Biomedical subjects

K Uekama

Publications and source records attributed to K Uekama.

At least 55 records · Page 3Linked to original sources

PubMed1997

Application of a beta-cyclodextrin sulfate-immobilized precolumn to selective on-line enrichment and separation of heparin-binding proteins by column-switching high-performance liquid chromatography.

A column-switching high-performance liquid chromatography (HPLC) system which consisted of a beta-cyclodextrin (beta-CD) sulfate-immobilized hydrophilic vinyl-polymer gel precolumn and a reversed-phase analytical column was developed for the selective on-line enrichment and separation of heparin-binding proteins. Of 15 proteins investigated, 10 proteins having heparin-binding activity were retained on the beta-CD sulfate precolumn almost quantitatively, in contrast 5 proteins having no heparin-binding activity were not retained. Calibration graphs for basic fibroblast growth factor constructed at various sample volumes were nearly identical, indicating that the protein could be enriched by this system. The system was successfully used for the selective separation of lysozyme in egg white. The beta-CD sulfate-immobilized precolumn showed no loss of analytical performance over 2 years during which about 400 samples were analysed.

Chromatography, High Pressure Liquid↗

PubMed1997

Pharmaceutical applications of cyclodextrins. III. Toxicological issues and safety evaluation.

The objective of this review is to summarize recent findings on the safety profiles of three natural cyclodextrins (alpha-, beta- and gamma-CDs) and several chemically modified CDs. To demonstrate the potential of CDs in pharmaceutical formulations, their stability against non-enzymatic and enzymatic degradations in various body fluids and tissue homogenates and their pharmacokinetics via parenteral, oral, transmucosal, and dermal routes of administration are outlined. Furthermore, the bioadaptabilities of CDs, including in vitro cellular interactions and in vivo safety profiles, via a variety of administration routes are addressed. Finally, the therapeutic potentials of CDs are discussed on the basis of their ability to interact with various endogenous and exogenous lipophiles or, especially for sulfated CDs, their effects on cellular processes mediated by heparin binding growth factors.

Animals↗

PubMed1997

Protection afforded by maltosyl-beta-cyclodextrin against alpha-chymotrypsin-catalyzed hydrolysis of a luteinizing hormone-releasing hormone agonist, buserelin acetate.

PURPOSE: The present study addresses how maltosyl-beta-cyclodextrin (G2-beta-CyD) impacts upon the alpha-chymotrypsin-catalyzed hydrolysis of buserelin acetate, an agonist of luteinizing hormone-releasing hormone with emphasis upon the direct effect of G2-beta-CyD on the activity of the protease. METHODS: Kinetic and solubility studies were performed in isotonic phosphate buffer (pH 7.4) at 25 degrees C and 37 degrees C. The interaction of alpha-chymotrypsin with G2-beta-CyD in the buffer solution was examined by differential scanning calorimetry. RESULTS: G2-beta-CyD decelerated the alpha-chymotrypsin-catalyzed hydrolysis of buserelin acetate to give the 1-3 tripeptide and the 4-9 hexapeptide fragments. This deceleration can be explained solely by a non-productive encounter between a complex of the substrate with G2-beta-CyD and the protease at relatively low CyD concentrations, while the direct inhibitory effect of G2-beta-CyD on the proteolytic activity made a considerable contribution to the overall deceleration of the hydrolysis at higher CyD concentrations. Calorimetric studies indicate the presence of intermediate states in the thermal unfolding of alpha-chymotrypsin, simultaneously accompanied by the autolysis. By contrast, a two-state thermal unfolding of alpha-chymotrypsin was observed in the presence of G2-beta-CyD, suggesting reduced proteolytic activity upon binding to G2-beta-CyD. CONCLUSIONS: These results suggest that G2-beta-CyD at higher concentrations inhibits the proteolytic action of alpha-chymotrypsin through direct interaction with the protease, as well as through the formation of a non-productive complex with the substrate.

Buserelin↗

PubMed1997

Spectroscopic characterization of the inclusion complex of a luteinizing hormone-releasing hormone agonist, buserelin acetate, with dimethyl-beta-cyclodextrin.

Inclusion complexation of buserelin acetate, and agonist of luteinizing hormone-releasing hormone, with dimethyl-beta-cyclodextrin (DM-beta-CyD) in aqueous solution was studied spectroscopically and its mode of interaction was assessed. Ultraviolet absorption and circular dichroism (CD) spectroscopies indicate that the aromatic side chains of buserelin acetate, L-tryptophan and L-tyrosine residues, are incorporated into the hydrophobic environment of the DM-beta-CyD activity. Furthermore, proton and carbon-13 nuclear magnetic resonance spectroscopies suggest that in addition to the two aromatic side chains, a tertiary butyl D-serine residue is inserted into the DM-beta-CyD cavity from the secondary hydroxyl side. On the other hand, the continuous variation plots for the buserelin acetate: DM-beta-CyD system showed a 1:1 stoichiometry of the complex. Therefore, the complexation should be initiated by the inclusion of one of the three binding sites on the buserelin molecule into DM-beta-CyD, which may in turn prevent the further access of the second cyclodextrin to the other binding sites, probably due to steric hindrance and/or conformational changes of the peptide. These structural features of the complex would account for the stabilizing effect of DM-beta-CyD on the enzymatic degradation of buserelin acetate.

Buserelin↗

PubMed1997

Varying effects of cyclodextrin derivatives on aggregation and thermal behavior of insulin in aqueous solution.

Maltosyl-beta-cyclodextrin (G2-beta-CyD) suppressed the aggregation of insulin in neutral solution, while the sulfate of beta-CyD (S-beta-CyD) accelerated the aggregation. On the other hand, the sulfobutyl ether of beta-CyD (SBE-beta-CyD) showed varying effects on insulin aggregation, depending on the degree of substitution of the sulfobutyl group: i.e., the inhibition at relatively low substitution and acceleration at higher substitution. Differential scanning calorimetric studies indicate that the self-association of insulin stabilized the native conformation of the peptide, as indicated by an increase in the mean unfolding temperature (Tm). G2-beta-CyD and SBE-beta-CyD decreased the Tm value of insulin oligomers, while S-beta-CyD increased the Tm value. 1H-Nuclear magnetic resonance spectroscopic studies suggest that G2-beta-CyD includes accessible hydrophobic side chains of insulin within the CyD cavity, and hence perturbs the intermolecular hydrophobic contacts between aromatic side chains across the monomer-monomer interfaces. By contrast, the electrostatic interaction between the positive charges of insulin and the concentrated negative charges of the sulfate and sulfonate groups of the anionic beta-CyDs seems to be more of a factor than the inclusion effects. These results suggest proper use of the CyD derivatives could be effective in designing rapid or long-acting insulin preparations.

Animals↗

PubMed1996

The interaction of charged and uncharged drugs with neutral (HP-beta-CD) and anionically charged (SBE7-beta-CD) beta-cyclodextrins.

PURPOSE: The objective of this work was to determine the role that charge might play in the interaction of charged and uncharged drugs with neutral (2-hydroxypropyl-beta-cyclodextrin, HP-beta-CD) and anionically charged (SBE7-beta-CD) modified beta-cyclodextrins. SBE7-beta-CD is a sulfobutyl ether, sodium salt, derivative variably substituted on the 2-, 3- and the 6-positions of beta-cyclodextrin. The number seven refers to the average degree of substitution. METHODS: The binding of the acidic drugs, indomethacin, naproxen and warfarin and the basic drugs, papaverine, thiabendazole, miconazole and cinnarizine with the two cyclodextrins was determined at 25 degrees C as a function of pH and cyclodextrin concentration by the phase-solubility method. RESULTS: Except for miconazole and cinnarizine (Ap-type diagrams), all other materials studied displayed AL-type diagrams. By comparing the binding constants of both the charged and uncharged forms of the same drugs to both HP-beta-CD and SBE7-beta-CD, the following conclusions could be drawn. The binding constants for the neutral forms of the drugs were always greater with SBE7-beta-CD than with HP-beta-CD. For the anionic agents, the binding constants between SBE7-beta-CD and HP-beta-CD were similar while the binding constants for the cationic agents with SBE7-beta-CD were superior to those of HP-beta-CD, especially when compared with the neutral form of the same drug. CONCLUSIONS: A clear charge effect on complexation, attraction in the case of cationic drugs and perhaps inhibition in the case of anionic drugs, was seen with the SBE7-beta-CD.

2-Hydroxypropyl-beta-cyclodextrin↗

PubMed1996

In-vitro evaluation of biphenylyl acetic acid-beta-cyclodextrin conjugates as colon-targeting prodrugs: drug release behaviour in rat biological media.

Biphenylyl acetic acid was selectively conjugated to one of the primary hydroxyl groups of beta-cyclodextrin through an ester- or amide-linkage, and the physicochemical properties (aqueous solubility and hydrolysis) were investigated. Aqueous solubility of the conjugates was lower than those of either drug or parent beta-cyclodextrin. The amide conjugate was stable in aqueous solution and in rat biological fluids and gastrointestinal contents. The ester conjugate was hydrolysed to beta-cyclodextrin and biphenylyl acetic acid at moderate rates resulting in a V-shaped rate-pH profile in aqueous solution. The ester conjugate released the drug preferentially when incubated with the contents of caecum or colon, whereas no appreciable drug release was observed on incubation with contents of stomach or intestine, nor on incubation with intestinal or liver homogenates, nor on incubation with rat blood. The present results suggest that the ester-type drug conjugate of beta-cyclodextrin may serve as a colon-targeting prodrug.

Animals↗

PubMed1996

Combination effects of alpha-cyclodextrin and xanthan gum on rectal absorption and metabolism of morphine from hollow-type suppositories in rabbits.

Pharmacokinetics of morphine and its glucuronides in plasma were studied after rectal administration of hollow-type oleaginous suppositories containing kneading mixtures of morphine hydrochloride, alpha-cyclodextrin, and/or xanthan gum in rabbits. In combination with xanthan gum, alpha-cyclodextrin reduced the first-pass metabolism of morphine in the rectal mucosa and by the liver and improved the apparent rectal bioavailability of the opioid about 4 fold. In vitro permeation studies using an isolated rectal mucosal preparation of rabbits revealed that alpha-cyclodextrin increased the transepithelial conductance and facilitated the transport of morphine through the rectal mucosa. Furthermore, alpha-cyclodextrin facilitated its own mucosal permeation and reduced the glucuronidation of morphine during the passage through the rectal mucosa, probably through restricting the formation of a catalytic complex of morphine with glucuronyltransferases, rather than because of the enzyme saturation. The present data suggest that alpha-cyclodextrin in combination with xanthan gum is particularly effective in improving the rectal bioavailability of morphine from hollow-type suppositories.

Analgesics, Opioid↗

PubMed1996

Possible enhancing mechanism of the cutaneous permeation of 4-biphenylylacetic acid by beta-cyclodextrin derivatives in hydrophilic ointment.

The enhancing effects of heptakis(2,6-di-O-methyl)-beta-cyclodextrin (DM-beta-CyD) and 2-hydroxypropyl-beta-cyclodextrin (HP-beta-CyD) on the percutaneous absorption of 4-biphenylylacetic acid (BPAA), a nonsteroidal anti-inflammatory drug, in hydrophilic ointment were studied and compared with the parent beta-cyclodextrin (beta-CyD). 13C-NMR measurements suggested that the biphenyl group of BPAA is preferably included within the cavity of three beta-CyDs. The three beta-CyDs remarkably enhanced the release of BPAA from the hydrophilic ointment base and the in vitro cutaneous permeation, depending on the increase in solubility of BPAA in the ointment base. Pretreatment of the ointment containing DM-beta-CyD or HP-beta-CyD onto the isolated skin of hairless mice, however, provided no effects on the skin permeation of BPAA. When propylene glycol was used as a vehicle, both the release rate and cutaneous permeation parameters showed no appreciable difference between BPAA alone and its HP-beta-CyD complex, because the solubilities of BPAA and its HP-beta-CyD complex were almost comparable in the vehicle. The present results suggested that the enhancing effect of beta-CyDs on the percutaneous absorption of BPAA can be mainly ascribed to an increase in the solubility of BPAA in the hydrophilic ointment.

Animals↗

PubMed1995

Modification of rectal absorption of morphine from hollow-type suppositories with a combination of alpha-cyclodextrin and viscosity-enhancing polysaccharide.

An attempt was made to optimize the rectal delivery of morphine, using cyclodextrins as an absorption enhancer and polysaccharides as a swelling hydrogel in Witepsol H-15 hollow-type suppositories, and this was tested in rabbits. alpha- and beta-cyclodextrins enhanced the rate and extent of bioavailability, the former being more effective; gamma-cyclodextrin decreased the absorption of morphine. The in-vitro membrane permeation studies using excised rectal sacs revealed that alpha-cyclodextrin enhanced the permeation of morphine through the rectal membranes. In contrast, viscous polysaccharides such as xanthan gum retarded the plasma morphine levels after the rectal administration, reflecting in-vitro slow release characteristics. A combination of alpha-cyclodextrin and xanthan gum produced sustained plasma profiles of morphine along with an increased rectal bioavailability (more than 4 times). From the observation of the distribution behavior of suppositories in rabbit rectum and colon after the rectal administration, xanthan gum was found to prevent the upward spread of the drug. Gross and microscopic observations suggested that this preparation was less irritating to the rectal mucosa.

Animals↗

PubMed1995

Improvement of nasal bioavailability of luteinizing hormone-releasing hormone agonist, buserelin, by cyclodextrin derivatives in rats.

The effects of chemically modified cyclodextrins on the nasal absorption of buserelin, an agonist of luteinizing hormone-releasing hormone, were investigated in anesthetized rats. Of the cyclodextrins tested, dimethyl-beta-cyclodextrin (DM-beta-CyD) was the most effective in improving the rate and extent of the nasal bioavailability of buserelin. Fluorescence spectroscopic studies indicated that the cyclodextrins formed inclusion complexes with buserelin, which may reduce the diffusibility of buserelin across the nasal epithelium and may participate in the protection of the peptide against enzymatic degradation in the nasal mucosa. Additionally, the cyclodextrins increased the permeability of the nasal mucosa, which was the primary determinant based on the multiple regression analysis of the nasal absorption enhancement of buserelin. Scanning electron microscopic observations revealed that DM-beta-CyD induced no remarkable changes in the surface morphology of the nasal mucosa at a minimal concentration necessary to achieve substantial absorption enhancement. The present results suggest that DM-beta-CyD could improve the nasal bioavailability of buserelin and is well-tolerated by the nasal mucosa of the rat.

Absorption↗

PubMed1995

Differential effects of sulfate and sulfobutyl ether of beta-cyclodextrin on erythrocyte membranes in vitro.

The hemolytic activity of beta-cyclodextrin (beta-CyD) on rabbit erythrocytes was reduced by the introduction of negatively-charged groups onto the hydroxyls of beta-CyD; the membrane disrupting abilities decreased in the order of beta-CyD > 2-hydroxypropyl-beta-CyD (HP-beta-CyD) > sulfobutyl-beta-CyD (SB-beta-CyD) >> beta-CyD sulfate (S-beta-CyD). Under pre-hemolytic concentrations, both beta-CyD and SB-beta-CyD induced shape changes of membrane invagination on the erythrocytes. In sharp contrast, S-beta-CyD showed biphasic effect on the shape of the erythrocytes; i.e. the crenation at relatively low concentrations and the invagination at higher concentrations. The S-beta-CyD-induced membrane crenation arose from a direct action on the membranes rather than cell metabolism-mediated effects. Unlike beta-CyD, S-beta-CyD was found to bind to the erythrocytes and may be confined to the outer surface of the membrane bilayer, which may expand the exterior layer relative to the cytoplasmic half, thereby inducing the cells to crenate. On the other hand, the membrane invagination mediated by the three beta-CyDs was initiated by extracting specific membrane lipids from the cells, depending upon their inclusion abilities, subsequently leading to the lysis of the cells. These results indicate that SB-beta-CyD and S-beta-CyD interact with the erythrocyte membranes in a differential manner and possess lower membrane disrupting abilities than the parent beta-CyD and HP-beta-CyD.

2-Hydroxypropyl-beta-cyclodextrin↗

PubMed1995

In-vivo and in-vitro correlation for delayed-release behaviour of a molsidomine/O-carboxymethyl-O-ethyl-beta-cyclodextrin complex in gastric acidity-controlled dogs.

The in-vivo absorption behaviour of molsidomine from the delayed-release tablets of an O-carboxymethyl-O-ethyl-beta-cyclodextrin complex was investigated using gastric acidity-controlled dogs under fasted and non-fasted conditions. The in-vitro release profiles were generated by changing the pH of the dissolution medium at different rotation paddle speeds. The absorptivity of molsidomine in the high acidity dog was correlated with the pH-changed release profile (pH 1.2 to 7.0 after 2 h), whereas that in the low acidity dog was correlated with the release profile at a constant pH of 7.0. The absorption in fasted dogs was well correlated with the in-vitro release at the low-rotation paddle speed (< 5 rev min-1), whereas that in the non-fasted dogs was correlated with that of high rotation (100 rev min-1). The present results suggested that the in-vivo delayed-release behaviour of the complex is predictable from the in-vitro release profiles generated using pH-variable dissolution testing apparatus at different rotation speeds of the paddle.

Animals↗

PubMed1995

Enhanced nasal delivery of luteinizing hormone releasing hormone agonist buserelin by oleic acid solubilized and stabilized in hydroxypropyl-beta-cyclodextrin.

The potential use of three 2-hydroxypropyl ether derivatives of cyclodextrins (HP-alpha-, HP-beta- and HP-gamma-CyDs) as biocompatible solubilizers and stabilizers for oleic acid, a lipophilic absorption enhancer, was assessed in the nasal absorption of buserelin, an agonist of luteinizing hormone-releasing hormone, in rats. HP-CyDs increased the aqueous solubility of oleic acid and protected it against oxidation through the formation of inclusion complexes with the efficacy increasing in the order: HP-gamma-CyD << HP-alpha-CyD < HP-beta- CyD. The bend structure due to a cis-double bond halfway along the acyl chain of oleic acid provided a better fit into the cavity of HP-beta-CyD, in which the double bond appears to be buried, and hence becomes less susceptible to oxidation. The rate and extent of nasal bioavailability of buserelin were remarkably increased by coadministration of oleic acid and HP-beta-CyD, compared with the sole use of the enhancer. This enhancement was ascribable to the lowering of both the enzymatic and physical barriers of the nasal epithelium to the peptide, probably through the facilitated transmucosal penetration of oleic acid solubilized in HP-beta-CyD.

2-Hydroxypropyl-beta-cyclodextrin↗

PubMed1995

Direct measurement of high-density lipoprotein cholesterol in serum with polyethylene glycol-modified enzymes and sulfated alpha-cyclodextrin.

We have developed an automated method for measuring high-density lipoprotein (HDL)-cholesterol in serum without prior separation, using polyethylene glycol (PEG)-modified enzymes and sulfated alpha-cyclodextrin. When cholesterol esterase and cholesterol oxidase enzymes were modified with PEG, they showed selective catalytic activities towards lipoprotein fractions, with the reactivity increasing in the order: low-density lipoprotein < very-low-density lipoprotein approximately chylomicron < HDL. In the presence of magnesium ions, alpha-cyclodextrin sulfate reduced the reactivity of cholesterol, especially in chylomicrons and very-low-density lipoprotein, without the need for precipitation of those lipoprotein fractions. The combination of PEG-modified enzymes with alpha-cyclodextrin sulfate provided selectivity for the determination of HDL-cholesterol in serum in the presence of a small amount of dextran sulfate without the need for precipitation of lipoprotein aggregates. The results of the HDL-cholesterol assayed in serum by this direct method correlated well with those obtained by precipitation-based methods and also that by an ultracentrifugation method.

Adult↗

PubMed1994

Effect of 2-hydroxypropyl-beta-cyclodextrin on crystallization and polymorphic transition of nifedipine in solid state.

The glassy state of nifedipine (NP) was prepared in the absence and presence of 2-hydroxypropyl-beta-cyclodextrin (HP-beta-CyD), and its crystallization and polymorphic transition behavior was investigated by differential scanning calorimetry (DSC) and powder X-ray diffractometry. In DSC thermograms, the glassy NP exhibited an endothermic peak at 48 degrees C representing the glass transition of NP, an exothermic peak at 105 degrees C for the crystallization to a metastable form of NP (Form B), an exothermic peak at 125 degrees C for the polymorphic transition of Form B to a stable form of NP (Form A), and an endothermic peak at 171 degrees C for the melting of Form A. The powder X-ray diffractogram of Form B was apparently different from that of Form A. In the presence of HP-beta-CyD, the exothermic peak at 125 degrees C for the Form B to A transition disappeared and a new endothermic peak appeared at 163 degrees C. This new peak was ascribed to the melting of Form B, and the conversion of Form B to Form A was significantly suppressed in HP-beta-CyD matrix. Upon storage at 60 degrees C, the glassy NP was converted to Form A with an activation energy of 18 kcal/mol. The apparent dissolution rate of the NP/HP-beta-CyD (molar ratio 1:1) increased in the order of glassy NP < Form A < Form B, because the glassy NP was readily converted to Form A upon contact with water, resulting in a lower dissolution rate.(ABSTRACT TRUNCATED AT 250 WORDS)

2-Hydroxypropyl-beta-cyclodextrin↗

PubMed1994

Aluminium beta-cyclodextrin sulphate as a stabilizer and sustained-release carrier for basic fibroblast growth factor.

The water-insoluble aluminium salt of beta-cyclodextrin sulphate (Al.beta-CyD-Sul) was used as a stabilizer and sustained-release carrier for recombinant human basic fibroblast growth factor (bFGF). An adsorbate of bFGF with Al.beta-CyD-Sul was prepared by incubating the protein with a suspension of Al.beta-CyD-Sul in water. The mitogenic activity of bFGF released from the adsorbate, as indicated by the proliferation of kidney cells of baby hamster (BHK-21), was almost comparable with that of the intact bFGF. Al.beta-CyD-Sul significantly protected bFGF from proteolytic degradation by pepsin and alpha-chymotrypsin, compared with the water-soluble sodium salt. The in-vitro release of bFGF from the adsorbate was sustained in proportion to a rise in the ratio of Al.beta-CyD-Sul to the protein in the adsorbate. Of the bFGF preparations evaluated, the adsorbate of bFGF with Al.beta-CyD-Sul, when given subcutaneously to the rat, showed the most prominent increase in the formation of granulation tissues, due to the stabilization and slow-release of the mitogen. The limited data presented here suggest that the adsorbate of bFGF with Al.beta-CyD-Sul has a potent therapeutic efficacy for wound healing, and may be applicable to oral protein formulations for the treatment of intestinal mucosal erosions.

Animals↗

PubMed1994

In-vivo and in-vitro evaluations of a modified-release oral dosage form of nifedipine by hybridization of hydroxypropyl-beta-cyclodextrin and hydroxypropylcelluloses in dogs.

To maintain a suitable blood level of nifedipine for a long period of time, double-layer tablets consisting of 2-hydroxypropyl-beta-cyclodextrin (HP-beta-CyD) and 3% nonionic surfactant (HCO-60) as a fast-release portion and hydroxypropylcelluloses (HPCs) with different viscosity grades (low, medium and high) as a slow-release portion were prepared, and their in-vitro and in-vivo release behaviours were investigated. Among the seven formulations, the tablet having the mean dissolution time of 0.8-1.3 h gave prolonged plasma nifedipine levels without decrease of AUC after oral administration to dogs. Consequently, the double-layer tablet consisting of HP-beta-CyD with 3% HCO-60/(HPC-low:HPC-medium) in a weight ratio 1/(1.5:1.5) was selected as an appropriate modified-release formulation because it elicited almost comparable retarding effects with superior oral bioavailability compared with those of a commercially available slow-release nifedipine product.

2-Hydroxypropyl-beta-cyclodextrin↗