Molecular modelling of the inclusion complexes between β-cyclodextrin and (R)/(S)-methylphenobarbitone and its application to HPLC

Molecular modelling of β-cyclodextrin and optimisation of its potential energy suggests that a favoured conformation is that distorted from a symmetrical torus. The inclusion of water molecules into the torus cavity simulates the increased stability in an aqueous solvent. Complexes of β-cyclodextrin with (R)- and (S)-enantiomers of methylphenobarbitone have been modelled and energetically optimised by the application of molecular mechanics. The simulations suggests that the guest molecules adopt an orientation in which the phenyl ring is projected into the torus cavity, with in each case the plane of the ring parallel to a longer axis of the distorted torus and slightly displaced from the axis through the torus cavity. It is suggested that the asymmetry in the macrocyclic ring contributes to chiral recognition as a result of additional discriminatory binding to the barbiturate ring residue of each enantiomer, which occupy different 3D geometries. The enantiomers form complexes of different minimum potential energies. The resulting difference in complex stability can be related to the behaviour of β-cyclodextrin, as a mobile phase additive in reverse-phase HPLC to effect chiral separation of rac-medthylphenobarbitone during chromatography. © 1994 Wiley-Liss, Inc.     

A highly sensitive probe detecting low pH area of HeLa cells based on rhodamine B modified β-cyclodextrins

Two kinds of rhodamine modified β-cyclodextrins (R-1 and R-2), which are coupled up ethylene diamine (EDA) and tetraethylene pentamine (TEPA) between Rh B and β-cyclodextrin, respectively, have been synthesized. R-1 and 2 work as a new fluorogenic probe for monitoring pH of Hela cells, and MTT of assay R-1, R-2, and rhodamine B indicate that less a cytotoxicity of those R-1 and R-2 than that of rhodamine B, where R-1 has much less one than that of R-2. The fluorogenetic probe capability of R-2 was recognized in an area of acidic area in living cell, which is lysosome.     

High-performance liquid chromatographic study of the interactions between immobilized β-cyclodextrin polymers and hydrophobically end-capped polyethylene glycols

The formation of inclusion complexes between polyethylene glycols (PEGs) bearing hydrophobic ends (naphtyl and phenyladamantyl) and β-cyclodextrin polymers (polyβ-CD) immobilized onto silica particles was studied by high-performance liquid chromatography (HPLC). It was shown that hydrophobic interactions were involved in the retention mechanism of these compounds, since retention volumes decreased when organic solvents were added to the mobile phase while it was the contrary in the presence of salts. Moreover, the association could be reversed by adding a competitor (hydroxypropylβ-cyclodextrin) to the mobile phase. A theoretical model permitted the evaluation of affinity constants of 1:1 complexes formed between the modified PEGs and the immobilized polyβ-CD which depended on the type of hydrophobic groups grafted to the PEG.     

Polar organic chiral separation of propranolol and analogues using a β-cyclodextrin bonded stationary phase

A β-cyclodextrin bonded stationary phase was employed for the enantioresolution of propranolol and several analogues in conjunction with various polar organic mobile phases. The effects of structural alterations in the non-polar regions of the analytes were found to exert profound changes upon chiral resolution and capacity values, indicating that features which cannot hydrogen-bond with the cyclodextrin molecule still play an important role in this chiral recognition process. This was linked to a repulsive steric effect facilitating the necessary conditions for chiral resolution. The degree of ionization of the analytes and the type and concentration of organic modifier used were also seen to influence the analytes¹ enantio-selectivity and capacity values. © 1996 Wiley-Liss, Inc.     

A novel preparation of methyl-β-cyclodextrin from dimethyl carbonate and β-cyclodextrin

A novel green synthesis process about methyl-β-cyclodextrin has been investigated through the reaction between β-cyclodextrin and dimethyl carbonate by anhydrous potassium carbonate as catalyst in DMF. The influence of experimental factors including the molar ratio of dimethyl carbonate to β-cyclodextrin, reaction temperature, and reaction time on the average degree of substitution of methyl-β-cyclodextrin was studied. The results show that the average degree of substitution of methyl-β-cyclodextrin can be dependent on the reaction temperature and the molar ratio of raw material primarily. The structures of methyl-β-cyclodextrin were characterized by TLC, IR, MS, ¹H NMR, and ¹³C NMR.     

Separating enantiomers by HPLC on silica dynamically coated with a chiral stationary phase of permethylated β-cyclodextrin

The paper demonstrates that the technique of solvent generated liquid--solid chromatography can be used to create normal phase systems for chiral separations. The chiral adsorption layer was generated by pumping a binary hexane:ethanol eluent containing a small fraction of permethylated β-cyclodextrin through a column packed with microparticulate silica. This technique leads to columns with good time stability and reproducibility. The possibility of generating normal phase systems with permethylated β-cyclodextrin as chiral component via the mobile phase broadens the range of phase system which can be used to separate enantiomers by HPLC.     

Quantification of trace levels of β-cyclodextrin and substitution patterns in hydroxypropyl-β-cyclodextrin using high-performance liquid chromatography with polarimetric detection

A high-performance liquid chromatographic (HPLC) method has been developed for separation and determination of components in hydroxypropyl-β-cyclodextrin (HP-β-CD). The method involves separation on an amino-bonded HPLC column using water–acetonitrile as a mobile phase with a polarimetric HPLC detector for quantification. It provides good selectivity and sensitivity and can also be used to compare different sources of HP-β-CD and to measure batch to batch variation. The similarity of the values of molar optical rotation for β-cyclodextrin (β-CD) and HP-β-CD suggests that a polarimetric HPLC detector may be used with a straightforward area normalization method, to quantify the proportion of β-CD in any HP-β-CD sample. Trace amounts of β-CD in HP-β-CD have been measured to a precision of 0.01%. © 1993 Wiley-Liss, Inc.     

γ-Cyclodextrin as chiral mobile phase additive in the HPLC separation of the atropisomers of some N-arylthiazoline-2-thiones and N-arylthiazoline-2-ones: Attempts to quantify the effect of selected structural parameters

The effect of the position and type of the substituent on the chromatographic separation of N-arylthiazoline-2-thione and arylthiazoline-2-one atropisomers are described in reversed-phase HPLC using β- or -β-cyclodextrin as chiral mobile phase additive. A quantitative approach to experimental design has been developed. © 1993 Wiley-Liss, Inc.     

H NMR study of rehydration/dehydration and water mobility in β-cyclodextrin

The processes of dehydration and rehydration of β-cyclodextrin were studied by analysis of the ¹H NMR (nuclear magnetic resonance) line shape. Dehydration was carried in an open ampoule as a function of temperature and above 400 K total dehydration of β-cyclodextrin was observed. This result was confirmed by the thermogravimetry (TG) measurements. Rehydration was studied as a function of time at room temperature. After 40 days, β-cyclodextrin was found to absorb eight water molecules. The analysis of temperature changes in the shape of the ¹H NMR line of β-cyclodextrin kept in a closed ampoule and its dielectric measurements provided information on the mobility of water molecules. The water molecules were found to perform complex molecular motions, that is, reorientational jumps below 200 K and additionally, translational motion (diffusion) above 200 K.     

Formulation and Development of Extended-Release Micro Particulate Drug Delivery System of Solubilized Rifaximin

Rifaximin (RFX), a semi-synthetic antibiotic belonging to BCS class IV category, has been used in the treatment of traveler’s diarrhea. An attempt has been made to improve aqueous solubility of RFX in the presence of β-cyclodextrin (β-CD) and hydroxy propyl β-cyclodextrin (HP-β-CD) and control its release in the gut by enteric coating. The stoichiometric proportion of RFX and complexing agent’s β-CD and HP-β-CD were determined by phase solubility studies. RFX–β-CD and RFX–HP-β-CD were prepared in 1:2 ratio by solvent evaporation technique using rota-evaporator with yield of 78% and 84% respectively followed by their evaluation using different techniques such as saturation solubility, Fourier transform infrared, differential scanning calorimeter, powder X-ray diffractometer, in vitro antimicrobial activity. The saturation solubility of RFX had improved from 0.0736 mg/ml to 0.2354 mg/ml and 0.5681 mg/ml in presence of β-CD and HP-β-CD respectively resulting in an increased zone of inhibition in the later complex during antimicrobial studies. The RFX–HP-β-CD complex particles were coated with eudragit L 100 (EL 100) by spray drying technique. The 3² factorial design was applied to formulate the micro particles. All formulations exhibited pH dependant drug release. The % EE was 69% and the release of RFX was retarded by enteric coating in the optimized batch FB2. Therefore, it can be concluded that solubility of some BCS class IV drugs can be improved by β-CD complexation and release of such inclusion complexes can be retarded to increase the residence time of RFX in the gastrointestinal tract.     

Use of chiral β-lactones for the synthesis of chiral LC phases

Chiral β-lactones offer an easy and economic approach to novel chiral R- or S-configurated LC phases. The preferred method for their preparation is based on conventional ready-to-use aminopropyl-functionalized silicas, e.g., aminopropyl HPLC columns. The new stationary phases can be used for analytical and preparative separations and are particularly suitable for the resolution of rotatory and heterocyclic stereoisomers. Applications in the LC, SFC, and TLC mode are possible, including the use of β-lactone-based mobile phase additives in a “push–pull” combination. A large variety of organic solvents can be used as eluents. © 1993 Wiley-Liss, Inc.     

Ring-Toss: Capping highly exposed tyrosyl or tryptophyl residues in proteins with β-cyclodextrin

We have used UV difference spectroscopy and fluorescence spectroscopy to study the perturbation by β-cyclodextrin of tyrosyl or tryptophyl residues located at each of the 10 variable consensus contact positions in the third domain of turkey ovomucoid. The goal was to monitor the accessibility of the side chain rings of these residues when located at these positions. The results indicated that the tyrosyl or tryptophyl rings are most highly exposed when located in the P₁ position followed by the P₄ position. It was possible to determine the association constants for β-cyclodextrin binding at these positions. When located at the P₂, P₅, P₆ and P₃′ positions, the rings of the tyrosyl or tryptophyl residues were exposed but less so than at the P₁ or P₄ positions. By contrast, when located at the P₁′, P₂′, P₁₄^′ and P₁₈^′ positions, the tyrosyl or tryptophyl residues were insufficiently exposed to be perturbed by β-cyclodextrin, although they reacted positively to dimethyl sulfoxide solvent perturbation. These findings indicate that β-cyclodextrin perturbation provides a convenient way to detect highly exposed tyrosyls or tryptophyls in proteins. Furthermore, we evaluated the ability of β-cyclodextrin to inhibit the interaction of turkey ovomucoid third domain variants with different P₁ residues. The results showed that the presence of β-cyclodextrin had little effect on the association constant when the P₁ residue was a glycyl residue, but greatly decreased the association constant when the P₁ residue was a tyrosyl or tryptophyl residue. Thus, β-cyclodextrin may be used to selectively modulate the interaction between proteinase inhibitors and their cognate enzymes.     

Determination of the enantiomeric purity of commercial Jacobsen's catalyst samples

A HPLC method is described for the chiral analysis of the commercially available Jacobsen's catalyst. A hydroxypropyl β-cyclodextrin stationary phase was used in conjunction with a nonaqueous, polar-organic mobile phase. The method can be applied to control the enantiomeric purity of the catalyst, which is of great importance for quality control of that product. High accuracy in the determination of trace levels of the unwanted enantiomer in the presence of large amounts of the desired enantiomer is demonstrated. Chirality 10:362–363, 1998. © 1998 Wiley-Liss, Inc.     

Binding of a chromen-4-one Schiff’s base with bovine serum albumin: capping with β-cyclodextrin influences the binding

This work deals with the synthesis of 6-methyl-3-[(4′-methylphenyl)imino]methyl-4H-chromen-4-one (MMPIMC), its binding to β-cyclodextrin, and the influence of the cyclodextrin complexation on the compound’s binding to bovine serum albumin (BSA). The 1:2 stoichiometry for the complexation of MMPIMC with β-cyclodextrin is determined with the binding constant of 1.90 × 10⁴ M^?2. The structure of host–guest complex plays a role in protein binding of MMPIMC. One- and two-dimensional NMR spectra are used to determine the mode of binding of the guest to β-cyclodextrin cavity and the structure of the inclusion complex is proposed. The binding of MMPIMC with BSA in the absence and the presence of β-cyclodextrin is studied. The binding strengths of MMPIMC–BSA (1.73 × 10⁵ M^?1) and β-cyclodextrin-complexed MMPIMC–BSA (9.0 × 10⁴ M^?1) show difference in magnitude. The Förster Resonance Energy Transfer efficiency and the proximity of the donor and acceptor molecules, are modulated by β-cyclodextrin. Molecular modeling is used to optimize the sites and mode of binding of MMPIMC with bovine serum albumin.     

Study on the Interaction between the Inclusion Complex of Hematoxylin with β-Cyclodextrin and DNA

Ultraviolet-visible (UV-vis) spectra, fluorescence spectra, electrochemistry, and the thermodynamic method were used to discuss the interaction mode between the inclusion complex of hematoxylin with β-cyclodextrin and herring sperm DNA. On the condition of physiological pH, the result showed that hematoxylin and β-cyclodextrin formed an inclusion complex with binding ratio n_hematoxylin:n_{β-cyclodextrin} = 1:1. The interaction mode between β-cyclodextrin-hematoxylin and DNA was a mixed binding, which contained intercalation and electrostatic mode. The binding ratio between β-cyclodextrin-hematoxylin and DNA was n_{β-cyclodextrin -hematoxylin}:n_DNA = 2:1, binding constant was K^? _298.15K = 5.29 × 10⁴ L·mol^?1, and entropy worked as driven force in this action.     

Absorption and excretion of tablet disintegrating β-cyclodextrin polymer in rats

Absorption and excretion of a new tablet disintegrating agent, a crosslinked β-cyclodextrin polymer was investigated following per os administration in the rat. The polymer, which is insoluble but swells in water, was prepared from β-cyclodextrin by reacting with [2-¹⁴C]epichlorohydrin in an alkaline medium. Radioactivity of blood, urine, faeces, exhaled carbon dioxide and the gastrointestinal tract was determined by a liquid scintillation method. No radioactivity could be detected in the blood up to 24 h after the administration of the polymer. Radioactivity of urine and exhaled carbon dioxide together did not exceed 0·11% of the total administered radioactivity, 98% of which was found in the large intestine and the faeces. Therefore, it is assumed that β-cyclodextrin polymer could not be absorbed from the gastrointestinal tract.     

Impact of β-cyclodextrin and resistant starch on bile acid metabolism and fecal steroid excretion in regard to their hypolipidemic action in hamsters1

To examine the impact on bile acid metabolism and fecal steroid excretion as a mechanism involved in the lipid-lowering action of β-cyclodextrin and resistant starch in comparison to cholestyramine, male golden Syrian hamsters were fed 0% (control), 8% or 12% of β-cyclodextrin or resistant starch or 1% cholestyramine. Resistant starch, β-cyclodextrin and cholestyramine significantly lowered plasma total cholesterol and triacylglycerol concentrations compared to control. Distinct changes in the bile acid profile of gallbladder bile were caused by resistant starch, β-cyclodextrin and cholestyramine. While cholestyramine significantly reduced chenodeoxycholate independently of its taurine–glycine conjugation, β-cyclodextrin and resistant starch decreased especially the percentage of taurochenodeoxycholate by ?75% and ?44%, respectively. As a result, the cholate:chenodeoxycholate ratio was significantly increased by 100% with β-cyclodextrin and by 550% with cholestyramine while resistant starch revealed no effect on this ratio. β-Cyclodextrin and resistant starch, not cholestyramine, significantly increased the glycine:taurine conjugation ratio demonstrating the predominance of glycine conjugated bile acids. Daily fecal excretion of bile acids was 4-times higher with 8% β-cyclodextrin and 19-times with 1% cholestyramine compared to control. β-Cyclodextrin and cholestyramine also induced a 2-fold increase in fecal neutral sterol excretion, demonstrating the sterol binding capacity of these two compounds. Resistant starch had only a modest effect on fecal bile acid excretion (80% increase) and no effect on excretion of neutral sterols, suggesting a weak interaction with intestinal steroid absorption. These data demonstrate the lipid-lowering potential of β-cyclodextrin and resistant starch. An impaired reabsorption of circulating bile acids and intestinal cholesterol absorption leading to an increase in fecal bile acid and neutral sterol excretion is most likely the primary mechanism responsible for the lipid-lowering action of β-cyclodextrin. In contrast, other mechanisms involving the alterations in the biliary bile acid profile or repressed hepatic lipogenesis, e.g., VLDL production, appear to be involved in the hypolipidemic effect of resistant starch.     

β-cyclodextrin production by the cyclodextrin glucanotransferase from Paenibacillus illinoisensis ZY-08: cloning, purification, and properties

The gene encoding the cyclodextrin glucanotransferase (CGTase, EC2.4.1.19) of Paenibacillus illinoisensis was isolated, cloned, sequenced and expressed in Escherichia coli. Sequence analysis showed that the mature enzyme (684 amino acids) was preceded by a signal peptide of 34-residues. The deduced amino acid sequence of the CGTase from P. illinoisensis ZY-08 exhibited highest identity (99 %) to the CGTase sequence from Bacillus licheniformis (P14014). The four consensus regions of carbohydrate converting domain and Ca²⁺ binding domain could be identified in the sequence. The CGTase was purified by using cold expression vector, pCold I, and His-tag affinity chromatography. The molecular weight of the purified enzyme was about 74 kDa. The optimum temperature and pH of the enzyme were 40 °C and pH 7.4, respectively. The enzyme activity was increased by the addition of Ca²⁺ and inhibited by Ba²⁺, Cu²⁺, and Hg²⁺. The K _m and V _max values calculated were 0.48 mg/ml and 51.38 mg of β-cyclodextrin/ml/min. The ZY-08 and recombinant readily converted soluble starch to β-cyclodextrin but ZY-08 did not convert king oyster mushroom powder and enoki mushroom powder. However the recombinant CGTase converted king oyster mushroom powder and enoki mushroom powder to β-cyclodextrin.     

Agglomerated Oral Dosage Forms of Artemisinin/β-Cyclodextrin Spray-Dried Primary Microparticles Showing Increased Dissolution Rate and Bioavailability

Artemisinin, a poorly water-soluble antimalarial drug, presents a low and erratic bioavailability upon oral administration. The aim of this work was to study an agglomerated powder dosage form for oral administration of artemisinin based on the artemisinin/β-cyclodextrin primary microparticles. These primary microparticles were prepared by spray-drying a water–methanol solution of artemisinin/β-cyclodextrin. β-Cyclodextrin in spray-dried microparticles increased artemisinin water apparent solubility approximately sixfold. The thermal analysis evidenced a reduction in the enthalpy value associated with drug melting, due to the decrease in drug crystallinity. The latter was also evidenced by powder X-ray diffraction analysis, while ¹³C-NMR analysis indicated the partial complexation with β-cyclodextrin. Agglomerates obtained by sieve vibration of spray-dried artemisinin/β-cyclodextrin primary microparticles exhibited free flowing and close packing properties compared with the non-flowing microparticulate powder. The in vitro dissolution rate determination of artemisinin from the agglomerates showed that in 10 min about 70% of drug was released from the agglomerates, whereas less than 10% of artemisinin was dissolved from raw material powder. Oral administration of agglomerates in rats yielded higher artemisinin plasma levels compared to those of pure drug. In the case of the agglomerated powder, a 3.2-fold increase in drug fraction absorbed was obtained.     

The Combined Usage of β-Cyclodextrin and Milk Proteins in Microencapsulation of Bifidobacterium bifidum BB-12

The present study aimed to determine the effects of combined usage of β-cyclodextrin with whey protein isolate and sodium caseinate on the microencapsulation of Bifidobacterium bifidum-BB12 by spray drying.

From the results, the highest count of B. bifidum was provided by whey protein isolate as 8.62 log CFU/g. The increasing concentration of β-cyclodextrin considerably increases gastric and intestinal resistance to B. bifidum cells. In the gastric and intestinal test, the highest protection was determined in whey protein isolate substituted with 10% β-cyclodextrin with reduction rates of 0.98 and 3.30%, respectively. Moreover, free cells did not survive in the same gastric conditions. The lowest hygroscopicity was determined in whey protein isolate as 8.57%. It must be noted that increasing β-cyclodextrin concentration in carrier material combination led to an increase in hygroscopicity of microcapsules. In general, substitution with β-cyclodextrin increased the particle size of microparticles, and microcapsules produced with whey protein isolate had a smaller size than that of sodium caseinate.