Multiple equilibria of phenothiazine dyes in aqueous cyclodextrin solutions

The dimerization and inclusion complexation equilibria of six phenothiazine (PN) dyes with cyclodextrins (alpha-, beta-, and gamma-CDs) in aqueous media have been studied using absorption and fluorescence spectroscopy. The PN dyes used in this study are thionine (TH), azure A (AZA), methylene blue (MB), toluidine blue (TB), new methylene blue (NMB), and 1,9-dimethylmethylene blue (DMMB). The dimerization constants (K-D) of the dyes having two methyl substituents at the phenothiazine ring (NMB and DMMB) are much greater than those of other dyes having unsubstituted rings, and the presence of methyl groups on the amine groups affects little the K-D values. The positions of the monomer/dimer equilibria do not change with the presence of alpha-CD, while the addition of beta-CD suppresses and gamma-CD enhances the dimerization of the dyes except DMMB. The equilibrium constants for the inclusion complexation of the dye monomers and dimers with CDs are determined from the analysis of the dependence of the absorption spectra of the dye solutions on the concentrations of the CDs using a multiple-equilibrium scheme. The results indicated that, except DMMB, which has methyl groups at the 1-position of the fused phenothiazine ring, the dye monomers fit better to beta-CD and the dimers fit snugly to gamma-CD. The DMMB monomer is too large to fit in beta-CD but forms stable complexes with gamma-CD, It appears that the inclusion complexes of the dye monomers and dimers are formed by deep insertion of the phenothiazine rings into the cavities of the CDs, with the 2-methyl groups (in TB and NMB) and the amine groups protruding from the cavities of the CDs. Fluorescence spectroscopic studies indicate that the dye dimers are not fluorescent and inclusion of the monomer in beta-CD results in a 3-5 times enhancement of fluorescence intensity. The determined equilibrium constants of the multiple-equilibrium scheme of the dyes in CD media and fluorescent properties of the dyes can be used to control the dye aggregation and the photophysical and photochemical properties of the phenothiazine dyes for various applications.