Controllable Nitric Oxide (NO) Donor NO-Rosa5 | Funakoshi

NO donor, controlled by visible light
Controllable NO Donor : NO-Rosa5

Nitric oxide (NO), a gaseous free radical, is one of the signaling molecules produced by NO synthases physiologically. NO plays a key role in a wide range of biological processes.
However, NO is extremely unstable in physiological condition, so it is difficult to handle NO molecule in biological experiments.
This made all tough to analyze further function of NO in cells.
To overcome this point, "NO donors" which are compounds releasing NO molecules in the aqueous buffers have been used in NO research. Although NO donors are valuable tools, NO donors are not controllable spatio-temporally.

Controllable NO donor (original name; NO-Rosa5) is a novel NO donor controllable by visible light.
Controllable NOdonor has two moieties, rosamine fluorescent dye as light harvesting group and N-nitrosoaminophenol structure as NO releasing moiety and releases NO molecules triggered by photoinduced electron transfer (PeT).
It shows low photo-toxicity, low cytotoxicity, and efficient NO releasing by yellowish green light irradiation.

This product has been commercialized with the support of Nagoya City University.

Due to Controllable NOdonor releasing NO molecule upon 500-600 nm photo-irradiation, fluorophores, including fluorescent dyes, fluorescent reagent, and fluorescent proteins which are excitable by 500-600 nm wavelength are not available.
Furthermore, Controllable NOdonor itself shows red fluorescent emission. When customers use any fluorophore with Controllable NOdonor, please refer to following information.

(1) Fluorophores not recommended
- Excitation with 500-600 nm such as red fluorophores (for example Cy3, TAMRA, RFP protein etc.)

(2) Suitable fluorophores
- Excitation with <500 nm such as blue fluorophores (for example coumarin dye, BFP protein etc.) or green fluorophores (for example fluorescein, GFP protein etc.)
- Excitation with >600 nm such as near infrared red or far red fluorophores (for example Cy5, Cy7 etc.)

Fig.1 In cellullo region-specific NO generation with Controllable NOdonor
HEK293T cells were firstly treated with DAF-FM DA (10 uM) for 30 min. After washing, the cells were subsequently treated with 10 uM Controllable NOdonor for 60 min.
The dish was photoirradiated inside the indicated white circle (r=31 um) using a 543 nm laser of confocal microscopy.
Left; before irradiation, Right; after irradiation.
Only inside the white circle, NO was detected. (scale bar = 40 um)

Fig.2 Visible light-induced NO releasing in vitro
Quantitative analysis of NO release from Controllable NOdonor with NO electrode. Controllable NOdonor (final 10uM) in 100 mM HEPES buffer was irradiated with a MAX-303 (Asahi Spectra)equipped with a 530-590 nm band-pass filter (light intensity: 70 mW/cm2) .
Left: Continuous irradiation. Right: Short pulse irradiation.

1. Ieda et al., Sci. Rep., 9, 1430 (2019)
Structure-efficiency relationship of photoinduced electron transfer-triggered nitric oxide releasers.
2. Ieda et al., Chem. Pharm. Bull., 67, 576-579 (2019) ,
In cellullo and ex vivo availability of yellowish-green-light-controllable NO releaser.
3. Okuno et al., Org. Biomol. Chem., 15, 2791-2796 (2017)
A yellowish-green-light-controllable nitric oxide donor based on N-nitrosoaminophenol applicable for photocontrolled vasodilation.