betadex and harman

Harman, a natural β-carboline alkaloid, has recently gained considerable interest due to its anticancer properties. However, its physicochemical characteristics and poor oral bioavailability have been limiting factors for its pharmaceutical development. In this paper, we described the complexation of harman (HAR) with β-cyclodextrin (βCD) as a promising alternative to improve its solubility and consequently its cytotoxic effect in chemoresistant melanoma cells (A2058 cell line). Inclusion complexes (βCD-HAR) were prepared using a simple method and then characterized by FTIR, NMR and SEM techniques. Through in silico studies, the mechanism of complexation of HAR with βCD was elucidated in detail. Both HAR and βCD-HAR promoted cytotoxicity, apoptosis, cell cycle arrest and inhibition of cell migration in melanoma cells. Interestingly, complexation of HAR with βCD enhanced its pro-apoptotic effect by increasing of caspase-3 activity (p < 0.05), probably due to an improvement in HAR solubility. In addition, HAR and βCD-HAR sensitized A2058 cells to vemurafenib, dacarbazine and 5FU treatments, potentializing their cytotoxic activity. These findings suggest that complexation of HAR with natural polymers such as βCD can be useful to improve its bioavailability and antimelanoma activity.

Topics: Antineoplastic Agents; Apoptosis; beta-Cyclodextrins; Cell Line, Tumor; Cell Movement; Cell Survival; Drug Resistance, Neoplasm; Harmine; Humans; Melanoma; Molecular Dynamics Simulation; Mutation; Proto-Oncogene Proteins B-raf; Skin Neoplasms

Photoinduced proton transfer reactions of harmane (1-methyl-9H-pyrido[3,4-b]indole) (HAR) in the presence of a proton donor/acceptor such as dihydrogen phosphate anions in aqueous solution have been studied by stationary and time-resolved fluorescence spectroscopy. The presence of high amounts of dihydrogen phosphate ions modifies the acid/base properties of this alkaloid. Thus, by keeping the pH constant at pH 8.8 and by increasing the amount of NaH(2)PO(4) in the solution, it is possible to reproduce the same spectral profiles as those obtained in high alkaline solutions (pH >12) in the absence of NaH(2)PO(4). Under these conditions, a new fluorescence profile appears at around 520 nm. This result could be related to the results of a recent investigation which suggests that a high intake of phosphates may promote skin tumorigenesis. The presence of β-cyclodextrin (β-CD) avoids the proton transfer reactions in this alkaloid by means the formation of an inclusion complex between β-CD and HAR. The formation of this complex originates a remarkable enhancement of the emission intensity from the neutral form in contrast to the cationic and zwitterionic forms. A new lifetime was obtained at 360 nm (2.5 ns), which was associated with the emission of this inclusion complex. At this wavelength, the fluorescence intensity decay of HAR can be described by a linear combination of two exponentials. From the ratio between the pre-exponential factors, we have obtained a value of K = 501 M for the equilibrium of formation of this complex.

Topics: Anions; beta-Cyclodextrins; Energy Transfer; Fluorescence; Harmine; Hydrogen-Ion Concentration; Kinetics; Molecular Structure; Phosphoric Acids; Photochemical Processes; Protons; Quantum Theory; Solutions; Spectrometry, Fluorescence; Thermodynamics; Water

The presence of cyclodextrins (CDs) in the mobile phase alters the chromatographic equilibria and induces a secondary chemical equilibrium associated to the chromatographic separation by HPLC. In this study the influence of the presence of CDs in the mobile phase as chemically modified beta-CDs, i.e. 2,3-di-O-methyl-beta-cyclodextrin (DMbeta-CD) and 2,3,6-tri-O-methyl-beta-cyclodextrin (TMbeta-CD) on the separation of the alkaloids norharmane, harmane and harmine is described. These beta-carboline alkaloids are chemically and structurally related and their quantitation by RP-HPLC is of interest due to their biological and pharmacological properties. Two stationary phases (methyl-, C(1) and octadecyl-, C(18)) were employed, with methanol:buffered aqueous solution and ethanol:buffered aqueous solution as mobile phases. The role of tert-butyl alcohol as a mobile phase modifier and also as an inclusion complex stabiliser was also considered. The concentrations of DMbeta-CD and TMbeta-CD vary from 0 to 17 mM. The presence of increasing amounts of CDs in the mobile phase reduces the retention factor. The changes observed in the retention factor allow the determination of the alkaloid/CD apparent association constants, whose magnitude is influenced by the chemical and structural properties of the guest molecules but also by the composition of the mobile phase. Assuming a 1:1 stoichiometry for the inclusion complexes, the apparent association constants obtained were higher for norharmane and for both DMbeta-CD and TMbeta-CD. The strength of the complexes is higher for DMbeta-CD than for TMbeta-CD and this behaviour can be explained considering the steric problems associated to the permethylated-beta-CDs. Besides significant differences in the magnitude of the apparent association constants were observed for the two stationary phases employed and thus can be related to the adsorption of CDs on the stationary phase. A significant reduction in the proportion of organic solvent in the mobile phase (50%) without a decrease in the selectivity or resolution of the separation is a favourable consequence of the presence of the CDs in the mobile phase.

Topics: beta-Cyclodextrins; Carbolines; Chromatography, High Pressure Liquid; Harmine; Indicators and Reagents; Reference Standards; Solvents; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet