betadex and 7-8-dimethylalloxazine

We report on photophysical studies of lumichrome (Lc) in water at different pHs, and interacting with the human serum albumin (HSA) protein and β-cyclodextrin (β-CD) in neutral aqueous solutions. We used steady-state and picosecond time-resolved emission spectroscopy to investigate the structural changes of Lc at the ground and excited states, as well as the rotational dynamics of the complexes with HSA and β-CD. In neutral water, the predominant neutral alloxazine-type structure of Lc coexists with a small population of the anionic form. In the presence of HSA, we observed an increase in the absorption band intensity at 450 nm. This increase is due to a preferential complexation (1:1 stoichiometry, K=8600 M(-1)) of the Lc anion structures within the protein. This change is not observed when β-CD is added, in which the Lc neutral form is exclusively complexed, giving a 1:1 stoichiometry. The fluorescence lifetimes of Lc in neutral water solutions are 4.2 and 2.3 ns, assigned to anionic and neutral alloxazinic forms, respectively. Using β-CD, the lifetime of the 1:1 complexes is 0.74 ns, while in the case of HSA complexes we observed two lifetimes (0.83 and 0.14 ns), which we explained in terms of different interactions of the anions with the protein. The rotational relaxation time of free Lc in neutral water is 75 ps. For Lc:β-CD complexes this time is 0.44 ns, in full agreement with the expected value from the hydrodynamic theory. For HSA solutions, we obtained a distribution of values between ∼1 and 4.5 ns, suggesting a site heterogeneity of complexation and a different strength of binding for the involved Lc anionic forms. Our results give information about the different photorelaxation behavior of Lc within chemical and biological cavities, and might help in a better design of nanosystems for drug carriers and delivery.

Topics: Absorption; beta-Cyclodextrins; Flavins; Fluorescence Polarization; Humans; Hydrogen-Ion Concentration; Hydrophobic and Hydrophilic Interactions; Models, Molecular; Molecular Conformation; Serum Albumin; Spectrometry, Fluorescence; Time Factors; Water

Complexation of the model drug lumichrome by 2-hydroxypropyl-beta-cyclodextrin (HPbetaCD), the most widely used cyclodextrin derivative in pharmaceutical preparations, was investigated in this study. The influence of frequently used pharmaceutical excipients, i.e. alcohols (ethanol, glycerol, propylene glycol), buffers (phosphate, citrate) and tonicity modulators (NaCl, MgCl2) was evaluated by phase solubility, absorption and fluorescence emission spectra and fluorescence lifetime studies. Further, complex formation constants and fluorescence quantum yields were calculated. The formation of a 1:1 complex was indicated by phase solubility studies. The shape of the absorption and emission spectra for lumichrome was nearly independent of dissolution medium. The intensity of the absorption peak was slightly decreasing by the addition of HPbetaCD, which indicates formation of an inclusion complex of lumichrome in the ground state. The intensity of the fluorescence emission peak (i.e. fluorescence quantum yield) was also steadily decreasing by the increase in HPbetaCD concentration. Monoexponential fluorescence decay was obtained in the absence of cyclodextrin. In the presence of cyclodextrin, bi-exponential decays were observed in all aqueous vehicles with the exception of plain water or samples containing salts. The longest decay time corresponds to the lifetime of free (uncomplexed) lumichrome, while the shortest decay time was attributed to the excited state of the complexed alloxazine form of lumichrome. The selected excipients influence the complexation constant and the lumichrome excited state deactivation pathways to various extents.

Topics: 2-Hydroxypropyl-beta-cyclodextrin; Absorption; Alcohols; beta-Cyclodextrins; Buffers; Chemistry, Pharmaceutical; Cyclodextrins; Electrolytes; Excipients; Flavins; Fluorescence; Solubility; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet; Water