Development of Novel Fluorescence Probes That Can Reliably Detect Reactive Oxygen Species and Distinguish Specific Species
Open accessAbstract
We designed and synthesized 2-[6-(4'-hydroxy)phenoxy-3H-xanthen-3-on-9-yl]benzoic acid (HPF) and 2- [6-(4'-amino)phenoxy-3H-xanthen-3-on-9-yl]benzoic acid (APF) as novel fluorescence probes to detect selectively highly reactive oxygen species (hROS) such as hydroxyl radical (*OH) and reactive intermediates of peroxidase. Although HPF and APF themselves scarcely fluoresced, APF selectively and dose-dependently afforded a strongly fluorescent compound, fluorescein, upon reaction with hROS and hypochlorite ((-)OCl), but not other reactive oxygen species (ROS). HPF similarly afforded fluorescein upon reaction with hROS only. Therefore, not only can hROS be differentiated from hydrogen peroxide (H(2)O(2)), nitric oxide (NO), and superoxide (O2*-) by using HPF or APF alone, but (-)OCl can also be specifically detected by using HPF and APF together. Furthermore, we applied HPF and APF to living cells and found that HPF and APF were resistant to light-induced autoxidation, unlike 2',7'-dichlorodihydrofluorescein, and for the first time we could visualize (-)OCl generated in stimulated neutrophils. HPF and APF should be useful as tools to study the roles of hROS and (-)OCl in many biological and chemical applications.
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The primary reactive species detected in our paradigm was ONOO−.
“Given the sensitivity of CM-H2DCFDA to oxidation by ONOO−, in combination with the propensity of ONOO− formation attributable to Aβ-induced stress, we reasoned that the primary RS detected in our paradigm was ONOO− (Setsukinai et al., 2003; Malinski, 2007).”
From The mitochondria-targeted antioxidant MitoQ prevents loss of spatial memory retention and early neuropathology in a transgenic mouse model of Alzheimer’s disease (2011) · cites this paper as Setsukinai et al., 2003
Table I reports a strong fluorescence increase when DCFH was exposed to ONOO−, supporting the peroxynitrite-sensitivity premise. The paper tested DCFH prepared from DCFH-DA, not CM-H2DCFDA, so it does not directly establish that probe’s sensitivity. The identification of ONOO− as the primary species in the citing authors’ paradigm is their own inference, not a finding attributed to this source. (LLM, conf=0.84)
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- Authors
- Setsukinai, K.-I., Urano, Y., Kakinuma, K., Majima, H. J., & Nagano, T.
- Affiliations
- The University of Tokyo (JP)
- Kagoshima University (JP)
- Corresponding author
- Nagano, Tetsuo
- Venue
- Journal of Biological Chemistry
- Publisher
- Elsevier BV
- Published
- 2003-01
- Volume
- 278
- Issue
- 5
- Pages
- 3170-3175
- ISSN
- 0021-9258 (ISSN-L)
- OpenAlex
- W1998199040
- Cited by
- 1,301
- References
- 1
- Citations by year
- 2026: 61 · 2025: 54 · 2024: 45 · 2023: 54 · 2022: 61 · 2021: 58 · 2020: 61 · 2019: 64 · 2018: 58 · 2017: 66 · 2016: 60 · 2015: 78 · 2014: 85 · 2013: 89 · 2012: 76
- FWCI
- 9.36
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Journal of Biological Chemistry (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Molecular Sensors and Ion Detection
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Redox biology and oxidative stress
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
- Keywords
- fluorescent probes, reactive oxygen species, hydroxyl radicals, hypochlorite, xanthenone derivatives, fluorescein, neutrophils, live-cell imaging, superoxide, hydrogen peroxide, nitric oxide
- MeSH terms
- Fluorescent Dyes, Carcinoma, Hepatocellular, Humans, Light, Liver Neoplasms, Microscopy, Fluorescence, Magnetic Resonance Spectroscopy, Oxidation-Reduction, Spectrometry, Fluorescence, Superoxide Dismutase, Tumor Cells, Cultured, Drug Design, Reactive Oxygen Species
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