Host-Guest Complexes of Flavanone and 4'-Chloroflavanone with Naturals and Modified Cyclodextrin: A Calorimetric and Spectroscopy Investigations.

The aim of the research was to investigate and compare the interaction between flavanones (flavanone, 4-chloro-flavanone) with potential anticancer activity and selected cyclodextrins. Measurements were made using calorimetric (ITC, DSC) and spectrophotometric (UV-Vis spectroscopy, FT-IR, 1H NMR) methods. The increase in the solubility in aqueous medium caused by the complexation process was determined by the Higuchi-Connors method. As a result of the study, the stoichiometry and thermodynamics of the complexation reaction were determined. The formation of stable inclusion complexes at a 1:1 M ratio between flavanone and 4-chloroflavanone and the cyclodextrins selected for research was also confirmed.

The Interaction of Heptakis (2,6-di-O-Methyl)-ß-cyclodextrin with Mianserin Hydrochloride and Its Influence on the Drug Toxicity.

One tetracyclic antidepressant, mianserin hydrochloride (MIA), has quite significant side effects on a patients' health. Cyclodextrins, which are most commonly used to reduce the undesirable features of contained drugs within their hydrophobic interior, also have the potential to alter the toxic behavior of the drug. The present paper contains investigations and the characteristics of interaction mechanisms for MIA and the heptakis (2,6-di-O-methyl)-ß-cyclodextrin (DM-ß-CD) system, and evaluated the effects of the complexation on MIA cytotoxicity. In order to assess whether there was an interaction between MIA and DM-ß-CD molecules, isothermal titration calorimetry (ITC) have been chosen. Electrospray ionization mass spectrometry (ESI-MS) helped to establish the complex stoichiometry, and circular dichroism spectroscopy was used to describe the process of complex formation. In order to make a wider interpretative perspective, the molecular docking results have been performed. The viability of Chinese hamster cells were investigated in the presence of DM-ß-CD and its complexes with MIA in order to estimate the cytotoxicity of the drug and the conjugate with the chosen cyclodextrin. The viability of B14 cells treated with MIA+DM-ß-CD is lower (the toxicity is higher) than with MIA alone, and no protective effects have been observed for complexes of MIA with DM-ß-CD in any ratio.

Dendrimeric HIV-peptide delivery nanosystem affects lipid membranes structure.

The aim of this study was to evaluate the nature and mechanisms of interaction between HIV peptide/dendrimer complexes (dendriplex) and artificial lipid membranes, such as large unilayered vesicles (LUV) and lipid monolayers in the air-water interface. Dendriplexes were combined as one of three HIV-derived peptides (Gp160, P24 and Nef) and one of two cationic phosphorus dendrimers (CPD-G3 and CPD-G4). LUVs were formed of 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine (DMPC) or of a mixture of DMPC and dipalmitoyl-phosphatidylglycerol (DPPG). Interactions between dendriplexes and vesicles were characterized by dynamic light scattering (DLS), fluorescence anisotropy, differential scanning calorimetry (DSC) and Langmuir-Blodgett methods. The morphology of formed systems was examined by transmission electron microscopy (TEM). The results suggest that dendriplexes interact with both hydrophobic and hydrophilic regions of lipid bilayers. The interactions between dendriplexes and negatively charged lipids (DMPC-DPPG) were stronger than those between dendriplexes and liposomes composed of zwitterionic lipids (DMPC). The former were primarily of electrostatic nature due to the positive charge of dendriplexes and the negative charge of the membrane, whereas the latter can be attributed to disturbances in the hydrophobic domain of the membrane. Obtained results provide new information about mechanisms of interaction between lipid membranes and nanocomplexes formed with HIV-derived peptides and phosphorus dendrimers. These data could be important for the choosing the appropriate antigen delivery vehicle in the new vaccines against HIV infection.

Studies of the Formation and Stability of Ezetimibe-Cyclodextrin Inclusion Complexes.

In the presented studies, the interactions between ezetimibe (EZE) and selected cyclodextrins were investigated. α-Cyclodextrin (αCD), ß-cyclodextrin (ßCD) and its modified derivatives, hydroxypropyl-ß-cyclodextrin (HPßCD) and sulfobutylether-ß-cyclodextrin (SBEßCD), were selected for the research. Measurements were carried out using calorimetric and spectroscopic methods. Additionally, the Hirshfeld surface and biochemical analysis were achieved. As a result of the study, the inclusion complexes with 1:1 stoichiometry were obtained. The most stable are the complexes of ß-cyclodextrin and its derivatives. The comparison of ßCD with its derivatives shows that the modifications have an affect on the formation of more durable and stable complexes.

Physicochemical and in vitro cytotoxicity studies of inclusion complex between gemcitabine and cucurbit[7]uril host.

Gemcitabine, a cytostatic drug from the pyrimidine antimetabolite group, exhibits limited storage stability and numerous side effects during therapy. One of the strategies to improve the effectiveness of therapy with such drugs is the use of supramolecular nano-containers, including dendrimers and macrocyclic compounds. The ability of gemcitabine to attach a proton in an aqueous environment necessitates the search for a carrier that is well-tolerated by an organism and capable of supramolecular binding of a ligand (drug) in a cationic form. In the current study a promising strategy was tested for using cucurbituril Q7 to bind gemcitabine cations for its efficient intracellular delivery on three selected cancer cell lines (MOLT4, THP-1 and U937). Based on physicochemical studies (equilibrium dialysis, UV and 1H NMR titrations, DOSY 1H NMR measurements, DSC calorimetry) and cytotoxicity tests on cells with a free and blocked hENT1 transporter, the conclusion was drawn about the binding and penetration of the cucurbituril-drug complex into cancer cells.

Poly(propyleneimine) glycodendrimers non-covalently bind ATP in a pH- and salt-dependent manner - model studies for adenosine analogue drug delivery.

Adenosine analogue drugs (such as fludarabine or cladribine) require transporter-mediated uptake into cells and subsequent phosphorylation for anticancer activity. Therefore, application of nanocarrier systems for direct delivery of active triphosphate forms has been proposed. Here, we applied isothermal titration calorimetry and zeta potential titration to determine the stoichiometry and thermodynamic parameters of interactions between 4th generation poly(propyleneimine) dendrimers (unmodified or sugar-modified for increased biocompatibility) and ATP as a model adenosine nucleotide. We showed that glycodendrimers have the ability to efficiently interact with nucleoside triphosphates and to form stable complexes via electrostatic interactions between the ionized phosphate and amino groups on the nucleotide and the dendrimer, respectively. The complexation process is spontaneous, enthalpy-driven and depends on buffer composition (strongest interactions in organic buffer) and pH (more binding sites in acidic pH). These properties allow us to consider maltose-modified dendrimers as especially promising carriers for adenosine analogues.

ß-Cyclodextrins incorporated multi-walled carbon nanotubes modified electrode for the voltammetric determination of the pesticide dichlorophen.

In this work, a glassy carbon electrode modified with ß-cyclodextrins and multi-walled carbon nanotubes (ß-CDs/MWCNTs/GCE) was constructed and applied for the square-wave adsorptive stripping voltammetric (SWAdSV) determination of the pesticide dichlorophen (Dcp). For the first time, this compound was electrochemically investigated. The voltammetric measurements were conducted in phosphate buffer (PBS) at pH 6.5 as a supporting electrolyte, and SWAdSV technique parameters were optimized. A linear calibration curve in the wide concentration range from 5.0 × 10-8molL-1 to 2.9 × 10-6molL-1 was obtained. Excellent analytical performance in terms of limit of detection (LOD) of 1.4 × 10-8molL-1 was achieved. The utility of the proposed method was verified by the quantitative analysis of Dcp in Pilica River water samples with satisfactory results. The characterization of modified electrodes was conducted by means of atomic force microscopy (AFM), electrochemical impedance spectroscopy (EIS), and cyclic voltammetry (CV). Moreover, in this work, the dissociation constants (pKa) of Dcp using potentiometric pH titration were estimated. The stoichiometry of the Dcp-ß-CDs inclusion complex formed in solution was determined by proton nuclear magnetic resonance (1H NMR) spectroscopy, and a binding constant (ß2) was estimated from NMR titration studies.

Quantum chemical study and isothermal titration calorimetry of ß-cyclodextrin complexes with mianserin in aqueous solution.

ß-Cyclodextrin (ß-CD) is studied as a carrier of the drug mianserin (MIA). ß-CD with MIA adducts with 1 : 1 and 2 : 1 stoichiometry are investigated in vacuo and in water using quantum chemical methods: PM6 and B3LYP/6-31G(d,p). An effect of the dispersion correction GD2 and the basis set superposition error on the complexation energies is also evaluated. Additionally, the interaction between MIA hydrochloride and ß-CD in aqueous solution at 298.15 K is examined experimentally by isothermal titration calorimetry. Interaction parameters, such as the binding constant, enthalpy, entropy and Gibbs free energy, are presented. Analysis of the obtained data led to the following conclusions: the interaction of MIA with ß-CD is rather strong; there is no significant energetic difference between the 1 : 1 complexes of ß-CD with S-MIA and R-MIA enantiomers; the 2 : 1 (ß-CD : MIA) adduct is energetically more favorable than 1 : 1; the complex formation of MIA + ß-CD is enthalpy and entropy driven.

Spectroscopic and calorimetric studies on the interaction between PAMAM G4-OH and 5-fluorouracil in aqueous solutions.

The results of spectroscopic measurements (an increase in solubility, equilibrium dialysis, 1H NMR titration) and calorimetric measurements (isothermal titration ITC) indicate spontaneous (ΔG<0) binding of 5-fluorouracil molecules by PAMAM G4-OH dendrimer with terminal hydroxyl groups in an aqueous solution. PAMAM G4-OH dendrimer bonds about n=8±1 molecules of the drug with an equilibrium constant of K=70±10. The process of saturating the dendrimer active sites by the drug molecules is exothermal (ΔH<0) and is accompanied by an advantageous change in entropy (ΔS>0). The parameters of binding 5-fluorouracil by PAMAM G4-OH dendrimer were compared with those of binding this drug by the macromolecules of PAMAM G3-OH and G5-OH.

Study of the interactions of PAMAM G3-NH2 and G3-OH dendrimers with 5-fluorouracil in aqueous solutions.

The results of spectroscopic measurements (increase in solubility, equilibrium dialysis, (1)H NMR titration) and calorimetric measurements (isothermal titration ITC) indicate spontaneous (ΔG<0) bonding of 5-fluorouracil by both cationic PAMAM G3-NH2 dendrimer and hydroxyl PAMAM G3-OH dendrimer in aqueous solutions. PAMAM G3-NH2 dendrimer bonds about n= 25±8 drug molecules. Some of them n1= 5±1 are bonded by terminal amine groups with equilibrium constant K1= 3890±930, while the remaining ones n2= 24 ±3 are bonded by amide groups with equilibrium constant K2= 110±30. Hydroxyl PAMAM G3-OH dendrimer bonds n=6.0±1.6 molecules of 5-fluorouracil through tertiary amine groups with equilibrium constant K= 65±10. The parameters of bonding 5-fluorouracil by PAMAM G3-NH2 and G3-OH dendrimer were compared with those of bonding this drug by the macromolecules of PAMAM of generations G4-NH2, G5-NH2 and G5-OH.

Generation-dependent effect of PAMAM dendrimers on human insulin fibrillation and thermal stability.

We have studied the effect of polyamidoamine (PAMAM) dendrimers of various generations on the thermal stability and fibrillation of human insulin. Thermostability of human insulin used differential scanning calorimetry (DSC), which showed two phase-transitions for insulin at 60 and 82°C. After adding dendrimers at 0.6 µmol/l, the first peaks disappeared and the second peaks were higher. We posited that, in the presence of dendrimers, the dimers in the solution were transformed into hexamers. The effect of dendrimers on insulin fibrillation was monitored by measuring ThT fluorescence, and visualization of insulin fibrils by transmission electron microscopy (TEM) and atomic force microscopy (AFM). The effect of PAMAM dendrimers on insulin fibrillation was strongly dependent on the dendrimers generation and dendrimer:protein ratio.

Cytoprotection of pancreatic ß-cells and hypoglycemic effect of 2-hydroxypropyl-ß-cyclodextrin: sertraline complex in alloxan-induced diabetic rats.

Sertraline, a selective inhibitor of serotonin reuptake, is widely used as antidepressant in diabetic patients for improvement of depression and glycemic control. Sertraline is poorly soluble in water, which limits its oral applicability. In this work we tried to improve the pharmaceutical properties of sertraline by complexation with 2-hydroxypropyl-ß-cyclodextrin (HPßCD) and evaluated the efficacy of the HPßCD:sertraline complex in prevention of alloxan-induced lesions in rats. The solubility of sertraline increased in the presence of HPßCD and the association constant for sertraline and HPßCD was equal to 4000 ± 1000 M(-1). Two-week treatment of diabetic animals with the HPßCD:sertraline complex improved pancreatic islet morphology and ß-cell survival, which, in turn, reduced the severity of diabetes, as evidenced by lowering of blood glucose and glycated hemoglobin contents as well as normalization of serum insulin level and insulin sensitivity (HOMA-IR). The effect of the HPßCD:sertraline complex was strongly expressed in comparison with the antidiabetic effect of both the monopreparations, HPßCD and sertraline. It is suggested that the cyclodextrin derivative increased the pharmacological effect of sertraline, probably due to enhanced drug bioavailability.

Spectroscopic and calorimetric studies of formation of the supramolecular complexes of PAMAM G5-NH2 and G5-OH dendrimers with 5-fluorouracil in aqueous solution.

The results of spectroscopic measurements (increase in solubility, equilibrium dialysis, (1)H NMR titration) and calorimetric measurements (isothermal titration ITC) indicate exothermic (ΔH<0) and spontaneous (ΔG < 0) combination of an antitumor drug, 5-fluorouracil, by both cationic PAMAM G5-NH2 dendrimer and its hydroxyl analog PAMAM G5-OH in aqueous solutions at room temperature. PAMAM G5-NH2 dendrimer combines about 70 molecules of the drug with equilibrium constant K ≅ 300, which is accompanied by an increase in the system order (ΔS < 0). Hydroxyl dendrimer, PAMAM G5-OH, combines about 14 molecules of 5-fluorouracil with equilibrium constant K ≅ 100. This process is accompanied by an increase in the system disorder (ΔS > 0).

Effect of inclusion of hydroxycinnamic and chlorogenic acids from green coffee bean in ß-cyclodextrin on their interactions with whey, egg white and soy protein isolates.

The aim of the study was to characterise the interactions of hydroxycinnamic and chlorogenic acids (CHAs) from green coffee, with isolates of proteins from egg white (EWP), whey (WPC) and soy (SPI), depending on pH and temperature. The binding degree was determined by liquid chromatography coupled to a diode array detector and an ultrahigh resolution hybrid quadruple-time-of-flight mass spectrometer with ESI source (LC-QTOF-MS/MS). As a result of binding, the concentration of CHAs in proteins ranged from 9.44-12.2, 11.8-13.1 and 12.1-14.4g/100g for SPI, WPC and EWP, respectively. Thermodynamic parameters of protein-ligand interactions were determined by isothermal titration calorimetry (ITC) and energetics of interactions at the atomic level by molecular modelling. The amount of CHAs released during proteolytic digestion was in the range 0.33-2.67g/100g. Inclusion of CHAs with ß-cyclodextrin strongly limited these interactions to a level of 0.03-0.06g/100g.

Biodegradation and utilization of 4-n-nonylphenol by Aspergillus versicolor as a sole carbon and energy source.

4-n-Nonylphenol (4-n-NP) is an environmental pollutant with endocrine-disrupting activities that is formed during the degradation of nonylphenol polyethoxylates, which are widely used as surfactants. Utilization of 4-n-NP by the filamentous fungus Aspergillus versicolor as the sole carbon and energy source was investigated. By means of gas chromatography-mass spectrometry, we showed that in the absence of any carbon source other than 4-n-NP in the medium, A. versicolor completely removed the xenobiotic (100 mg L(-1)) after 3 d of cultivation. Moreover, mass spectrometric analysis of intracellular extracts led to the identification of eight intermediates. The mineralization of the xenobiotic in cultures supplemented with 4-n-NP [ring-(14)C(U)] as a growth substrate was also assessed. After 3 d of incubation, approximately 50% of the initially applied radioactivity was recovered in the form of (14)CO2, proving that this xenobiotic was completely metabolized and utilized by A. versicolor as a carbon source. Based on microscopic analysis, A. versicolor is capable of germinating spores under such conditions. To confirm these observations, a microcalorimetric method was used. The results show that even the highest amount of 4-n-NP initiates heat production in the fungal samples, proving that metabolic processes were affected by the use of 4-n-NP as an energetic substrate.

Formation of complexes between PAMAM-NH2 G4 dendrimer and L-α-tryptophan and L-α-tyrosine in water.

Interactions between electromagnetic radiation and the side substituents of aromatic amino acids are widely used in the biochemical studies on proteins and their interactions with ligand molecules. That is why the aim of our study was to characterize the formation of complexes between PAMAM-NH2 G4 dendrimer and L-α-tryptophan and L-α-tyrosine in water. The number of L-α-tryptophan and L-α-tyrosine molecules attached to the macromolecule of PAMAM-NH2 G4 dendrimer and the formation constants of the supramolecular complexes formed have been determined. The macromolecule of PAMAM-NH2 G4 can reversibly attach about 25 L-α-tryptophan molecules with equilibrium constant K equal to 130±30 and 24±6 L-α-tyrosine molecules. This characterization was deduced on the basis of the solubility measurements of the amino acids in aqueous dendrimer solutions, the (1)H NMR and 2D-NOESY measurements of the dendrimer solutions with the amino acids, the equilibrium dialysis and the circular dichroism measurements of the dendrimer aqueous solutions with L-α-tryptophan. Our date confirmed the interactions of L-α-tryptophan and L-α-tyrosine with the dendrimer in aqueous solution and indicated a reversible character of the formed complexes.

Thermochemical and spectroscopic studies on the supramolecular complex of PAMAM-NH2 G4 dendrimer and 5-fluorouracil in aqueous solution.

The equilibrium of the formation of polyamidoamine dendrimer (PAMAM-NH(2) G4) and an oncological drug, 5-fluorouracil (FU) in water at room temperature has been examined. Using calorimetric titration, the number of active sites in the dendrimer combining the drug molecules and the equilibrium constant of the dendrimer-drug complex were estimated. The addition of the drug to the dendrimer active sites is an exothermic process. This process is accompanied by a beneficial change in entropy. The number of drug molecules combined by the polymer was confirmed by means of (1)H NMR spectroscopy. (1)HNMR measurements show that the dendrimer macromolecule binds the drug molecules with superficial protonated or unprotonated amine groups.

The influence of maltose modified poly(propylene imine) dendrimers on hen egg white lysozyme structure and thermal stability.

In this study the influence of dendrimers' surface modification upon the strength of interaction with proteins was examined. Unmodified, cationic poly(propylene imine) dendrimer of the fourth generation (PPI G4), two PPI G4 dendrimers, partially and fully coated with maltose residues, and anionic polyamidoamine dendrimer of the third and a half generation (PAMAM G3.5 dendrimer), were used in the study. Hen egg white lysozyme, which possesses a cationic net charge under physiological conditions, was chosen as a model protein. The influence of dendrimers on the thermal stability of lysozyme was studied using differential scanning calorimetry (DSC) and circular dichroism (CD) methods. Additionally, the effect of dendrimers on the availability of lysozyme tryptophan residues to fluorescence quenchers was examined. It was shown that modification of dendrimer surface with maltose reduced its influence on lysozyme properties. However, even full surface modification, resulting in a neutral surface charge, did not deprive dendrimer of the ability to interact with the protein. It was probably caused by the introduction of a large number of hydroxyl groups from maltose residues on the surface of the dendrimer. In the study a comparable strength of influence exerted on lysozyme by cationic PPI dendrimer and anionic PAMAM G3.5 dendrimer was observed. The possible explanation of this fact is the presence of both positively and negatively charged areas on the surface of lysozyme. Such areas allow dendrimers possessing opposite surface charges to interact with lysozyme.

Interaction of polyamidoamine (PAMAM) succinamic acid dendrimers generation 4 with human serum albumin.

Dendrimers, a relatively new group of highly branched three dimensional polymers, are intensively investigated to use them in biomedical and physicochemical sciences. Their specific architecture gives them the ability to interact with many different types of molecules. In our studies the interaction between PAMAM succinamic acid dendrimers generation 4 (PAMAM-SAH G4) and human serum albumin (HSA) was examined. Experiments showed that a single molecule of a HSA can bind approximately 6 particles of dendrimers. The fluorescence studies demonstrated that dendrimers lead to a decrease in protein fluorescence but changes in fluorescence anisotropy were not observed. Alterations in the spectrum of circular dichroism indicated changes in the secondary protein structure. The results clearly show that this generation of dendrimers possesses a strong ability to interact with human serum albumin.

Interaction between PAMAM-NH2 G4 dendrimer and 5-fluorouracil in aqueous solution.

The formation equilibrium of poly(amidoamine) dendrimer (PAMAM-NH2 G4) complex with an oncologic drug such as 5-fluorouracil (5-FU) in water at room temperature was examined. Using the results of the drug solubility in dendrimer solutions and the method of equilibrium dialysis, the maximal number of drug molecules in the dendrimer-drug complex and its equilibrium constant were evaluated. Solubility results show that PAMAM-NH2 G4 dendrimer can transfer tens 5-fluorouracil molecules in aqueous solution. The number of active sites in a dendrimer macromolecule being capable of combining the drug, determined by the separation method, amounts to n=30 ± 4. The calculated equilibrium constant of the 5-FU-active site bonding is equal to K=(400 ± 120).