Complexation of acridine orange by cucurbit[7]uril and beta-cyclodextrin: photophysical effects and pKa shifts.

The host-guest interactions of the neutral (AO) and cationic (AOH+) forms of the dye acridine orange with the macrocyclic hosts cucurbit[7]uril (CB7) and beta-cyclodextrin (beta-CD) were investigated by using ground-state absorption and steady-state as well as time-resolved fluorescence measurements. The cationic form undergoes no significant complexation with beta-CD, but binds strongly with CB7 (Keq = 2.0 x 10(5) M(-1)), causing a large enhancement in fluorescence intensity and lifetime of the dye in the latter host. The strong and selective binding of AOH+ with CB7 is attributed to ion-dipole interactions involving the ureido carbonyl rims of CB7 and the charged AOH+. In contrast, the neutral AO form of the dye shows quite similar binding with both CB7 and beta-CD, but the binding constants are lower by more than two orders of magnitude compared to that of the AOH+-CB7 system. CB7 and beta-CD show a contrasting behavior in modifying the acid-base character of the dye, shifting its pKa by about 2.6 units upward and about 0.7 units downward, in the two respective cases. These divergent pKa shifts of the dye arise from the differential affinity of the two host molecules to encapsulate the protonated and neutral form of the dye.     

Assembly, intracellular processing, and expression at the cell surface of the human alpha beta T cell receptor/CD3 complex. Function of the CD3-zeta chain

The TCR/CD3 complex is a multimeric protein complex composed of a minimum of seven transmembrane chains (TCR alpha beta-CD3 gamma delta epsilon zeta 2). Whereas earlier studies have demonstrated that both the TCR-alpha and -beta chains are required for the cell surface expression of the TCR/CD3 complex, the role of the CD3 chains for the TCR/CD3 expression have not been experimentally addressed in human T cells. In this study the function of the CD3-zeta chain for the assembly, intracellular processing, and expression of the TCR/CD3 complex in the human leukemic T cell line Jurkat was investigated. The results indicate that: 1) CD3-zeta is required for the cell surface expression of the TCR/CD3 complex; 2) the pentameric form (TCR alpha beta-CD3 gamma delta epsilon) of the TCR/CD3 complex and single TCR chains associated with CD3 (TCR alpha-CD3 gamma delta epsilon and TCR beta-CD3 gamma delta epsilon) are produced in the endoplasmic reticulum in the absence of CD3-zeta; 3) the CD3-zeta does not associate with TCR alpha-CD3 gamma delta epsilon or TCR beta-CD3 gamma delta epsilon complexes; 4) CD3-zeta associate with the pentameric form of the TCR/CD3 complex in the endoplasmic reticulum to form the heptameric complex (TCR alpha beta-CD3 gamma delta epsilon----TCR alpha beta-CD3 gamma delta epsilon 2); and 5) CD3-zeta is required for the export of the TCR/CD3 complex from the endoplasmic reticulum to the Golgi apparatus for subsequent processing.     

New approach for selective separation of dilute products from simulated clostridial fermentation broths using cyclodextrins

The ability of cyclodextrins (CD) to form crystalline insoluble complexes with organics was explored in this study in view of a selective separation of dilute products obtainable from three clostridial fermentation systems. To this purpose, a product or a product mixture at a concentration of 0.150 mM each were treated with alpha-CD or beta-CD (0.150 mM) in aqueous solutions as well as in a nutrient broth as a simulated fermentation medium. In the acetone-butanol-ethanol system, and in the butanol-iso-propanol system, alpha-CD was found to precipitate selectively 48% and 46% butanol after 1 h agitation at 30 degrees C. However, beta-CD was found to be superior for the butyric acid-acetic acid system because it selectively precipitated 100% butyric acid under the same conditions. Cooling the three product system with alpha-CD to 4 degrees C for 24 h increased significantly the crop of precipitates but decreased the selectivity for either butanol or butyric acid. Cyclodextrins were thus shown to offer potentially a new exciting possibility for downstream processing of low-concentration fermentation products.     

Kinetic and thermodynamic analysis of the cyclodextrin-allyl isothiocyanate inclusion complex in an aqueous solution

The decomposition of allyl isothiocyanate (AITC) in an aqueous solution was depressed in the presence of cyclodextrin (CD), it's suppression effect increasing in the order of none < beta-CD < alpha-CD. The results of kinetic and thermodynamic analysis of the CD-AITC inclusion complexes showed that the inclusion process was mostly governed by an enthalpy change (delta H degree) rather than by an entrophy change (delta S degree), and that Van der Waals forces played a primary role int he inclusion. Steric factors were important for the reaction activity of AITC inclusion into the CD cavity, especially significant being the stereospecificity between the size of the CD cavity and the AITC molecule which is the main factor concerning it's activity. Our results suggest that the association stability and activity of the included AITC molecule are important factors in the suppression mechanism for CDs. Therefore, both these factors would make an alpha-CD-AITC system more advantageous than a beta-CD-AITC system, and the marked suppression effect of alpha-CD on the decomposition of AITC can be attributed to the formation of inclusion complexes in an aqueous solution.     

Solubility of cyclomaltooligosaccharides (cyclodextrins) in H2O and D2O: a comparative study

Cyclomaltooligosaccharides (cyclodextrins, CDs) are cyclic oligomers having six, seven, or eight units of alpha-D-glucose, named as cyclomaltohexaose (alpha-CD), cyclomaltoheptaose (beta-CD) and cyclomaltooctaose (gamma-CD), respectively. The molecule of CD has a cavity in which the interior is hydrophobic relative to its outer surface. The solubility of cyclodextrins in water is unusual, as an irregular trend is observed in the series of the cyclic oligomers of glucose. beta-CD is at least nine times less soluble than the others CDs. This intriguing behavior has been investigated, and some interesting explanations in terms of the effect caused by CD on the water lattice structure have been proposed. In this work a comparative study on the solubility of alpha, beta, and gamma-cyclodextrins was carried out in H2O and D2O and reveals a much lower solubility of the three CDs in D2O. The solid-phase structure of the CDs in equilibrium with the solution is quite similar with both solvents. The results are discussed in terms of the CD molecular structure and the differences in the hydrogen bonds formed between H2O and D2O.     

Quantitative analysis of natural cyclodextrins by high-performance liquid chromatography with pulsed amperometric detection: application to cell permeation study

Simple HPLC-PAD methods were developed for quantitation of cyclodextrins (CDs) in aqueous matrices from in vitro cell permeation studies. C-18 solid-phase extraction was used for sample pretreatment. Samples were analysed using acetonitrile-water mobile phase with post-column alkalization by 0.5M NaOH. Zorbax SB-Aq (for alpha-CD) and Zorbax SB-Phenyl (for beta-CD and gamma-CD) columns gave excellent peak shape and sufficient resolution of CD to glucose (2.7-3.2). The methods showed good concentration-response relationship (r > or = 0.999), precision (RSD% 0.7-5.1), repeatability (RSD% 3.4-13.7) and accuracy (87-107%). The limits of quantitation were 0.78, 0.46 and 0.52 microg/ml for alpha-CD, beta-CD and gamma-CD (RSD% of 10.6, 8.1 and 16.3, respectively).     

Chlorin p6 as a fluorescent probe for the investigation of surfactant-cyclodextrin interactions.

Cyclodextrins (CD) are often proposed as potential vehicles in targeted drug delivery. However, if the membrane structure is disrupted by CD, then it cannot be considered to be a good drug delivery vehicle. When an extrinsic fluorescence probe is used to monitor such interactions, there are no less than three possible equilibria that can operate simultaneously: surfactant-cyclodextrin, surfactant-fluorophore and cyclodextrin-fluorophore. The fluorescence intensity/lifetime might be affected by all these and so, the results depend strongly on the fluorophore used as well as the nature of the surfactant. This aspect highlights the importance of the suitability of the fluorescence probe to be used to study complicated systems and interaction. In the present work, chlorin p6, prepared from chlorophyll from spinach leaves, has been used as the fluorescence probe to investigate the interaction between alpha-CD and beta-CD with the neutral surfactants Triton X 100 (TX 100) and cetyl trimethyl ammonium bromide (CTAB). The fluorophore is found to be a sensitive one for the study of the interaction of alpha, beta and gamma-CD with the surfactants TX 100 and CTAB. It is found that contrary to earlier reports, a complex between alpha-CD and TX 100 is formed, even though the binding constant is not very high. This observation can be obtained with chlorin p6, which does not bind to the CDs, but not with a fluorophore, which binds to the CD as well and thus complicates the situation as the binding with CD is stronger than that between TX 100 and alpha-CD as compared to that between TNS and CD.     

Separation of cefoperazone enantiomers using beta-cyclodextrin as chiral additive by capillary zone electrophoresis

A capillary electrophoresis method was developed to separate the enantiomers of cefoperazone. Different cyclodextrins, including alpha-cyclodextrin (alpha-CD), beta-cyclodextrin (beta-CD), gamma-cyclodextrin (gamma-CD), 2-hydroxypropyl-beta-cyclodextrin (HP-beta-CD), and methyl-beta-cyclodextrin (Me-beta-CD), were tested as chiral additives in the running buffer. The effect of various parameters on enantioseparation such as concentration of NaH(2)PO(4), buffer pH, and CD concentration was also studied. The cefoperazone enantiomers were baseline separated under conditions of 0.04 mmol/L beta-CD, 75 mmol/L NaH(2)PO(4) buffer at pH 4.0. A fused silica capillary (40 cm effective length x 75 microm ID) was used. The applied voltage and capillary temperature were 20 kV and 25 degrees C, respectively. Under these conditions, linear calibration curves were obtained in the 5-500 microg/ml range using UV detection at 280 nm. The limit of detection for both isomers was 0.1 microg/ml. The method was used for the analysis of different pharmaceutical preparations (dose) and biological samples containing cefoperazone.     

The first chemical synthesis of a cyclodextrin heteroduplex

The synthesis of the first heteroduplex of cyclodextrin (CD) 11, i.e., a compound in which the two primary rims of alpha- and beta-CDs are doubly connected, was achieved. The selected strategy involved a Sonogashira coupling of propargylated beta-CD 6 and iodo-alkenyl alpha-CD 4 to singly connect the two CDs. A ring-closing metathesis (RCM) of the heterodimer 9 afforded the second bridge, final deprotection and reductions giving access to 11.     

Complexation of polymethine dyes with human serum albumin: a spectroscopic study

Non-covalent interactions between polymethine dyes of various types (cationic and anionic thiacarbocyanines as well as anionic oxonols and tetracyanopolymethines) and human serum albumin (HSA) were studied by means of absorption, fluorescence and circular dichroism (CD) spectroscopies. Complexation with the protein leads to a red shift of the dye absorption spectra and, in most cases, to a growth of the fluorescence quantum yield (Phif; for oxonols this growth is very small). The binding constants (K) obtained from changing the absorption spectra and Phif vary from 10(4) to (5-6) x 10(7) M(-1). K for the anionic dyes is much higher than for the cationic dyes (the highest K was found for oxonols). Interaction of meso-substituted anionic thiacarbocyanines with HSA results in cis-->trans isomerization and, as a consequence, an appearance and a steep rise of dye fluorescence. Binding to HSA gives rise to dye CD signals and in many cases is accompanied by aggregation of the dyes. These aggregates often exhibit biphasic CD spectra. The aggregates formed by the dyes alone are decomposed in the presence of HSA.     

Crystalline beta-cyclodextrin.12H2O reversibly dehydrates to beta-cyclodextrin.10.5 H2O under ambient conditions.

In contact with mother liquor, crystalline beta-cyclodextrin (beta-CD) hydrate has composition approximately beta-CD.12H2O. If crystals are dried at ambient conditions (18 degrees C, approximately 50% humidity), the unit cell volume diminishes approximately 30 to 50 A3. X-ray structure analysis of a dry crystal (0.89 A resolution, 4617 data, R = 0.059) showed the composition beta-CD.10.5 H2O, with approximately 5.5 water molecules in the beta-CD cavity (7 partially and 2 fully occupied sites) and approximately 5.0 between the beta-CD molecules. The positions of the beta-CD host and of most of the hydration waters are conserved during dehydration, but the occupancies of the waters in the beta-CD cavity diminish. Dry crystals put into solvent re-hydrate to the original form. The mechanism of de- and re-hydration is not evident.     

Characterization of thermostable cyclodextrinase from Clostridium thermohydrosulfuricum 39E.

Clostridium thermohydrosulfuricum 39E produced a cell-bound cyclodextrin (CD)-degrading enzyme (cyclodextrinase). It was partially purified 205-fold (specific activity, 14.5 U/mg of protein) by solubilizing with Triton X-100, ammonium sulfate treatment, and DEAE-Sepharose CL-6B column chromatography. The enzyme activity was found to be stable at pH 5.5 and 60 degrees C and optimally active at pH 6.0 and 65 degrees C. The enzyme preparation hydrolyzed CDs, with alpha-CD greater than beta-CD greater than gamma-CD, and displayed a putative multiple attack pattern. The enzyme activity was inhibited by p-chloromercuribenzoate but not by N-bromosuccinimide.     

Characterization of thermostable cyclodextrinase from Clostridium thermohydrosulfuricum 39E

Clostridium thermohydrosulfuricum 39E produced a cell-bound cyclodextrin (CD)-degrading enzyme (cyclodextrinase). It was partially purified 205-fold (specific activity, 14.5 U/mg of protein) by solubilizing with Triton X-100, ammonium sulfate treatment, and DEAE-Sepharose CL-6B column chromatography. The enzyme activity was found to be stable at pH 5.5 and 60 degrees C and optimally active at pH 6.0 and 65 degrees C. The enzyme preparation hydrolyzed CDs, with alpha-CD greater than beta-CD greater than gamma-CD, and displayed a putative multiple attack pattern. The enzyme activity was inhibited by p-chloromercuribenzoate but not by N-bromosuccinimide.     

Retarding effects of cyclodextrins on the decomposition of organic isothiocyanates in an aqueous solution

One- and two-dimensional (1)H nuclear magnetic resonance ((1)H NMR) spectra were measured in order to estimate the dissociation constants (K(diss)) and molecular geometries of cyclodextrin (CD) with three organic isothiocyanates (ITC), allyl-ITC, 3-butenyl-ITC, and 4-pentenyl-ITC, in an aqueous solution (pH 5.0, I(c)=0.75 M). In every ITC, the K(diss) values decreased in the order of alpha-CD>beta-CD>gamma-CD, and the three rod-like linear molecules of ITCs were accommodated with the best fit into the smallest cavity of alpha-CD. By rotating-frame nuclear Overhauser enhancement spectroscopy, cross peaks were found between the H-3 of alpha-CD and the H-1 of AITC. From the result, the molecular geometry of alpha-CD-AITC complex presume that the isothiocyanate group of AITC is located some where around the widening rim and a hydrophobic allyl group is included within the hydrophobic cavity of alpha-CD.     

Cyclodextrin production by cyclodextrin glycosyltransferase from Bacillus circulans DF 9R

Cyclodextrins (CD) are cyclic oligosaccharides with multiple applications in the food, pharmaceutical, cosmetic, agricultural and chemical industries. In this work, the conditions used to produce CD with cyclodextrin glycosyltransferase from Bacillus circulans DF 9R were optimized using experimental designs. The developed method allowed the partial purification and concentration of the enzyme from the cultural broth and, subsequently, the CD production, using the same cassava starch as enzyme adsorbent and as substrate. Heat-treatment of raw starch at 70 degrees C for 15 min in the presence of adsorbed cyclodextrin glycosyltransferase allowed the starch liquefaction without enzyme inactivation. The optimum conditions for CD production were: 5% (w/v) cassava starch, 15 U of enzyme per gram of substrate, reaction temperature of 56 degrees C and pH 6.4. After 4h, the proportion of starch converted to CD reached 66% (w/w) and the weight ratio of alpha-CD:beta-CD:gamma-CD was 1.00:0.70:0.16.     

Spectral properties and inclusion of a hetero-chalcone analogue in organized media of micellar solutions and beta-cyclodextrin

The absorption and fluorescence emission spectral properties of 3-(4'-dimethylaminophenyl)-1-(2-thienyl)prop-2-en-1-one, abbreviated as DMATP, have been investigated in organized media of aqueous micellar and beta-cyclodextrin (CD) solutions. While the absorption spectra are less sensitive to the nature of the added surfactant or CD, the characteristics of the intramolecular charge transfer (ICT) fluorescence are highly sensitive to the properties of the medium. The ICT maximum is strongly blue-shifted with a great enhancement in the fluorescence quantum yield on adding micellar or CD solutions. This indicates the solubilization of DMATP in the micellar core and formation of an inclusion complex with beta-CD. The critical micelle concentration (CMC) as well as the polarity of the micellar core of SDS, CTAB and TX-100 have been determined. The CMC values are in good agreement with the reported values while the polarity is lower indicating that DMATP molecules are incorporated in the micellar core not at the micellar interface. The inclusion constants of binding of DMATP in micellar or CD have been also determined. The thermodynamic parameters of formation of DMATP:CD inclusion complex have been calculated from the temperature dependence of the fluorescence spectra of the formed complex. The negative enthalpy and free energy of formation indicate that the inclusion process is energetically favorable. The highly negative value of formation entropy (DeltaS = -162.3 J mol(-1) K(-1)) reflects the high restrictions imposed on the movement of both the host and included guest molecules which is consistent with the increase of the fluorescence yield and blue shift of the fluorescence maximum.     

Study on effects of cyclodextrins on the photolysis of dissolved anthracene by fluorometry.

A simple, fast and effective fluorescence method for studying on photolysis of dissolved anthracene (An) in cyclodextrins (CDs) aqueous solutions was developed. With this method the effects of five kinds of cyclodextrin on photolysis of dissolved An were investigated. It was found that all of the CDs promoted the photolysis of An, due to the incorporation of An into the cavity of CDs, in which self-quenching of triplet-state An was inhibited and the lifetime of singlet-state An was prolonged. Hence, the possibility of the attack of oxygen on An was enhanced and more singlet-state oxygen was produced. The effect of cavity size and steric hindrance of substituents of CDs on photolysis of An was explored. The order of the promotion effect for the five kinds of CDs was as follows: HPCD(hydroxypropyl-beta-CD) > DMCD(heptakis(2,6-di-O-methyl)-beta-CD > beta-CD > gamma-CD > alpha-CD.     

Supramolecular Polypseudorotaxanes composed of star-shaped porphyrin-cored poly(epsilon-caprolactone) and alpha-cyclodextrin

Star-shaped porphyrin-cored poly(epsilon-caprolactone) (SPPCL) was synthesized using a tetrahydroxyethyl-terminated porphyrin as a core initiator and stannous octoate as a catalyst in bulk at 120 degrees C. The molecular weight of as-synthesized polymer could be adjusted linearly by controlling the molar ratio of epsilon-caprolactone to porphyrin core initiator, and the molecular weight distribution was reasonably narrow. Supramolecular polypseudorotaxanes were prepared by inclusion complexation of SPPCL with alpha-cyclodextrin (alpha-CD) and thoroughly characterized by means of FT-IR, 1H NMR, 13C CP/MAS NMR, DSC, TGA, and WAXD. The results demonstrated that the porphyrin-cored polypseudorotaxanes formed through alpha-CD molecules threading onto the branch chains of star-shaped SPPCL polymers, and they had a channel-type crystalline structure. Meanwhile, the original crystallization of SPPCL polymers within the polypseudorotaxanes was completely suppressed in the alpha-CD cavities. Moreover, inclusion complexation between SPPCL and alpha-CD enhanced the thermal stability of both the guest SPPCL polymers and the host alpha-CD. Furthermore, both the SPPCL polymers and the polypseudorotaxanes showed similar fluorescent and UV-vis spectra compared with porphyrin core initiator. Consequently, this will not only provide potentially porphyrin-cored poly(epsilon-caprolactone) and its polypseudorotaxanes for photodynamic therapy but also improve the compatibility between poly(epsilon-caprolactone) and peptide drugs for drug delivery.     

Inclusion complexes of bisphenol A with cyclomaltoheptaose (beta-cyclodextrin): solubilization and structure

The inclusion complexation behavior and the solubilization effects of Bisphenol A (BPA, an endocrine-disrupting chemical) by cyclomaltohexaose, -heptaose, and -octaose (alpha-, beta-, and gamma-cyclodextrins) were investigated by (1)H NMR spectroscopy and by elemental analysis. The results showed that beta- and gamma-cyclodextrins gave the satisfactory solubilization ability to BPA up to 7.2x10(3)mgL(-1) and 9.0x10(3)mgL(-1), respectively. X-ray crystallographic diffraction and ROESY spectroscopy were also employed to investigate the structure of the beta-CD/BPA inclusion complex in both aqueous solution and the solid state. The result showed that this complex adopted a 2:2 stoichiometry in the solid state, that is, a head-to-head beta-CD dimer accommodated two BPA molecules. The inclusion of BPA led to the desolvation of the beta-CD cavity and the destruction of the circularly closed hydrogen-bond network in the secondary side of beta-CD, which made the complex more soluble.