Development of pullulanase mutants to enhance starch substrate utilization for efficient production of ß-CD.

The inhibitory effect of ß-CD on pullulanase which hydrolyzes α-1,6 glycosidic bond in starch to release more available linear substrates, limited substrate utilization thus influencing the yield of ß-CD. Here, an aspartic acid residue (D465) which interacted with cyclodextrin ligand by hydrogen bond, was mutated to explore its contribution to bind inhibitors and obtain mutants with lower affinity to ß-CD. Enzyme activity results showed that mutants D465E and D465N retained higher activity than wild-type pullulanase in presence of 10 mM ß-CD. Circular dichroism spectra and fluorescence spectra results showed that D465 was related to structure stability. Chain length distribution results confirmed the improvement of substrate utilization by the addition of D465E. The conversion rate from potato starch, cassava starch, and corn starch into ß-CD, increased to 56.9%, 55.4% and 54.7%, respectively, when synchronous using ß-CGTase and D465E in the production process.

In vivo preclinical evaluation of the new 68Ga-labeled beta-cyclodextrin in prostaglandin E2 (PGE2) positive tumor model using positron emission tomography.

The cyclooxygenase-2 (COX-2)/prostaglandin E2 (PGE2) pathway plays an important role in tumor development and formation of metastases. It was earlier reported that cyclodextrin derivatives have a high affinity to form complexes with PGE2. Based on these results radiolabeled cyclodextrins - as new radiopharmaceuticals - may open a new pathway in the in vivo imaging and diagnosis of PGE2 positive tumors. The aims of this study were to synthetize the PGE2 specific 68Ga-labeled NODAGA-randomly methylated beta-cyclodextrin (68Ga-NODAGA-RAMEB) and investigate its tumor-targeting properties. NODAGA-RAMEB was labeled with Gallium-68 (68Ga), and the radiochemical purity (RCP%), partition coefficient (logP values), and in vitro-in vivo stability of 68Ga-NODAGA-RAMEB were determined. After intravenous injection of 68Ga-NODAGA-RAMEB the accumulation in organs and tissues was monitored in vivo by positron emission tomography (PET) and ex vivo by gamma counter in BxPC-3 and PancTu-1 tumor-bearing CB17 SCID mice. The RCP% of the newly synthesized 68Ga-NODAGA-RAMEB was higher than 98%. The molar activity was 15.34 ± 1.93 GBq/µmol. The logP of 68Ga labeled NODAGA-RAMEB was - 3.63 ± 0.04. Biodistribution studies showed high accumulation of 68Ga-NODAGA-RAMEB in PGE2 positive BxPC-3 tumors; approximately 15-20-fold higher radiotracer uptake was observed, than that of the background. 68Ga-labeled RAMEB is a promising radiotracer in PET diagnostics of PGE2 positive tumors.

Involvement of the PKA pathway and inhibition of voltage gated Ca2+ channels in antihyperalgesic activity of Lippia grata/ß-cyclodextrin.

Neuropathic pain (NP) is a difficult condition to treat because of the modest efficacy of available drugs. New treatments are required. In the study we aimed to investigate the effects of the essential oil from Lippia grata alone or complexed in ß-cyclodextrin (LG or LG-ßCD) on persistent inflammatory and neuropathic pain in a mouse model. We also investigated Ca2+ currents in rat dorsal root ganglion (DRG) neurons. Male Swiss mice were treated with LG or LG/ß-CD (24 mg/kg, i.g.) and their effect was evaluated using an acute inflammatory pleurisy model and nociception triggered by intraplantar injection of an agonist of the TRPs channels. We also tested their effect in chronic pain models: injection of Freund's Complete Adjuvant and partial sciatic nerve ligation (PSNL). In the pleurisy model, LG reduced the number of leukocytes and the levels of TNF-α and IL-1ß. It also inhibited cinnamaldehyde and menthol-induced nociceptive behavior. The pain threshold in mechanical and thermal hyperalgesia was increased and paw edema was decreased in models of inflammatory and neuropathic pain. PSNL increased inflammatory protein contents and LG and LG-ßCD restored the protein contents of TNF-α, NF-κB, and PKA, but not IL-1ß and IL-10. LG inhibited voltage gated Ca2+ channels from DRG neurons. Our results suggested that LG or LG-ßCD produce anti-hyperalgesic effect in chronic pain models through reductions in TNF-α levels and PKA, and inhibited voltage-gated calcium channels and may be innovative therapeutic agents for the management of NP.

The nature of ß-cyclodextrin inhibition of potato polyphenol oxidase-catalyzed reactions.

There is general interest in strategies to control polyphenol oxidase (PPO)-initiated enzymatic browning because it is often associated with declining food quality. Cyclodextrins are cyclic glucan oligosaccharides that form inclusion complexes with a number of PPO substrates. This study focuses on the effect of ß-cyclodextrins (ßCyD) on PPO-catalyzed reactions. Potato enzyme extracts and semi-purified potato PPO served as enzyme sources. Substrates included phenolics endogenous to potatoes. Reaction time-courses were followed spectrophotometrically; rates were compared by analysis of variance. Extents of ßCyD inhibition of PPO-catalyzed reactions are shown to be substrate specific and can be quantitatively accounted for based on degrees of ßCyD substrate sequestration. There was no evidence for direct irreversible ßCyD inactivation of potato PPO. An apparent "direct PPO inactivation" by ßCyD is shown to result from a sequence of sequestration-dependent reactions that occur in commonly employed assay systems for the quantification of PPO in fruits and vegetables.

Construction of a small-caliber tissue-engineered blood vessel using icariin-loaded ß-cyclodextrin sulfate for in situ anticoagulation and endothelialization.

The rapid endothelialization of tissue-engineered blood vessels (TEBVs) can effectively prevent thrombosis and inhibit intimal hyperplasia. The traditional Chinese medicine ingredient icariin is highly promising for the treatment of cardiovascular diseases. ß-cyclodextrin sulfate is a type of hollow molecule that has good biocompatibility and anticoagulation properties and exhibits a sustained release of icariin. We studied whether icariin-loaded ß-cyclodextrin sulfate can promote the endothelialization of TEBVs. The experimental results showed that icariin could significantly promote the proliferation and migration of endothelial progenitor cells; at the same time, icariin could promote the migration of rat vascular endothelial cells (RAVECs). Subsequently, we used an electrostatic force to modify the surface of the TEBVs with icariin-loaded ß-cyclodextrin sulfate, and these vessels were implanted into the rat common carotid artery. After 3 months, micro-CT results showed that the TEBVs modified using icariin-loaded ß-cyclodextrin sulfate had a greater patency rate. Scanning electron microscopy (SEM) and CD31 immunofluorescence results showed a better degree of endothelialization. Taken together, icariin-loaded ß-cyclodextrin sulfate can achieve anticoagulation and rapid endothelialization of TEBVs to ensure their long-term patency.

Amisulpride-CD-Loaded Liposomes: Optimization and In Vivo Evaluation.

Amisulpride (AMS) is an atypical antipsychotic agent used for the treatment of schizophrenia. The effect of different variables, i.e., the type of cyclodextrins (CDs), ratio of drug/CDs, and type of loading on the prepared AMS-CD liposomes (single and double loaded) was studied by applying 23 full factorial design. Double-loaded liposomes are loaded with AMS-hydroxyl propyl-ß-cyclodextrin (HP-ß-CD) in the aqueous phase and free drug in the lipophilic bilayer, while single-loaded liposomes are loaded only with AMS-HP-ß-CD in the aqueous phase. Entrapment efficiency, particle size, polydespersibility, and zeta potential were selected as dependent variables. Design Expert® software was used to obtain an optimized formulation with high entrapment efficiency (64.55 ± 1.27%), average particle size of 40.1 ± 2.77 nm, polydespersibility of 0.44 ± 0.37, and zeta potential of - 48.8 ± 0.28. Optimized formula was evaluated for in vitro release, surface morphology and stability study was also conducted. AMS-HP-ß-CD in double-loaded liposomes exhibited higher drug release than those in the conventional liposomes and in the single-loaded liposomes. The maximum plasma concentration (Cmax) of AMS in optimized AMS-HP-ß-CD double-loaded liposomal formulation increased by 1.55- and 1.29-fold, as compared to the commercial tablets and conventional liposomes, respectively. However, the relative bioavailability of AMS double-loaded liposomes was 1.94- and 1.28-folds of commercial tablet and conventional liposomes, respectively.

Dietary α-cyclodextrin reduces atherosclerosis and modifies gut flora in apolipoprotein E-deficient mice.

SCOPE: α-Cyclodextrin (α-CD), a cyclic polymer of glucose, has been shown to lower plasma cholesterol in animals and humans; however, its effect on atherosclerosis has not been previously described. METHODS AND RESULTS: apoE-knockout mice were fed either low-fat diet (LFD; 5.2% fat, w/w), or Western high fat diet (21.2% fat) containing either no additions (WD), 1.5% α-CD (WDA); 1.5% ß-CD (WDB); or 1.5% oligofructose-enriched inulin (WDI). Although plasma lipids were similar after 11 weeks on the WD vs. WDA diets, aortic atherosclerotic lesions were 65% less in mice on WDA compared to WD (P < 0.05), and similar to mice fed the LFD. No effect on atherosclerosis was observed for the other WD supplemented diets. By RNA-seq analysis of 16S rRNA, addition of α-CD to the WD resulted in significantly decreased cecal bacterial counts in genera Clostridium and Turicibacterium, and significantly increased Dehalobacteriaceae. At family level, Comamonadaceae significantly increased and Peptostreptococcaceae showed a negative trend. Several of these bacterial count changes correlated negatively with % atherosclerotic lesion and were associated with increased cecum weight and decreased plasma cholesterol levels. CONCLUSION: Addition of α-CD to the diet of apoE-knockout mice decreases atherosclerosis and is associated with changes in the gut flora.

Defective Cytochrome P450-Catalysed Drug Metabolism in Niemann-Pick Type C Disease.

Niemann-Pick type C (NPC) disease is a neurodegenerative lysosomal storage disease caused by mutations in either the NPC1 or NPC2 gene. NPC is characterised by storage of multiple lipids in the late endosomal/lysosomal compartment, resulting in cellular and organ system dysfunction. The underlying molecular mechanisms that lead to the range of clinical presentations in NPC are not fully understood. While evaluating potential small molecule therapies in Npc1-/- mice, we observed a consistent pattern of toxicity associated with drugs metabolised by the cytochrome P450 system, suggesting a potential drug metabolism defect in NPC1 disease. Investigation of the P450 system in the context of NPC1 dysfunction revealed significant changes in the gene expression of many P450 associated genes across the full lifespan of Npc1-/- mice, decreased activity of cytochrome P450 reductase, and a global decrease of multiple cytochrome P450 catalysed dealkylation reactions. In vivo drug metabolism studies using a prototypic P450 metabolised drug, midazolam, confirmed dysfunction in drug clearance in the Npc1-/- mouse. Expression of the Phase II enzyme uridinediphosphate-glucuronosyltransferase (UGT) was also significantly reduced in Npc1-/- mice. Interestingly, reduced activity within the P450 system was also observed in heterozygous Npc1+/- mice. The reduced activity of P450 enzymes may be the result of bile acid deficiency/imbalance in Npc1-/- mice, as bile acid treatment significantly rescued P450 enzyme activity in Npc1-/- mice and has the potential to be an adjunctive therapy for NPC disease patients. The dysfunction in the cytochrome P450 system were recapitulated in the NPC1 feline model. Additionally, we present the first evidence that there are alterations in the P450 system in NPC1 patients.

Development of a high-throughput assay for measuring phospholipase A activity using synthetic 1,2-α-eleostearoyl-sn-glycero-3-phosphocholine coated on microtiter plates.

To date, several sensitive methods, based on radiolabeled elements or sterically hindered fluorochrome groups, are usually employed to screen phospholipase A (PLA) activities. With the aim of developing a convenient, specific, sensitive, and continuous new ultraviolet (UV) spectrophotometric assay for PLA, we have synthesized a specific glycerophosphatidylcholine (PC) esterified at the sn-1 and sn-2 positions, with α-eleostearic acid (9Z, 11E, 13E-octadecatrienoic acid) purified from Aleurites fordii seed oil. The conjugated triene present in α-eleostearic acid constitutes an intrinsic chromophore and, consequently, confers the strong UV absorption properties of this free fatty acid as well as of the glycerophospholipids harboring it. This coated PC film cannot be desorbed by the various buffers used during PLA assays. Following the action of PLA at the oil-water interface, α-eleostearic acid is freed and desorbed from the film and then solubilized with ß-cyclodextrin. The UV absorbance of the α-eleostearic acid is considerably enhanced due to the transformation from an adsorbed to a water-soluble state. The PLA activity can be measured continuously by recording the variations with time of the UV absorption spectra. The rate of lipolysis was monitored by measuring the increase of absorption at 272 nm, which was found to be linear with time and proportional to the amount of added PLA. This continuous high-throughput PLA assay could be used to screen new PLA and/or PLA inhibitors present in various biological samples.

Inhibitory effect of Angelica sinensis extract in the presence of 2-hydroxypropyl-ß-cyclodextrin.

Angelica sinensis (AS) is a traditional Chinese medicinal herb. Its ethanolic extract contains active ingredients, such as ferulic acid, ligustilide, and butylidenephthalide, which are hydrophobic and have inhibitory effects on hepatoma cells. To increase the aqueous solubility/dispersibility of AS extract and study the consequent inhibitory effect on hepatoma cells, the ethanolic extract of AS was complexed with 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD), a cyclic oligosaccharide that has a hydrophilic outer surface and a hydrophobic central cavity. The AS-HP-ß-CD complex (weight ratio of AS extract: HP-ß-CD=1:5) was prepared and characterized. The effect of complexing the AS extract with HP-ß-CD on Hep3B cell growth was investigated by analyzing cytotoxicity. Our results showed that cytotoxicity inhibition of AS-HP-ß-CD complex was up to 94% and higher than that of AS extract (about 68%). These observations suggested that the use of HP-ß-CD to stabilize AS extract in aqueous solution was possible for herbal medicine application.

Regulation of the high-affinity choline transporter activity and trafficking by its association with cholesterol-rich lipid rafts.

The sodium-coupled, hemicholinium-3-sensitive, high-affinity choline transporter (CHT) is responsible for transport of choline into cholinergic nerve terminals from the synaptic cleft following acetylcholine release and hydrolysis. In this study, we address regulation of CHT function by plasma membrane cholesterol. We show for the first time that CHT is concentrated in cholesterol-rich lipid rafts in both SH-SY5Y cells and nerve terminals from mouse forebrain. Treatment of SH-SY5Y cells expressing rat CHT with filipin, methyl-ß-cyclodextrin (MßC) or cholesterol oxidase significantly decreased choline uptake. In contrast, CHT activity was increased by addition of cholesterol to membranes using cholesterol-saturated MßC. Kinetic analysis of binding of [(3)H]hemicholinium-3 to CHT revealed that reducing membrane cholesterol with MßC decreased both the apparent binding affinity (KD) and maximum number of binding sites (Bmax ); this was confirmed by decreased plasma membrane CHT protein in lipid rafts in cell surface protein biotinylation assays. Finally, the loss of cell surface CHT associated with lipid raft disruption was not because of changes in CHT internalization. In summary, we provide evidence that CHT association with cholesterol-rich rafts is critical for transporter function and localization. Alterations in plasma membrane cholesterol cholinergic nerve terminals could diminish cholinergic transmission by reducing choline availability for acetylcholine synthesis. The sodium-coupled choline transporter CHT moves choline into cholinergic nerve terminals to serve as substrate for acetylcholine synthesis. We show for the first time that CHT is concentrated in cholesterol-rich lipid rafts, and decreasing membrane cholesterol significantly reduces both choline uptake activity and cell surface CHT protein levels. CHT association with cholesterol-rich rafts is critical for its function, and alterations in plasma membrane cholesterol could diminish cholinergic transmission by reducing choline availability for acetylcholine synthesis.

Effects of inulin and di-D-fructose dianhydride-enriched caramels on intestinal microbiota composition and performance of broiler chickens.

In vitro and in vivo experiments were designed to evaluate the effectiveness of laboratory-made di-d-fructose dianhydride (DFA)-enriched caramels. The DFA-enriched caramels were obtained from d-fructose (FC), d-fructose and sucrose (FSC), or d-fructose and ß-cyclodextrin (FCDC). In the in vitro experiment, raftilose and all caramels increased (P<0.05) l-lactate concentration and decreased (P<0.05) pH. Total short-chain fatty acid concentration was higher (P<0.05) than controls in tubes containing raftilose, FSC, FCDC and commercial sucrose caramel (CSC). Raftilose, and all caramels tested except FSC and FC (1%), increased (P<0.01) lactobacilli log10 number of copies compared with the non-additive control. FSC, FCDC and CSC increased (P<0.01) the bifidobacteria number of copies as compared with controls. All additives, except FCDC, decreased (P<0.01) Clostridium coccoides/Eubacterium rectale log number of copies. Compared with controls, raftilose, FC and CSC led to lower (P<0.01) Escherichia-Shigella and enterobacteria. For the in vivo experiment, a total of 144 male 1-day-old broiler chickens of the Cobb strain were randomly assigned to one of the three dietary treatments for 21 days. Dietary treatments were control (commercial diet with no additive), inulin (20 g inulin/kg diet) and FC (20 g FC/kg diet). Final BW of birds fed FC diet was higher (P<0.01) than controls or inulin-fed birds, although feed: gain values were not different. Feed intake of chickens fed FC was higher (P<0.01) than that of inulin-fed birds but not statistically different from controls. Crop pH values were lower (P<0.01) in birds fed FC diet as compared with control diet, with inulin-fed chickens showing values not different from control- or FC-fed birds. Lower (P<0.05) lactobacilli number of copies was determined in the crop, ileum and caeca of birds fed the inulin diet compared with the control diet. Inulin supplementation also resulted in lower (P<0.05) C. coccoides/E. rectale, bacteroides and total bacteria in caecal contents. Addition of FC to broiler diets gave place to lower (P<0.05) enterobacteria and Escherichia-Shigella in crop and caecal contents compared with controls. The bacteroides number of copies increased (P<0.05) as compared with controls in the ileum, but decreased (P<0.05) in the caeca of chickens fed the FC diet. Energy, ADF, NDF and non-starch polysaccharides faecal digestibilities were greater (P<0.05) than controls in chickens fed diets containing inulin or FC. Fat digestibility was higher (P<0.05) in FC-fed birds compared with controls or inulin-fed chickens. In conclusion, DFA-enriched caramels tested here, particularly FC, may represent a type of new additives useful in poultry production.

Development of low methoxy amidated pectin-based mucoadhesive patches for buccal delivery of triclosan: effect of cyclodextrin complexation.

A novel mucoadhesive buccal patch formulation of triclosan (TR), a broad spectrum antibacterial agent, was developed using low methoxy amidated pectin (AMP). The integrity of AMP matrix was improved by addition of 20% (w/w) Carbopol (CAR). The efficiency of ß-cyclodextrin-epichlorohydrin polymer (EPIßCD) and anionic carboxymethylated ß-cyclodextrin-epichlorohydrin polymer (CMEPIßCD) in optimization of TR solubility and release from such a matrix was investigated and confronted to that of parent ß-cyclodextrin (ßCD). Loading of TR/ßCD co-ground complex into AMP/CAR matrix resulted in a biphasic release profile which was sensitive upon the hydration degree of the matrix, due to lower solubilizing efficiency of ßCD, while the drug release from patches loaded with TR/EPIßCD complex was significantly faster with a constant release rate. Microbiological studies evidenced faster onset and more pronounced antibacterial action of TR/EPIßCD loaded patches, clearly demonstrating their good therapeutic potential in eradication of Streptococcus mutans, a cariogenic bacteria, from the oral cavity.

Preparation, characterization, and in vitro intestinal permeability evaluation of thalidomide-hydroxypropyl-ß-cyclodextrin complexes.

Thalidomide is emerging as a therapeutic agent with renewed clinical importance, presenting anti-inflammatory, immunomodulatory, and antineoplasic properties. In this work, we studied the complexation of thalidomide with cyclodextrins as a strategy to circumvent the poor aqueous solubility of the drug. Thalidomide-hydroxypropyl-ß-cyclodextrin complexes were obtained by kneading method and were characterized by differential scanning calorimetry, powder X-ray diffractometry, and scanning electronic microscopy. The aqueous solubility and in vitro dissolution of thalidomide were significantly improved through the complexation. Physicochemical analysis of the complexes in solid state revealed a decreased crystallinity of the complexed drug in comparison with free thalidomide. Thalidomide was able to dissociate from the complexes and permeates across intestinal epithelial Caco-2 cells with a favorable high permeability profile equivalent to that of the free drug. In summary, the present results suggest that thalidomide-hydroxypropyl-ß-cyclodextrin complexes could be regarded as a promising strategy for improving the gastrointestinal absorption of thalidomide.

[Study on excretion of stilbene glycoside (THSG) and its beta-cyclodextrin inclusion].

The excretion characteristics of stilbene glycoside (THSG) and its beta-cyclodextrin inclusion in bile, urine and feces after oral administration to rats were studied. Bile for 24 h, urine and feces for 72 h were collected. The content of THSG was determined by HPLC-UV. The established HPLC-UV method was available for the analysis of THSG in excreta and corresponded to the requirement of biological sample analysis. After given THSG and its beta-cyclodextrin inclusion, the amount of prototype THSG in feces were 3.27% and 0.61%, meanwhile THSG in bile were 0.20% and 0.18%, respectively. Only a little THSG was found in urine. The result showed that beta-cyclodextrin inclusion reduced the fecal excretion of THSG. However, the characteristic of urinary and biliary excretion wasn't changed.

Sensory properties of ginseng solutions modified by masking agents.

Ginseng is one of the most popular functional ingredients found in energy drink formulations. Although ginseng is known for its health benefits, ginseng is also notorious for imparting a bitter taste. Incorporating ginseng into beverages without the bitterness, while still maintaining its health benefits, is necessary for developing an acceptable product. Thus, the objectives of this study were to (1) identify effective treatments for minimizing the bitterness of ginseng in water base and model energy drink base solutions and (2) determine the sensory effects of incorporating different treatment levels to minimize the bitterness of ginseng. A series of pilot studies investigating bitterness reducing treatments were conducted, which included: congruent flavor addition, bitterness blocking agent incorporation, enzymatic modification, ingredient interaction, and complexation. Based on the results of a series of pilot studies, γ-cyclodextrin (γ-CD) and ß-cyclodextrin (ß-CD) complexation agents were identified as having the most potential. Effectiveness of the γ-CDs, ß-CDs, and combinations of γ- and ß-CDs were tested in 100 mL water and in 100 mL model energy drink base solutions containing 0.052 g 80% ginsenosides panax ginseng, using descriptive sensory analysis. Twelve trained panelists evaluated 42 solution treatments (3 treatments × 7 levels × 2 bases) for bitter attributes with and without nose clips. Overall, the most effective treatments were 0.09 g γ-CDs in 100 mL of solution and 1 g ß-CDs in 100 mL solution, which both reduced the bitterness intensity of the solutions by half. Incorporation of these levels of CDs in water and model energy drink base solutions containing ginseng will aid in the development of functional beverages that are more acceptable to a wider range of consumers.

Pretreatment of Candida rugosa lipase with soybean oil before immobilization on beta-cyclodextrin-based polymer.

In this study, Candida rugosa lipase was pretreated with soybean oil in order to allow fatty acids to bond to the active site before immobilization. The pretreated C. rugosa lipase was immobilized on beta-cyclodextrin-based polymer by crosslinking with hexamethylene diisocyanate (HMDI). This pretreated lipase exhibited steric hindrance around the active site such that during immobilization, covalent bonds were formed between the carrier and the lipase region far from the active site. The pretreated lipase was immobilized on a beta-cyclodextrin-based polymer by crosslinking with hexamethylene diisocyanate with the activity of 42U/g-matrix, which is 5.7 times higher than that of the immobilized non-pretreated lipase. In addition, the immobilized lipase activity was maintained at levels exceeding 60% of its original activity after 10 reuses.

Microspheres for colonic delivery of ketoprofen-hydroxypropyl-beta-cyclodextrin complex.

A new multiparticulate system, with the potential for site-specific delivery to the colon, has been developed using ketoprofen as model drug. The system simultaneously exploits cyclodextrin complexation, to improve drug solubility, and vectorization in microspheres (MS) based on Ca-pectinate and chitosan. The effect of complexation with hydroxypropyl-beta-cyclodextrin (HPbetaCyd) and of chitosan presence on drug entrapment efficiency and release properties, as well on the drug permeation rate across Caco-2 cells has been investigated. Solid-state interactions between the components have been investigated by FTIR spectroscopy, differential scanning calorimetry and X-ray powder diffractometry. The morphology of MS was examined by scanning electron microscopy. Release studies revealed a different behaviour for MS containing drug alone or as complex: drug alone was released faster than in the presence of cyclodextrin from MS without chitosan, due to a reservoir effect. The opposite was found for MS containing chitosan, due to a competition effect between polymer and drug for the cyclodextrin. Cytotoxicity tests demonstrated the safety of these formulations. Permeation studies showed an increased permeation of the drug formulated as MS, particularly marked when it was used as complex, thus revealing an enhancing power of both cyclodextrin and chitosan with a synergistic effect in improving drug permeation.

A novel dicyclodextrinyl diselenide compound with glutathione peroxidase activity.

A 6A,6A'-dicyclohexylamine-6B,6B'-diselenide-bis-beta-cyclodextrin (6-CySeCD) was designed and synthesized to imitate the antioxidant enzyme glutathione peroxidase (GPX). In this novel GPX model, beta-cyclodextrin provided a hydrophobic environment for substrate binding within its cavity, and a cyclohexylamine group was incorporated into cyclodextrin in proximity to the catalytic selenium in order to increase the stability of the nucleophilic intermediate selenolate. 6-CySeCD exhibits better GPX activity than 6,6'-diselenide-bis-cyclodextrin (6-SeCD) and 2-phenyl-1,2-benzoisoselenazol-3(2H)-one (Ebselen) in the reduction of H(2)O(2), tert-butyl hydroperoxide and cumenyl hydroperoxide by glutathione, respectively. A ping-pong mechanism was observed in steady-state kinetic studies on 6-CySeCD-catalyzed reactions. The enzymatic properties showed that there are two major factors for improving the catalytic efficiency of GPX mimics. First, the substrate-binding site should match the size and shape of the substrate and second, incorporation of an imido-group increases the stability of selenolate in the catalytic cycle. More efficient antioxidant ability compared with 6-SeCD and Ebselen was also seen in the ferrous sulfate/ascorbate-induced mitochondria damage system, and this implies its prospective therapeutic application.

Biodegradation of cyclodextrins in soil.

Cyclodextrins, especially random methylated betaCD (RAMEB) and hydroxypropyl betaCD (HPbetaCD), are becoming common enhancing additives in the bioremediation of soils formerly contaminated by hydrocarbons and/or other poorly bioavailable organic pollutants. Therefore, their degradation in the soil, particularly the most persistent RAMEB, has been of great concern. Like oil contaminants, these additives should be biodegradable via an environmentally safe technology. Hence, in this paper, the biodegradability of eight different cyclodextrins (CDs) in four different soils was examined under various treatment conditions in laboratory and pilot scale field experiments. This paper is the first report on the potential biological fate of CDs studied under a large variety of environmental conditions and in different soil ecosystems. Data on the potential relationship between CD biodegradation and the biological removal of hydrocarbons in the CD-amended contaminated soils are also given. All CDs were found to be more or less biodegradable; even the most persistent RAMEB was depleted from soils under favourable conditions. In the field experiments, the depletion of RAMEB to about 40% of its initial level was observed for a period of 2 years in hydrocarbon-contaminated soils of high organic matter and cell concentration.