Study to Probe Subsistence of Host-Guest Inclusion Complexes of α and ß-Cyclodextrins with Biologically Potent Drugs for Safety Regulatory Dischargement.

Host-guest interaction of two significant drugs, phenylephrine hydrochloride and synephrine with α and ß-cyclodextrins were studied systematically. Initially two simple but reliable physicochemical techniques namely conductance and surface tension were employed to find out saturation concentration for the inclusion and its stoichiometry. The obtained 1:1 stoichiometry was further confirmed by two spectrometric methods, UV-Vis study and spectrofluorimetry. Significant shifts in IR stretching frequency also support the inclusion process. Relative stabilities of the inclusion complexes were established by the association constants obtained from UV-Vis spectroscopic measurements, program based mathematical calculation of conductivity data. Calculations of the thermodynamic parameters dictates thermodynamic feasibility of the inclusion process. Spectrofluorometric measurement scaffolds the UV-Vis spectroscopic measurement validating stability of the ICs once again. Mass spectroscopic measurement gives the molecular ion peaks corresponding to the inclusion complex of 1:1 molar ratio of host and guest molecules. The mechanism of inclusion was drawn by 1H-NMR and 2D ROESY spectroscopic analysis. Surface texture of the inclusion complexes was studied by SEM. Finally, the cytotoxic activities of the inclusion complexes were analyzed and found, Cell viability also balances for non-toxic behavior of the ICs. Moreover, all the studies reveal the formation of inclusion complexes of two ephedra free, alternatively emerging drugs (after their banned product having ephedra) SNP, PEH with α and ß-CD which enriches the drug delivery system with their regulatory release without any chemical modification.

Improvement of Cellular Uptake and Transfection Ability of pDNA Using α-Cyclodextrin-Polyamidoamine Conjugates as Gene Delivery System.

Polyamidoamine (PAMAM) dendrimers are a class of unique nanomaterials which attracted attention because of their extraordinary properties, such as highly branched structure and types of terminal primary groups. In addition, development in PAMAM chemical modification has broadened its biological application especially for drug and gene delivery. In this study, PAMAMs are covalently conjugated onto α-Cyclodextrin (α-CD) via amide bonds obtaining the starburst cationic polymers (CD-PG2). The chemical structure and composition of CD-PG2 was characterized by IH NMR. Physicochemical and biological properties of CD-PG2/pDNA polyplex were evaluated by agarose gel retardation, stability test against DNasecñ, MTT assay, DLS measurement, CLSM observation, LDH leakage test, cellular uptake route analysis and in-vitro cell transfection. Results showed that CD-PG2 can efficiently condense pDNA into nanoscale particles with a narrow size distribution, and protect pDNA form DNase I degradation. Compared with free PEI-25K and commercial product Lipofectamine2000, CD-PG2 shows excellent gene transfection efficiency without serum interference as well as relatively low cytotoxicity. Cellular uptake of CD-PG2/pDNA polyplex is mainly through CME and CvME route and further investigations demonstrate that α-CD can regulate CvME pathway to improve polyplex transfection behavior. In conclusion, CD-PG2 can be considered as a versatile tool for gene delivery, especially for gene transfer in-vivo.

Thiolated α-Cyclodextrin: The Invisible Choice to Prolong Ocular Drug Residence Time.

It was the aim of this study to develop cysteamine-conjugated α-cyclodextrin (α-CD) enabled to form disulfide bonds with cysteine-rich substructures of the ocular mucus layer to provide a prolonged residence time of incorporated drugs at the site of action. Cysteamine was covalently attached to oxidized α-CD via reductive amination. The resulting α-CD-cysteamine conjugates (α-CD-Cys) were characterized regarding the amount of free thiol groups attached to the oligomer backbone via Ellman's reagent; resazurin assay was conducted for cytotoxicity, and mucoadhesive properties were evaluated on porcine intestinal and ocular mucosal tissues. Furthermore, albino rabbits were used for assessing the irritation-masking effects of α-CD-Cys. Free thiol groups attached to the backbone were in the range of 558 ± 24-1143 ± 92 µmol/g. None of these α-CD-Cys unduly affected the viability of Caco-2 cells in a concentration of 0.5%. Mucoadhesive properties of α-CD-Cys were up to 32-fold improved compared to unmodified α-CD. Encapsulation of cetirizine into α-CD-Cys resulted in significantly reduced local ocular mucosal irritation of this model drug. According to these results, α-CD-Cys is a promising new tool to prolong drug residence time on the ocular mucosal surface.

Evaluation of the potential toxicity of unmodified and modified cyclodextrins on murine blood-brain barrier endothelial cells.

In this study, we investigated the cytotoxic effects of unmodified α-cyclodextrin (α-CD) and modified cyclodextrins, including trimethyl-ß-cyclodextrin (TRIMEB) and hydroxypropyl-ß-cyclodextrin (HPßCD), on immortalized murine microvascular endothelial (cEND) cells of the blood-brain barrier (BBB). A CellTiter-Glo viability test, performed on the cEND cells showed significant differences among the different cyclodextrins. After 24 hr of incubation, TRIMEB was the most cytotoxic, and HPßCD was non-toxic. α-CD and TRIMEB exhibited greater cytotoxicity in the Dulbecco's modified Eagle's medium than in heat-inactivated human serum indicating protective properties of the human serum. The predicted dynamic toxicity profiles (Td) for α-CD and TRIMEB indicated higher cytotoxicity for these cyclodextrins compared to the reference compound (dimethylsulfoxide). Molecular dynamics simulation of cholesterol binding to the CDs suggested that not just cholesterol but phospholipids extraction might be involved in the cytotoxicity. Overall, the results demonstrate that HPßCD has the potential to be used as a candidate for drug delivery vector development and signify a correlation between the in vitro cytotoxic effect and cholesterol binding of cyclodextrins.

Effects of dendrimer/cyclodextrin conjugates as gene transfer carriers on nitric oxide production from macrophages.

OBJECTIVES: The development of safe gene transfer carriers with high transfection efficiency, which does not affect the cell function, is a challenging issue. In this study, we examined the effects of α-cyclodextrin (α-CyD)/dendrimer conjugate (α-CDE (G3)) on nitric oxide (NO) production in murine macrophages J774.1 cells stimulated with toll-like receptors (TLR) ligands. METHODS: NO production from macrophages stimulated with TLR ligands was determined by the Griess method. Transfection efficiency of α-CDE (G3)/plasmid DNA (pDNA) complex was quantified by a luminometer. KEY FINDINGS: α-CDE (G3) significantly inhibited NO production from J774.1 cells stimulated with TLR ligands. α-CyD molecules in α-CDE (G3) had no effect on NO production. The inhibitory effect of α-CDE (G3) on NO production might be attributed to the dendrimer (G3). Increasing the degree of substitution (DS) of α-CyD in the α-CDE (G3) molecule was accompanied by a significant decrease in the inhibition of NO production. Furthermore, higher gene transfection efficiency of α-CDE (G3)/pDNA complex was observed upon increasing the DS of α-CyD. CONCLUSIONS: α-CDE (G3) with high DS value of α-CyD may be considered as a safe gene transfer carrier that does not adversely affect NO production from macrophages stimulated with TLR ligands.

Evaluation of the Cytotoxicity of α-Cyclodextrin Derivatives on the Caco-2 Cell Line and Human Erythrocytes.

Cyclodextrins, even the 6-membered α-cyclodextrin, are approved in the various pharmacopoeias as pharmaceutical excipients for solubilizing and stabilizing drugs as well as for controlling drug release. Recently α-cyclodextrin has also been marketed as health food with beneficial effects on blood lipid profiles. However, the concentration of α-cyclodextrin used may be very high in these cases, and its toxic attributes have to be seriously considered. The objective of this study was to investigate the cytotoxicity of various, differently substituted α-cyclodextrin derivatives and determine relationship between the structures and cytotoxicity. Three different methods were used, viability tests (MTT assay and Real Time Cell Electronic Sensing on Caco-2 cells) as well as hemolysis test on human red blood cells. The effect of α-cyclodextrin derivatives resulted in concentration-dependent cytotoxicity, so the IC50 values have been determined. Based on our evaluation, the Real Time Cell Electronic Sensing method is the most accurate for describing the time and concentration dependency of the observed toxic effects. Regarding the cytotoxicity on Caco-2 cells, phosphatidylcholine extraction may play a main role in the mechanism. Our results should provide help in selecting those α-cyclodextrin derivatives which have the potential of being used safely in medical formulations.