**Conflict of interest**

*Phytochemicals in Human Health*

**Coumarin Ex/Em** 

1-(4-(7-(diethylamino)-4-methylcoumarin-3-yl)phenyl)-1H-pyrrole-2,5-dione

7-(diethylamino)-N-(2-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)ethyl)coumarin-3-carboxamide

*Fluorescent coumarin maleimides used for biomolecule labelling.*

tion of a phenolate anion [115].

**3. Concluding remarks**

**Table 5.**

imaging techniques.

**Acknowledgements**

grant SFRH/BD/128807/2017.

hydroxyl groups do not allow highly specific labelling reactions due to the existence of several hydroxyl groups in biomolecules (serine, threonine, and tyrosine) [113]. One of the well-known labelling methods is the reaction with diazonium salts

**(nm)**

**Physicochemical features and biological applications**

groups in tissue sections. Fluorescence probe for glutathione intact cells Used to monitor release of thiols, to quantitate thiol in microplate reactions, and to distinguish proliferating cancer cells by nuclear protein staining

biological sensing device and for real-time measurements for the release of inorganic phosphates during enzymatic reaction. Also, used for intramolecular fluorescence energy transfer (FRET) experiments

391/472 Labelling of protein thiol

419/467 Used as a fluorescent

**Reference**

[68, 105]

[68, 110–112]

Reactive fluorescent coumarins have been increasingly attracting special interest as fluorescent labels, with a wide range of applications in bioimaging and biolabelling, due to their extremely attractive and stable scaffold. Coumarins will allow the development of new low-cost fluorescent dyes due to its easy synthesis with high yields, large Stokes shift, pH independence of absorbance and emission, and excellent photostability, which represents a great value for the biological fluorescence

The authors thank the European Regional Development Fund ALENTEJO 2020

through the project "MEDUSA-Microorganisms Monitoring and Mitigation— Developing and Unlocking novel Sustainable Approaches (ALT20-03-0145- FEDER-000015)" for financial support. Sérgio Martins acknowledge Fundação para a Ciência e a Tecnologia (FCT) for the economic support through the doctoral

resulting in the formation of azo compounds, as 4-trifluoromethylcoumarin-7-diazonium chloride [114]. Although these aryl diazonium ions are promising for the desired application, their storage and delivery are challenging, and they often require in situ generation. The pH range should be between 8 and 10 for the forma-

**168**

There are no conflicts of interest to declare.
