
J Clin Invest
Luminescent chemodosimeters for bioimaging
Chem Rev
Dynamic and reversible fluorescence imaging of superoxide anion fluctuations in live cells and in vivo
J Am Chem Soc
The common biology of cancer and ageing
Nature
Evidence for rapid Inter- and intramolecular chlorine transfer reactions of histamine and carnosine chloramines: implications for the prevention of hypochlorous-acid-mediated damage
Biochemistry
Characterization of the oxidation products of the reaction between reduced glutathione and hypochlorous acid
Biochem J
Cited by (26)
A novel hepatocyte-targeting fluorescent probe of rhodamine analogues to detect hypochlorite in lysosomes: Design, synthesis and evaluation
2024, Journal of Photochemistry and Photobiology A: ChemistrySynthesis and application of hypochlorite ratiometric fluorescence probe based on cellulose
2024, International Journal of Biological MacromoleculesLysosome-targeting benzothiazole-based fluorescent probe for imaging viscosity and hypochlorite levels in living cells and zebrafish
2022, Spectrochimica Acta - Part A: Molecular and Biomolecular SpectroscopyCitation Excerpt :Hypochlorite (OCl-) is an important reactive oxygen species (ROS) produced in myeloperoxidase (MPO)-promoted reactions between hydrogen peroxide and chloride ions in living systems. [1–2]