Cyclodextrin-grafted polysaccharides as supramolecular carrier systems for naproxen

Dextran, mannan and carboxymethylcellulose, previously activated by periodate oxidation, were grafted with beta-cyclodextrin moieties by reductive alkylation in the presence of sodium borohydride. These polymers were used as supramolecular carriers for naproxen, improving the "in vivo" anti-inflammatory properties of this drug.     

Effect of oppositely charged polymer and dissolution medium on swelling, erosion, and drug release from chitosan matrices

The purpose of this research was to investigate the potential use of anionick-carrageenan and nonionic hydroxypropyl-methylcellulose (HPMC, K4) to improve the matrix integrity of directly compressed chitosan tablets containing naproxen sodium, an anionic drug. The influence of buffer pH and drug:polymer ratio on the water uptake, matrix erosion, and drug release were studied. The rapid release of naproxen sodium was seen from matrices containing 100% chitosan due to loss in the matrix cohesiveness; whereas, it was relatively slow for matrices containing optimum concentration ofk-carrageenan. In-situ interaction between oppositely charged moieties resulted in the formation of polyelectrolyte complexes with stoichiometric charge ratios of unity. Fourier transform in frared (FTIR) spectroscopy and powder x-ray diffraction (PXRD) data confirmed the importance of ionic bonds in polyelectrolyte complexation. The ionic interactions between polymers were absent in matrices containing HPMC and the integrity of tablets was improved owing to the presence of viscous gel barrier. The reasons for retarded release of naproxen sodium from the chitosan matrices at different pH include poor aqueous solubility of drug, the formation of a rate-limiting polymer gel barrier along the periphery of matrices, the interaction of naproxen sodium with protonated amino, groups of chitosan, and the interaction of ionized amino groups of chitosan with ionized sulfate groups ofk-carrageenan. Published: June 15, 2007     

Stability of the dimerization domain effects the cooperative DNA binding of short peptides

The basic region peptide derived from the basic leucine zipper protein GCN4 bound specifically to the native GCN4 binding sequences in a dimeric form when the beta-cyclodextrin/adamantane dimerization domain was introduced at the C-terminus of the GCN4 basic region peptide. We describe here how the structure and stability of the dimerization domain affect the cooperative formation of the peptide dimer-DNA complex. The basic region peptides with five different guest molecules were synthesized, and their equilibrium dissociation constants with a peptide possessing beta-cyclodextrin were determined. These values, ranging from 1.3 to 15 microM, were used to estimate the stability of the complexes between the dimers with various guest/cyclodextrin dimerization domains and GCN4 target sequences. An efficient cooperative formation of the dimer complexes at the GCN4 binding sequence was observed when the adamantyl group was replaced with the norbornyl or noradamantyl group, but not with the cyclohexyl group that formed a beta-cyclodextrin complex with a stability that was 1 order of magnitude lower than that of the adamantyl group. Thus, cooperative formation of the stable dimer-DNA complex appeared to be effected by the stability of the dimerization domain. For the peptides that cooperatively formed dimer-DNA complexes, there was no linear correlation between the stability of the inclusion complex and that of the dimer-DNA complex. With the beta-cyclodextrin/adamantane dimerization domain, the basic region peptide dimer preferred to bind to a palindromic 5'-ATGACGTCAT-3' sequence over the sequence lacking the central G.C base pair and that with an additional G.C base pair in the middle. Changing the adamantyl group into a norbornyl group did not alter the preferential binding of the peptide dimers to the palindromic sequence, but slightly affected the selectivity of the dimer for other nonpalindromic sequences. The helical contents of the peptides in the DNA-bound dimer with the adamantyl group were decreased by reducing the stability of the dimer-DNA complex, which was possibly caused by deformation of the helical structure proximal to the dimerization domain.     

NMR spectroscopic study of cyclodextrin inclusion complexes with A-007 prodrugs

One- and two-dimensional NMR spectroscopy was used to demonstrate the formation of inclusion cyclodextrin complexes with several A-007 prodrugs. These complexes are comprised from the encapsulation of the two phenol moieties of the A-007 prodrugs within the cyclodextrin cavity. Considering the size of the two phenol moieties of the A-007 prodrugs compared to the sizes of alpha-, beta-, and gamma-cyclodextrin cavities, we observed complementary binding of the A-007 prodrug with only beta-cyclodextrin, which was also demonstrated spectroscopically. The beta-cyclodextrin inclusion complexes increased the prodrug solubility and modified the prodrug half-life in water. Therefore, beta-cyclodextrin inclusion complexes can be used as an essential form of A-007 prodrug delivery.     

Structural effect of phenyl ring compared to thiadiazole based adamantyl-sulfonamides on carbonic anhydrase inhibition

We investigated the inhibitory activity of sulfonamides incorporating adamantyl moieties against the physiologically relevant human (h) CA (EC 4.2.1.1) isoforms hCA I, II III (cytosolic), IX and XII (transmembrane, tumor-associated). The presence of a benzenesulfonamide instead of an 1,3,4-thiadiazole-sulfonamide fragment in the molecule of CA inhibitors (CAIs) drastically affects both inhibition efficacy and binding within the enzyme active site, as rationalized by means of X-ray crystallography of the adduct of hCA II with 4-(1-adamantylcarboxamidomethyl)benzenesulfonamide. Comparing the present X-ray structure with that of the corresponding 1,3,4-thiadiazole-sulfonamide compound possessing the 1-adamantylcarboxamide moiety, important differences of binding emerged, which explain the highly different inhibition profile of the two compounds against the investigated CA isoforms, most of which (CA I, II, IX and XII) are important drug targets.     

The reversible binding of anti-human serum albumin to poly beta-cyclodextrin-coated porous silica supports

A supramolecular system involving host-guest interactions between immobilized beta-cyclodextrin (beta-CD) cavities and adamantyl groups was evaluated for the preparation of immunosorbents which can be regenerated after use. First a dextran layer bearing both adamantyl groups and carboxylic functions is immobilized onto beta-CD-modified porous silica particles (400 nm) by formation of inclusion complexes. Then, antibody molecules are grafted to the polymer layer. The stationary phases can be prepared in batch or directly in the column. They are stable in aqueous media and are able to trap specifically the corresponding antigen. In case of alteration of the antibody layer, it is possible to remove it by passing a SDS solution through the column. The feasibility of the procedure was evaluated, using the anti-HSA/HSA system.     

Stable sol-gel microstructured and microfluidic networks for protein patterning

We demonstrate the formation of micropatterned sol-gel structures containing active proteins by patterning with polydimethylsiloxane (PDMS) microchannels. To transport sol solution efficiently into the hydrophobic PDMS microchannels, a hydrophilic-hydrophobic block copolymer was used to impart hydrophilicity to the PDMS microchannels. Poor adhesion of the micropatterned gel structure onto glass slides was improved by treating the glass surface with a polymeric substrate. To minimize cracks in the gel microstructure, hybrid matrices of interpenetrating organic and inorganic networks were prepared containing the reactive organic moieties polyvinylalcohol or polyvinylpyrrolidone. Retention of biochemical activity within the micropatterned gel was demonstrated by performing immunobinding assays with immobilized immunoglobulin G (IgG) antibody. The potential application of microfluidics technology to immobilized-enzyme biocatalysis was demonstrated using PDMS-patterned microchannels filled with trypsin-containing sol-gels. This work provides a foundation for the microfabrication of functional protein chips using sol-gel processes.     

Enantiomeric separation of drugs and herbicides on a beta-cyclodextrin-bonded stationary phase.

A chemically bonded beta-cyclodextrin chiral stationary phase for HPLC was prepared in a "one pot" process by the reaction of a phenylated beta-cyclodextrin with silica gel. Various racemic analytes such as drugs (aminoalcohol adrenergic beta-blockers, benzodiazepine anxiolytics, arylpropionic acid antiinflammatory agents) and herbicides (aryloxypropionic acids and esters) were separated on the prepared material. The column showed good chiral recognition ability for most of the solutes tested when using heptane and either 2-propanol or chloroform as organic mobile phase modifiers.     

Effect of Drying Methods on Swelling,Erosion and Drug Release from Chitosan–Naproxen Sodium Complexes

The purpose of this research was to explore theapplication of ionic interactions between naproxen sodium (NS) and chitosan (CH) in complexes (NSC) prepared by tray drying (TD) and spray drying (SD) methods. Drug–polymer ratio (1:1) in the NSC was optimized on the basis of dialysis studies. The particulate systems of NSC were prepared by tray drying (TD) and spray drying (SD) methods. Release retarding polymers were added to the NSC and to the physical mixtures containing NS–CH and their effects on water uptake, matrix erosion and drug release at different pH were compared. Spray dried complexes (SDC) were spherical, free flowing, light and fine amorphous particles in contrast to the crystalline, hard, tenacious, irregularly shaped, denser tray dried complexes (TDC) with poor flowability. Differential scanning calorimetry (DSC), powder X-ray diffraction (PXRD) and Fourier transform infrared (FTIR) patterns confirm the conversion of crystalline to high energy amorphous phase suitable for ionic interactions in NSC. Presence of release retarding polymers, kappa carrageenan and hydroxypropylmethylcellulose (HPMC) in the NSC compacts retarded the drug release and improved the matrix integrity. Carrageenan matrices exhibited more retardation than HPMC tablets. FTIR patterns, erosion, swelling and drug release from matrices support ionic interactions between NS and CH in NSC. The reasons for retarded drug release from the chitosan matrices at acidic pH include poor solubility of drug at acidic pH, formation of a rate limiting polymer gel barrier along the periphery of matrices and the ionic interactions between oppositely charged moieties.     

Mucoadhesive, thermosensitive, prolonged-release vaginal gel for clotrimazole:beta-cyclodextrin complex

The purpose of this study was to achieve a better therapeutic efficacy and patient compliance in the treatment for vaginitis. Clotrimazole (1%) has been formulated in a vaginal gel using the thermosensitive polymer Pluronic F127 (20%) together with mucoadhesive polymers such as Carbopol 934 and hydroxypropylmethylcellulose (0.2% for both). To increase its aqueous solubility, clotrimazole was incorporated as its inclusion complex with 1:1 molar ratio with beta-cyclodextrin. The inclusion complex was thoroughly characterized using various techniques, including 1H NMR spectroscopy, FT IR spectrophotometry, differential scanning calorimetry, scanning electron microscopy, phase solubility studies, and determination of stability constant (k(1:1)). The gelation temperature and rheological behavior of different formulations at varying temperatures were measured. In vitro release profiles of the gels were determined in pH 5.5 citrate buffer. It was observed that complexation with cyclodextrin slowed down the release of clotrimazole considerably. Carbopol 934, on the other hand, was found to interact with beta-cyclodextrin, inducing precipitation. As far as rheological properties are concerned, thermosensitive in situ gelling was obtained with formulations containing drug:cyclodextrin complex rather than with free drug. Thus, the optimum formulation for a controlled-release thermosensitive and mucoadhesive vaginal gel was determined to be clotrimazole:beta-cyclodextrin 1% with 0.2% hydroxypropylmethylcellulose in Pluronic F127 gel (20%) providing continuous and prolonged release of active material above MIC values.     

Novel anticancer polymeric conjugates of activated nucleoside analogues

Inherent or therapy-induced drug resistance is a major clinical setback in cancer treatment. The extensive usage of cytotoxic nucleobases and nucleoside analogues in chemotherapy also results in the development of specific mechanisms of drug resistance, such as nucleoside transport or activation deficiencies. These drugs are prodrugs; and being converted into the active mono-, di-, and triphosphates inside cancer cells following administration, they affect nucleic acid synthesis, nucleotide metabolism, or sensitivity to apoptosis. Previously, we actively promoted the idea that the nanodelivery of active nucleotide species, e.g., 5'-triphosphates of nucleoside analogues, can enhance drug efficacy and reduce nonspecific toxicity. In this study, we report the development of a novel type of drug nanoformulations, polymeric conjugates of nucleoside analogues, which are capable of the efficient transport and sustained release of phosphorylated drugs. These drug conjugates have been synthesized, starting from cholesterol-modified mucoadhesive polyvinyl alcohol or biodegradable dextrin, by covalent attachment of nucleoside analogues through a tetraphosphate linker. Association of cholesterol moieties in aqueous media resulted in intramolecular polymer folding and the formation of small nanogel particles containing 0.5 mmol/g of a 5'-phosphorylated nucleoside analogue, e.g., 5-fluoro-2'-deoxyuridine (floxuridine, FdU), an active metabolite of anticancer drug 5-fluorouracyl (5-FU). The polymeric conjugates demonstrated rapid enzymatic release of floxuridine 5'-phosphate and much slower drug release under hydrolytic conditions (pH 1.0-7.4). Among the panel of cancer cell lines, all studied polymeric FdU-conjugates demonstrated an up to 50× increased cytotoxicity in human prostate cancer PC-3, breast cancer MCF-7, and MDA-MB-231 cells, and more than 100× higher efficacy against cytarabine-resistant human T-lymphoma (CEM/araC/8) and gemcitabine-resistant follicular lymphoma (RL7/G) cells as compared to free drugs. In the initial in vivo screening, both PC-3 and RL7/G subcutaneous tumor xenograft models showed enhanced sensitivity to sustained drug release from polymeric FdU-conjugate after peritumoral injections and significant tumor growth inhibition. All these data demonstrate a remarkable clinical potential of novel polymeric conjugates of phosphorylated nucleoside analogues, especially as new therapeutic agents against drug-resistant tumors.     

Associating hyaluronan derivatives: a novel horizon in viscosupplementation of osteoarthritic joints

The association of two high-molecular-weight hyaluronan (HA) derivatives, namely a beta-cyclodextrin (HA-beta-CD) and an N-acylurea (EDC-HA), dissolved in aqueous NaCl, was studied. The weight-average of the molecular weights (Mw) of HA-beta-CD and of EDC-HA was 185.3 and 86.8 kDa, respectively. However, the Mw value determined for the equimolar mixture of the two biopolymers equaled 556.0 kDa. Similarly, the radius of gyration (dimension) Rg = 80.6 nm of the above equimolar mixture was significantly greater than the values found for the single macromolecules, i.e., 40.2 nm for HA-beta-CD and 23.8 nm for EDC-HA. These data indicate that the two kinds of substituents, borne by the HA polymeric chains, form host-guest inclusion complexes resulting in polymacromolecular associates/aggregates.     

Synthesis of mono-glucose-branched cyclodextrins with a high inclusion ability for doxorubicin and their efficient glycosylation using Mucor hiemalis endo-beta-N-acetylglucosaminidase

The mono-glucose-branched cyclodextrins having an appropriate spacer between the beta-cyclodextrin and a glucose moiety were synthesized from beta-cyclodextrin and arbutin. They had the significantly high association constants for doxorubicin, the anticancer agent, in the range of 10(5)-10(6)M(-1), and worked as highly reactive glycosyl acceptors for the transglycosylation reaction by endo-beta-N-acetylglucosaminidase of Mucor hiemalis to produce sialo-complex type oligosaccharide-branched cyclodextrins in the high yields of 65-67%.     

The development of Cutina lipogels and gel microemulsion for topical administration of fluconazole

The influence of the vehicle on the release and permeation of fluconazole, a topical antifungal drug dissolved in Jojoba oil was evaluated. Series of Cutina lipogels (Cutina CPA [cetyl palmitate], CBS [mixture of glyceryl stearate, cetearyl alcohol, cetyl palmitate, and cocoglycerides], MD [glyceryl stearate], and GMS [glyceryl monostearate]) in different concentrations as well as gel microemulsion were prepared. In-vitro drug release in Sorensens citrate buffer (pH 5.5) and permeation through the excised skin of hairless mice, using a modified Franz diffusion cell, were performed. The rheological behavior and the apparent viscosity values for different gel bases were measured before and after storage under freezing conditions at −4 °C and were taken as measures for stability of network structure.Candida albicans was used as a model fungus to evaluate the antifungal activity of the best formula achieved. The results of in vitro drug release and its percutaneous absorption showed that the highest values from gel microemulsion were assured. The rheological behavior of the prepared systems showed pseudoplastic (shear-thinning) flow indicating structural breakdown of the existing intermolecular interactions between polymeric chains. Moreover, the stability study revealed no significant difference between viscosity before and after storage for different formulae except for CPA Cutina lipogel (using analysis of variance [ANOVA] test at level of significance .05). The antifungal activity of fluconazole showed the widest zone of inhibition with gel microemulsion. The gel microemulsion is an excellent vehicle for fluconazole topical drug delivery.     

Synthesis of ethylenediamine linked beta-cyclodextrin dimer and its analytical application for tranilast determination by spectrofluorimetry

A synthesis of beta-cyclodextrin (beta-CD) dimer, containing two beta-CD moieties that are linked through their sides by ethylenediamine, was presented. The dimer was characterized by means of IR, (1)H NMR, (13)C NMR, and elemental analysis. The inclusion complexation behavior of beta-cyclodextrin dimer with tranilast was studied in an aqueous KH(2)PO(4)-citric acid buffer solution of pH 2.00 at room temperature by spectrofluorimetry. Based on the significant enhancement of fluorescence intensity of tranilast, a spectrofluorimetric method with high sensitivity and selectivity was developed for the determination of tranilast in bulk aqueous solution in the presence of ethylenediamine beta-CD dimer. The apparent association constant of the complex was 8.39 x 10(3) L mol(-1), and the linear range was 10.8-1.40 x 10(4) ng mL(-1) with the detection limit 3.2 ng mL(-1). There was no interference from the excipients normally used in tablets and serum constituents. The proposed method was successfully applied to the determination of tranilast in serum.     

Danazol-beta-cyclodextrin binary system: a potential application in emergency contraception by the oral route

This study explored the potential of beta-cyclodextrin to improve the aqueous solubility and dissolution of danazol, investigated a simple and less expensive method for preparation of a danazol-beta-cyclodextrin binary system, and explored the potential application of a danazol-beta-cyclodextrin binary system as a single-dose emergency contraceptive. Phase solubility analysis indicated formation of a first-order soluble complex with stability constant 972.03 M(-1), while Job's plot affirmed 1:1 stoichiometry. The hyperchromic shift in the UV-Vis spectrum of danazol in the presence of beta-cyclodextrin indicated solubilization capability of beta-cyclodextrin for danazol. The extrinsic Cotton effect with a negative peak at 280.7 nm confirmed the inclusion of danazol in the asymmetric locus of beta-cyclodextrin. (1)H-nuclear magnetic resonance analysis suggested that the protons of the steroidal skeleton of danazol display favorable interactions with the beta-cyclodextrin cavity. The danazol-beta-cyclodextrin binary system was prepared by kneading, solution, freeze-drying, and milling methods. The extent of the enhancement of dissolution rate was found to be dependent on the preparation method. Dissolution studies showed a similar relative dissolution rate (2.85) of the danazol-beta-cyclodextrin binary system prepared by the freeze-drying and milling (in the presence of 13% moisture) methods. In a mouse model, the danazol-beta-cyclodextrin binary system at 51.2 mg/kg (equivalent to a 400-mg human dose) showed 100% inhibition of implantation when given postcoitally. Moreover, the danazol-beta-cyclodextrin binary system is safe up to 2000 mg/kg in the mouse (15.52 g/70 kg human) as a single oral dose. Thus, the danazol-beta-cyclodextrin binary system could serve as a new therapeutic application: an oral emergency contraceptive at a physiologically acceptable single dose.     

Molecular state of cy tochrome c oxidase on polyacrylamide gel electrophoresis

The molecular state of cytochrome oxidase, complex IV (EC 1.9.3.1), was studied by polyacrylamide gel electrophoresis. Cytochrome oxidase in the presence of non-ionic detergent Emasol 1130 ran as a single band under conditions where there is a small sieving effect as in a 2.5% polyacrylamide gel. This polymeric form is an association of different molecular species that can be dissociated on a preparative electrophoresis system. In such a system, five species of different molecular size with enzymic activity are obtained, though the specific activity is lower in the first running fractions. After treatment in highly dissociating conditions (phenol-acetic acid-water (2:1:1, w/v/v) the polymeric form and the different species present two main fractions on analytical polyacrylamide electrophoresis (7.5% in acrylamide, 35% acetic acid, and 5 m urea). The original form after treatment with 2.5% sodium dodecylsulfate (SDS) also presents two fractions on analytical electrophoresis in the presence of 2.5% sodium dodecylsulfate. These results suggest that the basic subunit structure of the various forms of the cytochrome oxidase consists of two closely related polypeptides. The highest activity was found with the polymeric form of cytochrome oxidase, a fact which may have physiological significance in relation to the natural state of this enzyme in the mitochondrion.     

Synthesis and evaluation of carbaborane derivatives of indomethacin as cyclooxygenase inhibitors

Nonsteroidal anti-inflammatory drugs (NSAIDs) exert their pharmacological activities by inhibiting cyclooxygenase (COX)-1 and COX-2. Previous studies have shown that esters and amides of non-selective inhibitors such as indomethacin are selective against COX-2, which is the therapeutically relevant isoform. Structure-activity analysis indicates that substituted phenyl rings are tolerated as ester components. In the present study, the introduction of inorganic ortho- and meta-carbaborane moieties was explored with the aim to create COX-2 inhibitors and more importantly to investigate the validity of using these boron clusters as drug entities. Interestingly, only the ortho-carbaborane ester was active whereas the meta isomer was not. A similar lack of inhibitory potency was observed when an adamantyl substituent or alkylene spacers at the carbaborane were introduced in the ester functionality.     

Detection of DNA sites damaged by 1-(4-amino-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride (ACNU) using a DNA sequencing procedure

A DNA sequencing technique was applied to the highly reiterated DNA from HeLa S3 cells in order to detect DNA damage induced by the antitumor drug, 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride (ACNU). A DNA reiterated fragment of 92 base pairs (bp) was isolated by gel electrophoresis after EcoRI and EcoRI restriction endonuclease digestion. In the defined sequence of the 92 bp fragment, ACNU caused damage and modifications primarily at guanine moieties, leading to alkali-labile sites as determined by subsequent piperidine reaction on an extended Maxam-Gilbert sequencing gel. These results indicate that guanine moieties in double-stranded DNA are preferentially vulnerable to ACNU over other base moieties.     

Interaction of beta-cyclodextrin with the granular starch binding domain of glucoamylase

The granular starch binding domain of glucoamylase 1 (EC 3.2.1.3 1,4-alpha-D-glucan glucohydrolase) binds two molecules of beta-cyclodextrin, with a dissociation constant (Kd) for the second ligand of 1.68 microM. The catalytic domain showed no interaction with beta-cyclodextrin. Beta-cyclodextrin competitively inhibited the adsorption of the binding domain onto granular starch with an inhibition constant (Ki) of 11.0 +/- 1.9 microM. The results show that beta-cyclodextrin binds to the binding domain of glucoamylase at the same site(s) as granular starch.