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Small Molecules (Tool Compounds)

AzuFluorTM 483-Bpin: Azulene-Based Fluorescent Probe for ROS/RNS small molecule (tool compound)

Invented by Lloyd Murfin , Tony James , Simon Lewis
Invented at University Of Bath
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Image thumbnail for AzuFluor<sup>TM</sup> 483-Bpin: Azulene-Based Fluorescent Probe for ROS/RNS small molecule (tool compound)
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Figure S5. Fluorescence emission spectrum of 2 at 500 nM in different solvents, λmax = 480, 482, 487, and 491 nm for acetone, MeOH, MeCN, and DMSO respectively. Fluorescence emissions were measured with λex = 350 (ex. slit 10, em. slit 20) nm on an Agilent Technologies Cary Eclipse Fluorescence Spectrometer.
Figure 5. Selectivity data for 1 (500 nM) in the presence of ONOO−, H2O2, HOCl, OH•, O2•−, 1O2, and ROO• (each 500 nM). Data acquired 5 min after mixing, in PBS buffer 52% H2O: MeOH, pH 8.2 at 25 °C, at λmax = 483 nm. Fluorescence intensities were measured with λex = 350 (bandwidth 20) nm, on a BMG Labtech CLARIOstar plate reader.
Figure 6. TPM images of RAW 264.7 macrophages stained with 1 (5 μM, 30 min). (a) Control image. (b−h) Cells were pretreated with (b) exogenous added H2O2 (2 mM, 30 min), (c) exogenous added ONOO− (100 μM, 30 min), (d) PMA (1 μg mL−1, 30 min), (e) LPS (1 μg mL−1, 4 h) and IFN-γ (50 ng mL−1, 1 h), (f) LPS, IFN-γ and ebselen (50 μM, 30 min), (g) SIN-1 (50 μM, 30 min), and (h) SIN-1 and ebselen. (i) Average fluorescence intensity in the corresponding TPM images. Images were obtained using 800 nm as the excitation wavelength and 400−600 nm emission windows. Scale bars = 20 μm. Asterisks represent statistical significance (***p < 0.001) and n is the number of counted cells.
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Catalogue Number 157848
Antigen/Gene or Protein Targets Reactive Oxygen and Nitrogen Species
Synonyms AzuFluorTM 483-Bpin, Diethyl 2-amino-6-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)azulene-1,3- dicarboxylate
Type Fluorescent Probe
Relevance Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are important mediators in many physiological and pathological processes.

The one-electron reduction of O2 in vivo leads to the formation of O2•− (superoxide), which in turn undergoes disproportionation catalyzed by superoxide dismutase to give O2 and H2O2 (hydrogen peroxide). Another fate of O2•− is to react with endogenously produced NO (nitric oxide) to form ONOO− (peroxynitrite) via a nonenzymatic process. Ordinarily, the flux of H2O2 is tightly regulated; aberrant H2O2 production or overexposure is implicated in the pathogenesis of many diseases such as cancer and neurodegenerative conditions.
Similarly, while ONOO− has roles in signal transduction, its strongly oxidizing and nitrating properties mean it can react in an uncontrolled manner with various biomolecules. Elevated levels of ONOO− have been linked to cardiovascular, neurodegenerative, and inflammatory diseases as well as cancer.

In view of this, there is significant need for tools and techniques to elucidate the roles of ONOO−, H2O2, and other ROS/RNS in biological systems.
Molecular Formula C22H28BNO6
Key Attributes AzuFluorTM 483B-pin shows high selectivity for peroxynitrite and hydrogen peroxide over other reactive oxygen species. It is cell-permeable, photostable, and noncytotoxic in vitro on the time scale of the two-photon fluorescence microscopy experiments.
Molecular Weight (g/mol) 412.2046
Solubility DMSO, MeCN, Acetone, MeOH
Research Area Cancer, Cardiovascular, Neurobiology, Other
Storage ambient
Notes pH: Fluorescence intensity greatest at pH 7-8.

Optimal two-photon excitation wavelength λex = 800nm.

References

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References: 1 entry

Murfin et al. 2019. J Am Chem Soc. 141(49):19389-19396. PMID: 31773957.

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References: 1 entry

Murfin et al. 2019. J Am Chem Soc. 141(49):19389-19396. PMID: 31773957.

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Inventor Information

Inventors

Lloyd Murfin

Lloyd Murfin

Tony James

Tony James

Simon Lewis

Simon Lewis

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Invented at

University Of Bath
University Of Bath

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