Sonochemical synthesis, optical properties and DFT studies on novel (N-arylamino)phenothiazinium dyes suitable for fluorescence cells imaging

• Published in: Spectrochimica acta. Part A, Molecular and biomolecular spectroscopy Molecular and biomolecular spectroscopy, 2024-12, Vol.322, p.124768, Article 124768
• Main Authors: Gal, Melinda, Gaina, Luiza, Lovasz, Tamas, Gal, Emese, Craciun, Ana-Maria, Focsan, Monica, Turza, Alexandru, Rugina, Dumitrita, Brânzanic, Adrian M.V., Pesek, Szilard, Silaghi-Dumitrescu, Radu, Cristea, Castelia
• Format: Article
• Language: English
• Subjects: DFT; FLIM; Methylene blue; Sonochemistry; UV–vis spectroscopy
• ISSN: 1386-1425
• DOI: 10.1016/j.saa.2024.124768

[Display omitted] •Sonochemical synthesis of methylene blue analogues.•X-Ray Diffraction analysis of 3,7-bis(N-(meta-bromoaryl)amino)phenothiazinium dye.•UV–vis absorption/emission properties of N-(arylamino)phenothiazinium cationic dyes.•Absorption/emission properties by TD-DFT using B3PW91 versus CAM-B3LYP functionals.•Fluorescence Imaging of N-(arylamino)phenothiazinium cationic dyes. Novel (N-arylamino)phenothiazinium dyes containing meta-substituted-arylamine auxochrome units were successfully obtained by applying a sonochemical protocol designed for a more efficient energy usage in the preparation of methylene blue (MB) analogues. Single crystal X-ray diffraction analysis revealed the spatial arrangement in aggregated crystalline state of (N-(meta-bromoaryl)amino)phenothiazinium dye with minor variances induced by the nature of the halogenide counterion (iodide or chloride). The optical UV–vis properties of the novel (N-arylamino)phenothiazinium dyes were comparable to those of the parent MB, with the longest wavelength absorption maxima situated in the visible range (640–680 nm), large molar extinction coefficients (log ε = 4.5–5.1) and weak solvatochromism in polar solvents. Their fluorescence emission in solid state was evidenced by One Photon Excited Fluorescence Lifetime Imaging (OPE-FLIM) and Two Photon Excited Fluorescence Lifetime Imaging (TPE-FLIM) experiments. Theoretical calculations based on Time Dependent-Density Functional Theory (TD-DFT) at B3PW91 and CAM-B3LYP/def2-SV(P) level of theory predicted absorption and fluorescence emission wavelength maxima in reasonable agreement with experimental data. Computational results suggest that the electronic excitations imply a departure from the planar molecular ground state towards geometrically rearranged excited states disfavoring the vibronic couplings due to a high degree of flexibility induced by the conformational motion of the N-arylamino auxochromes. Preliminary studies regarding the dyes’ relevance in biological environment indicated lipophilicity (log P octanol/water 0.5–2.3), no aggregation tendency in diluted solutions in the concentration range 10–50 microM and ability for cytoplasmatic staining of D407 human retinal pigment epithelial cells.