RESUMEN
Thermofluorochromism and photochromism of salicylideneaniline (SA) in alcohol were investigated using steady-state and time-resolved fluorescence and absorption spectroscopy. The planar trans-enol form of SA in alcohols is converted into the twisted trans-enol form on heating. This conversion results change in the emission maximum from the 530 to the 440 nm region with an increase in fluorescence intensity, which confirms the absence of intramolecular hydrogen bonding between imine nitrogen and phenolic hydrogen in the twisted trans-enol form. The activation barrier for thermal-induced formation of the twisted trans-enol form in methanol was determined experimentally and was found to be 20.15 ± 2.22 kcal/mol. The rotation of the phenolic C7-C8 and C7-N1 bond followed by breaking of the intramolecular hydrogen bond and formation of an intermolecular hydrogen bond with alcohol solvent molecules results in the thermally stable twisted trans-enol form in alcohol solvents. The biexponential nature of the fluorescence decay of the twisted trans-enol form of SA confirms that the emission originates from multiple (π-π* and n-π*) excited states. On photolysis under UV light, the twisted trans-enol form is converted back into the planar trans-enol form. The time-resolved absorption and excitation-resolved fluorescence spectrum of SA in methanol confirm the existence of the twisted cis-keto form as a transient photochromic intermediate in the light-induced planarization of SA in alcohols. In alcohols, an interplay between the intra- and intermolecular hydrogen-bonding controls excited-state reaction dynamics and conformational relaxation of SA, which are responsible for the photochromism of salicylideneaniline.
RESUMEN
Excitation-resolved area-normalized emission spectroscopy (ERANES) is proposed as a new steady-state fluorescence technique for the investigation of heterogeneous fluorescence (HGF) from a mixture of fluorophores and fluorophores present in various environments and proteins. The presence of a single isoemissive point was used to confirm the presence of two absorbing and emitting species in the system. The isoemissive point was found to occur at the wavelength where the ratio of wavelength dependent fluorescence quantum yield of the emissive species equals to the ratio of their total fluorescence quantum yield. The application of the ERANES method for resolving HGF from a mixture of fluorophores having similar or different fluorescence lifetimes with a relatively high degree of fluorescence spectral overlap was demonstrated. When compared to excitation fluorescence (EF) matrix and time-resolved methods, ERANES was found to be a simple analytical method for analyzing HGF from a mixture of fluorophores, and from fluorophores present in heterogeneous media, such as cells, membranes, etc., and for analyzing protein fluorescence, without the requirement for sophisticated instrumentation and data analysis.