Ekaterini Antoniadou-Vyza

Antimicrobial activity of β-lactam antibiotics against clinical pathogens after molecular inclusion in several cyclodextrins. A novel approach to bacterial resistance

Recognition and uptake by specific cellular receptors and transport systems for cyclodextrins have been demonstrated. Based on this concept, natural and synthetically modified cyclodextrins were used as drug carriers. Several β‐lactam antibiotics were selected and their inclusion complexes with different cyclodextrins were prepared (molar ratio ranging from 1:1 to 1:3). The complex formation, in aqueous solution, was monitored and optimum complexation conditions were selected. The inclusion of the active molecules in the cyclodextrin cavity was confirmed by 1H NMR spectroscopy. Specific HPLC methods for the quantitation of antibiotics in the presence of cyclodextrins were developed and their chemical stability under complexation conditions was confirmed. Antimicrobial activity of drug‐cyclodextrin complexes, in terms of minimum inhibitory concentration (MIC), were compared with the corresponding values of uncomplexed free molecules. A wide range of clinical pathogens and known β‐lactamase‐producing strains were tested. The activity of the cyclodextrin‐included antibiotics was increased, particularly against Gram‐negative clinical strains. The nature and degree of substitution on cyclodextrin macromolecules may be the predominant factor in the observed improvement in antimicrobial activity. We believe that the proposed methodology is a novel approach to the microbial resistance problem and will trigger research towards the development of new cyclodextrin derivatives bearing the ability to increase the uptake of included antimicrobial molecules through intensification of the corresponding molecular recognition phenomena.

An improved HPLC method overcoming Beer’s law deviations arising from supramolecular interactions in tolfenamic acid and cyclodextrins complexes

Inclusion complexes of tolfenamic acid (TA), a non-steroidal anti-inflammatory drug, with methyl-beta cyclodextrin and hydroxypropyl-beta cyclodextrin were prepared and characterised. Spectrophotometric, chromatographic (RP-HPLC) and 1H NMR studies of the complexes were conducted. It was observed that cyclodextrins influence TAs molar absorptivity leading to Beers law deviation. Consequently, the accuracy problem arose, urged for the application of specific chromatographic conditions for the determination of TA in the presence of CDs. A new HPLC method was developed and validated. TA was analysed on a C18 column 5 microm (150 x 4.6 mm), using a column thermostat regulated at 30 degrees C. The mobile phase consisted of methanol-phosphate buffer solution (pH 3.2; 0.07 M) (90:10 v/v) and the flow rate was set at 2.0 ml min(-1). The detector was operated at 286 nm. TA was successfully determined, overcoming the problems arising from the presence of cyclodextrins.

The self-association of the drug acemetacin and its interactions and stabilization with β-cyclodextrin in aqueous solution as inferred from NMR spectroscopy and HPLC studies

Strongly concentration dependent, (1)H NMR chemical shifts of the non-steroidal anti-inflammatory drug acemetacin sodium salt (sodium [[1-(4-chlorobenzoyl)-5-methoxy-2-methylindol-3-yl]acetoxy]acetate), were observed in aqueous solution. Self-titration and nOe experiments, point to a self-association model where stacking takes place via the indole portion of the drug. In addition, conformational isomerism (atropisomerism) of the anti to syn form was confirmed. Further increase of the concentration eventually led to stable chemical shifts and nearly simultaneous appearance of microcrystals. In the presence of betaCD, 1:1 inclusion complexation occurred through the p-chlorobenzoyl part of the drug, whereas with excess betaCD the indole part seemed to participate to a minor degree. The anti isomer is suggested to be involved in the inclusion process. In addition, aggregation of acemetacin was also evident, as competing with the conformational and inclusion equilibria. The present case demonstrates that many competitive processes are simultaneously active in a seemingly simple system. The measurements were strongly dependent upon the pH and use of buffered solutions was mandatory. Finally, for the quantitative analysis of acemetacin in the presence of betaCD, a special HPLC method was developed. The stability of the drug, studied by the identification of the degradation products and the pseudo-first order rate of hydrolysis, was found to be unaffected by the presence of betaCD.

The formation of an inclusion complex of methocarbamol with hydroxypropyl-β-cyclodextrin: the effect on chemical stability, solubility and dissolution rate

Abstract The inclusion complex of the hydrophobic drug methocarbamol (ML) with hydroxypropyl- β -cyclodextrin (HP β CD) was prepared and characterized in the solid state and in aqueous solution. The prepared complex was studied by chromatographic, spectral and phase solubility methods to determine its structure, solubility, chemical stability and dissolution rate. Host-guest interactions in aqueous solution were studied by proton nuclear magnetic resonance spectroscopy ( 1 H-NMR) and in the solid state by differential scanning calorimetry (DSC) and infrared spectroscopy (IR). The stoichiometry of the isolated complex was determined by reversed phase high pressure liquid chromatography (RP-HPLC), 1 H-NMR spectroscopy and elemental analysis. The solubility and dissolution rate of ML in free and complexed form were examined in aqueous solution. The stability constant of the complex was determined by the classical solubility techniques. The chemical stability of free and complexed ML, in buffered solution at pH 7.4 and at two different temperatures, 37 and 60°C, was monitored using an HPLC method and resulted in a 2-fold increase in the stability of complexed ML compared to free ML.

Original triazine inductor of new specific molecular targets, with antitumor activity against nonsmall cell lung cancer.

Despite our growing insight into carcinogenesis, treatment of tumors, especially nonsmall cell lung cancer (NSCLC), remains limited and it is urgent to develop strategies that target tumor cells and their genetic features. Drug discovery efforts have historically focused on the search for compounds that modulate the protein products of genes. Current drug therapy targets only a few hundred endogenous targets, mainly proteins, such as receptors and enzymes. But now, the interest in specifically targeting RNA is increasing, both for target validation and/or therapeutic purposes. In this regard, our work was concerned with the induction of new molecular targets correlated to a cytostatic effect on NSCLC cell line, after treatment with a new triazin named A190. The in vitro study of cell cycle and apoptosis induction demonstrated the antiproliferative potential of this new compounds, and the use of quantitative RT‐PCR analysis permit to display an original mechanism of action involving 2 genes: HEF1 and B2. The antitumor effect was also confirmed by the good results in vivo on nude mice xenografts.

Competitive complexation and SPE techniques combined with liquid chromatography for the separation and determination of cyclodextrin encapsulated drug substances

SummaryA new analytical procedure, combining sample pretreatment with solid phase extraction and high performance liquid chromatography, was developed and applied to ketoprofen:cyclodextrin (CD) complexes. The significant differences in molar absorptivity between free and CD bound drugs lead to analytical errors up to 30%. Procedures routinely used to remove most of the common excipients proved to be inadequate, creating the necessity of more specific (HPLC) analytical methods including the step of dissociation and separation of drugs from CDs before the quantitation by UV detector. In the case of complexes such as Ketoprofen: CDs, strong binding to CDs inhibits the dissociation. Prior to the analysis, the addition of molecules competing the binding in the CD cavity fairly decreased the strength of the complexation. When molecules, competing the binding to CD such as 1-adamantanol, were added to the samples the results were improved. The extent of the selective binding and the drug displacement were monitored with H-NMR spectroscopy and it was revealed that only partial dissociation of the complexes was obtained. The combination of the sample pre-treatment with a solid phase extraction technique was proved to be inevitable for the dissociation of the complex. When analyzing supramolecules spectral changes, arising from intramolecular and/or intermolecular interactions, are expected. Consequently, analytical methods utilized should be carefully revised.

Supramolecular Interactions Between Tolfenamic Acid and Various Cyclodextrins: Effects of Complexation on Physicochemical and Spectroscopic Data

Tolfenamic acid is a non-steroidal inflammatory drug with associated gastrointestinal sideeffects and problems with bioavailability. Furthermore, it is extremely insoluble in water. Tolfenamic acid was complexed with various cyclodextrins (β-CyD, HP-β-CyD, methyl-β-CyD) in order to circumvent some of these problems. During the characterization of the inclusion complexes with spectroscopic methods, significant variations in the UV and NMR spectra were observed and attributed to the supramolecular interactions between the guest and host molecules. A validated HPLC method was applied to obtain accurate measurements. The complexation process was further examined; the time needed for the completion of the complexation was determined, the binding constants of the complexes were calculated and the energetics of the system were evaluated. The solubility of tolfenamic acid was 30-fold in the presence of HP-β-CyD and twofold in the presence of methyl-β-CyD.

Thermal properties of adamantanol derivatives and their β-cyclodextrin complexes in phosphatidylcholine bilayers

Differential Scanning Calorimetry (DSC) has been applied to study the thermal properties of the membrane perturbing antibacterial octyl- and dodecyl-bromide salts of quaternary dimethylamino adamantanol (ADM-8 and ADM-12 correspondingly) incorporated in free or complexed form with beta-cyclodextrin (beta-CD) into dipalmitoylphosphatidylcholine (DPPC) containing bilayers. The DSC results showed that the studied compounds exert pronounced thermotropic changes in DPPC bilayers when inserted as free molecules. These effects are reduced when are present in a complex form with beta-CD. Since the studied compounds exert destructive effects in membrane bilayers their insertion in membrane bilayers as complexes with cyclodextrin may result in differentiation of their activity. The obtained results suggest that their complexation with beta-CD may improve their biological profile. It also increases their aqueous solubility, a limited factor for their use as drugs.

High-performance liquid chromatography in stability studies of an organophosphorus insecticide in free form and after formulation as emulsifiable conce

Abstract DCPE [α-(diethoxyphosphinoximino)dicyclopropylmethane] is an organophosphorus compound with good insecticidal activity. It is a yellow oily liquid, p

Molecular, crystal and solution structure of a β-cyclodextrin complex with the bromide salt of 2-(3-dimethylaminopropyl)tricyclo[3.3.1.13,7]decan-2-ol, a potent antimicrobial drug

The pharmacological properties of a cyclomaltoheptaose (beta-cyclodextrin) series of adamantane-group-bearing compounds that exhibit potent antibacterial activity have been studied, both isolated and in complex with beta-cyclodextrins (betaCDs). In this work, the structure of the bromide salt of 2-(3-dimethylaminopropyl)-tricyclo[3.3.1.1(3,7)]decan-2-ol(A DM-10) complexed with betaCD and ten water molecules was studied in the solid state by X-ray crystallography and in solution by NMR spectroscopy. X-ray crystallographic studies of the complex were performed both at room and cryogenic temperatures. The long aliphatic chain of ADM-10 adopts a single conformation at low temperature in contrast to what is observed at room temperature, where two side chain conformations are seen. Both NMR and X-ray crystallography studies indicate that the adamantane moiety of ADM-10 is buried in the betaCD cavity. Chemical shifts in NMR experiments can be explained on the basis of the crystal structure of the complex.