Application of commercially available fluorophores as triplet spin probes in EPR spectroscopy

In recent years, photoexcited molecular triplet states became increasingly popular in magnetic resonance, e.g. as spin probes to measure distances relative to other electron-paramagnetic species or as moieties that transfer light-generated electron–spin polarisation of the triplet state to surrounding magnetic nuclei. In this study, the triplet states of three commercially available dyes, Erythrosin B, Rose Bengal and Atto Thio 12, all typically utilised as fluorophores in optical spectroscopies and microscopies, are investigated in aqueous solutions by using transient absorption spectroscopy and transient electron paramagnetic resonance (EPR). From these methods, the triplet-state lifetimes as well as their zero-field splitting parameters, D and E, which reflect the electronic structures of the triplet state wavefunctions, were obtained. Atto Thio 12 exhibits much smaller D and E values as compared to Rose Bengal and Erythrosin B. On the basis of density functional theory calculations of the triplets’ energy splittings at zero magnetic field, these findings were rationalised. As a proof of concept for applications, the triplet-state properties of Atto Thio 12 bound to an aptamer were also determined and the results are discussed.