Cytoprotective effects of α-linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, oleic acid and α-tocopherol on 7-ketocholesterol - Induced oxiapoptophagy: Major roles of PI3-K / PDK-1 / Akt signaling pathway and glutathione peroxidase activity in cell rescue.

On murine N2a cells, 7-ketocholesterol induced an oxiapotophagic mode of cell death characterized by oxidative stress (reactive oxygen species overproduction on whole cells and at the mitochondrial level; lipid peroxidation), apoptosis induction (caspase-9, -3 and -7 cleavage, PARP degradation) and autophagy (increased ratio LC3-II / LC3-I). Oxidative stress was strongly attenuated by diphenyleneiodonium chloride which inhibits NAD(P)H oxidase. Mitochondrial and peroxisomal morphological and functional changes were also observed. Down regulation of PDK1 / Akt signaling pathways as well as of GSK3 / Mcl-1 and Nrf2 pathways were simultaneously observed in 7-ketocholesterol-induced oxiapoptophagy. These events were prevented by α-linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, oleic acid and α-tocopherol. The inhibition of the cytoprotection by LY-294002, a PI3-K inhibitor, demonstrated an essential role of PI3-K in cell rescue. The rupture of oxidative stress in 7-ketocholesterol-induced oxiapoptophagy was also associated with important modifications of glutathione peroxidase, superoxide dismutase and catalase activities as well as of glutathione peroxidase-1, superoxide dismutase-1 and catalase level and expression. These events were also counteracted by α-linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, oleic acid and α-tocopherol. The inhibition of the cytoprotection by mercaptosuccinic acid, a glutathione peroxidase inhibitor, showed an essential role of this enzyme in cell rescue. Altogether, our data support that the reactivation of PI3-K and glutathione peroxidase activities by α-linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, oleic acid and α-tocopherol are essential to prevent 7KC-induced oxiapoptophagy.

Activity of Na+/K+- and Ca2+-ATPases in human erythrocyte membranes: Protocol improvement, relation to cholesterol content, and effects of polyphenols.

It is known that the activities of Na+/K+- and Ca2+-ATPases in the plasma membrane with an excess of cholesterol are compromised. Our main goal was to find out whether quercetin, resveratrol, or caffeic acid, in the nano- and low micromolar concentration ranges, can improve the ATPase activity in human erythrocyte membranes with excess cholesterol. These molecules belong to different chemical classes of polyphenols and are widely present in plant foods. Also, due to some variations in the protocol for determining the ATPase activity, we first analyzed several key parameters of the protocol to improve the accuracy of the results. The activities of Na+/K+- and Ca2+-ATPases were reduced in membranes with moderate and high cholesterol levels compared to membranes from normocholesterolemic subjects (p < 0.01). All three polyphenols affected the ATPase activity in a similar biphasic manner. Namely, the ATPase activity gradually increased with increasing polyphenol concentration up to 80-200 nM, and then gradually decreased with further increase in polyphenol concentration. Moreover, the stimulating effect of the polyphenols was highest in membranes with high cholesterol content, making ATPase activity values close/equal to those in normal cholesterol membranes. In other words, quercetin, resveratrol, and caffeic acid at nanomolar concentrations were able to improve/restore the functioning of Na+/K+- and Ca2+-ATPases in erythrocyte membranes with high cholesterol levels. This suggests a common membrane-mediated mechanism of action for these polyphenols, related to the content of membrane cholesterol.

Prevention by Dietary Polyphenols (Resveratrol, Quercetin, Apigenin) Against 7-Ketocholesterol-Induced Oxiapoptophagy in Neuronal N2a Cells: Potential Interest for the Treatment of Neurodegenerative and Age-Related Diseases.

The Mediterranean diet is associated with health benefits due to bioactive compounds such as polyphenols. The biological activities of three polyphenols (quercetin (QCT), resveratrol (RSV), apigenin (API)) were evaluated in mouse neuronal N2a cells in the presence of 7-ketocholesterol (7KC), a major cholesterol oxidation product increased in patients with age-related diseases, including neurodegenerative disorders. In N2a cells, 7KC (50 µM; 48 h) induces cytotoxic effects characterized by an induction of cell death. When associated with RSV, QCT and API (3.125; 6.25 µM), 7KC-induced toxicity was reduced. The ability of QCT, RSV and API to prevent 7KC-induced oxidative stress was characterized by a decrease in reactive oxygen species (ROS) production in whole cells and at the mitochondrial level; by an attenuation of the increase in the level and activity of catalase; by attenuating the decrease in the expression, level and activity of glutathione peroxidase 1 (GPx1); by normalizing the expression, level and activity of superoxide dismutases 1 and 2 (SOD1, SOD2); and by reducing the decrease in the expression of nuclear erythroid 2-like factor 2 (Nrf2) which regulates antioxidant genes. QCT, RSV and API also prevented mitochondrial dysfunction in 7KC-treated cells by counteracting the loss of mitochondrial membrane potential (ΨΔm) and attenuating the decreased gene expression and/or protein level of AMP-activated protein kinase α (AMPKα), sirtuin 1 (SIRT1) and peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) implicated in mitochondrial biogenesis. At the peroxisomal level, QCT, RSV and API prevented the impact of 7KC by counteracting the decrease in ATP binding cassette subfamily D member (ABCD)3 (a peroxisomal mass marker) at the protein and mRNA levels, as well as the decreased expresssion of genes associated with peroxisomal biogenesis (Pex13, Pex14) and peroxisomal ß-oxidation (Abcd1, Acox1, Mfp2, Thiolase A). The 7KC-induced decrease in ABCD1 and multifunctional enzyme type 2 (MFP2), two proteins involved in peroxisomal ß-oxidation, was also attenuated by RSV, QCT and API. 7KC-induced cell death, which has characteristics of apoptosis (cells with fragmented and/or condensed nuclei; cleaved caspase-3; Poly(ADP-ribose) polymerase (PARP) fragmentation) and autophagy (cells with monodansyl cadaverine positive vacuoles; activation of microtubule associated protein 1 light chain 3-I (LC3-I) to LC3-II, was also strongly attenuated by RSV, QCT and API. Thus, in N2a cells, 7KC induces a mode of cell death by oxiapoptophagy, including criteria of OXIdative stress, APOPTOsis and autoPHAGY, associated with mitochondrial and peroxisomal dysfunction, which is counteracted by RSV, QCT, and API reinforcing the interest for these polyphenols in prevention of diseases associated with increased 7KC levels.

Prevention of 7-Ketocholesterol-Induced Overproduction of Reactive Oxygen Species, Mitochondrial Dysfunction and Cell Death with Major Nutrients (Polyphenols, ω3 and ω9 Unsaturated Fatty Acids) of the Mediterranean Diet on N2a Neuronal Cells.

The brain, which is a cholesterol-rich organ, can be subject to oxidative stress in a variety of pathophysiological conditions, age-related diseases and some rare pathologies. This can lead to the formation of 7-ketocholesterol (7KC), a toxic derivative of cholesterol mainly produced by auto-oxidation. So, preventing the neuronal toxicity of 7KC is an important issue to avoid brain damage. As there are numerous data in favor of the prevention of neurodegeneration by the Mediterranean diet, this study aimed to evaluate the potential of a series of polyphenols (resveratrol, RSV; quercetin, QCT; and apigenin, API) as well as ω3 and ω9 unsaturated fatty acids (α-linolenic acid, ALA; eicosapentaenoic acid, EPA; docosahexaenoic acid, DHA, and oleic acid, OA) widely present in this diet, to prevent 7KC (50 µM)-induced dysfunction of N2a neuronal cells. When polyphenols and fatty acids were used at non-toxic concentrations (polyphenols: ≤6.25 µM; fatty acids: ≤25 µM) as defined by the fluorescein diacetate assay, they greatly reduce 7KC-induced toxicity. The cytoprotective effects observed with polyphenols and fatty acids were comparable to those of α-tocopherol (400 µM) used as a reference. These polyphenols and fatty acids attenuate the overproduction of reactive oxygen species and the 7KC-induced drop in mitochondrial transmembrane potential (ΔΨm) measured by flow cytometry after dihydroethidium and DiOC6(3) staining, respectively. Moreover, the studied polyphenols and fatty acids reduced plasma membrane permeability considered as a criterion for cell death measured by flow cytometry after propidium iodide staining. Our data show that polyphenols (RSV, QCT and API) as well as ω3 and ω9 unsaturated fatty acids (ALA, EPA, DHA and OA) are potent cytoprotective agents against 7KC-induced neurotoxicity in N2a cells. Their cytoprotective effects could partly explain the benefits of the Mediterranean diet on human health, particularly in the prevention of neurodegenerative diseases.

Isocitrate dehydrogenase 1 mutation is associated with reduced levels of inflammation in glioma patients.

Background: Glioma patients with mutant isocitrate dehydrogenase have improved survival; this could be in part due to the suppressive effect of mutant IDH on the level of chronic inflammation. This study aimed to prospectively analyze the association of IDH1 mutation status with preoperative levels of blood inflammatory markers: neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR), C-reactive protein (CRP), and red cell distribution width (RDW) in gliomas. Patients and methods: Receiver operating characteristic curves for cutoff value determination, various bivariate tests, and survival analyses (Kaplan-Meier curves and Cox regression) were performed. Results: Patients with mutant IDH1 had reduced levels of NLR (P<0.032) and CRP (P<0.008). Moreover, these patients showed better median overall survival compared to those without IDH1 mutation (P<0.000). In univariate analysis, IDH1 mutation status (P<0.000), NLR (P<0.000), PLR (P<0.008), and CRP (P<0.001) were among the factors associated with survival. By multivariate analysis, IDH1 mutation (P<0.044) and NLR<2.65 (P<0.022) remained independent factors associated with better survival; other independent variables were tumor grade (P<0.000) and location in noneloquent area (P<0.015). Conclusion: The obtained results show that IDH1 mutation is associated with lower levels of chronic inflammation that could account for an improved prognosis in this group of patients.

Cyclodextrin-membrane interaction in drug delivery and membrane structure maintenance.

Cyclodextrins (CDs) are cyclic oligosaccharides able to improve drug water solubility and stability by forming CD/drug inclusion complexes. To further increase drug entrapment and delay its release, the CD/drug inclusion complex can be embedded in the aqueous phase of a liposome, a lipid vesicle composed of phospholipid bilayer surrounding an aqueous compartment. The resulting carrier is known as drug-in-cyclodextrin-in-liposome (DCL) system. CDs and DCLs are recognized as effective drug delivery systems; therefore, understanding the interaction of CDs with liposomal and biological membranes is of great importance. CDs are able to extract phospholipids, cholesterol, and proteins from membranes; the effect depends on the membrane structure and composition as well as on the CD type and concentration. Under definite conditions, CDs can affect the membrane fluidity, permeability, and stability of liposomes and cells, leading to the leakage of some of their internal constituents. On the other side, CDs demonstrated their beneficial effects on the membrane structure, including preservation of the membrane integrity during freeze-drying. In this paper, we review the literature concerning the interaction of CDs with biomimetic and biological membranes. Moreover, the impact of CDs on the membrane properties, mainly fluidity, stability, and permeability, is highlighted.

First evaluation of drug-in-cyclodextrin-in-liposomes as an encapsulating system for nerolidol.

Nerolidol, a naturally occurring sesquiterpene with antimicrobial activities, is a promising candidate as a natural alternative for synthetic preservatives in food. However, its application is limited by low aqueous solubility and stability. In this study, conventional liposomes and drug-in-cyclodextrin-in-liposomes (DCLs) were evaluated for the first time as encapsulating materials for nerolidol. The size, encapsulation efficiency (EE%), loading rate (LR%), photo- and storage stabilities of both systems were characterized. Moreover, the in vitro release of nerolidol from liposomes and DCLs was investigated over time. Nerolidol was efficiently entrapped in both carriers with high EE% and LR% values. In addition, DCLs prolonged the release of nerolidol over one week and enhanced the photostability more effectively than conventional liposomes. Finally, all formulations were stable after 12 months of storage at 4 °C (>60% incorporated nerolidol). Therefore, DCLs are promising carriers for new applications of sesquiterpenes in the pharmaceutical and food industries.

Nootkatone encapsulation by cyclodextrins: Effect on water solubility and photostability.

Nootkatone (NO) is a sesquiterpenoid volatile flavor, used in foods, cosmetics and pharmaceuticals, possessing also insect repellent activity. Its application is limited because of its low aqueous solubility and stability; this could be resolved by encapsulation in cyclodextrins (CDs). This study evaluated the encapsulation of NO by CDs using phase solubility studies, Isothermal Titration Calorimetry, Nuclear Magnetic Resonance spectroscopy and molecular modeling. Solid CD/NO inclusion complex was prepared and characterized for encapsulation efficiency and loading capacity using UV-Visible. Thermal properties were investigated by thermogravimetric-differential thermal analysis and release studies were performed using multiple headspace extraction. Formation constants (Kf) proved the formation of stable inclusion complexes. NO aqueous solubility, photo- and thermal stability were enhanced and the release could be insured from solid complex in aqueous solution. This suggests that CDs are promising carrier to improve NO properties and, consequently, to enlarge its use in foods, cosmetics, pharmaceuticals and agrochemicals preparations.

The effect of cyclodextrin complexation on the solubility and photostability of nerolidol as pure compound and as main constituent of cabreuva essential oil.

Nerolidol (Ner), a major component of many plant essential oils, is known for its various biological properties. However, the low solubility of Ner in water and its susceptibility to degradation limit its application. The aim of our study was to improve the solubility and photostability of Ner through its encapsulation in different cyclodextrins (CDs). The formation constants of cis-, trans-Ner and their commercial mixture with various CDs (α-CD, ß-CD, γ-CD, HP-ß-CD, RAMEB, CRYSMEB and SBE-ß-CD) were determined by phase solubility studies and confirmed by the spectral displacement UV-visible method. The solubility of cabreuva essential oil (EO) rich in trans-Ner was also evaluated by total organic carbon (TOC) analysis. The encapsulation efficiency (EE %) of Ner in HP-ß-CD solid complexes was assessed by HPLC. The structural characterization of CD/trans-Ner inclusion complex was then conducted by NMR spectroscopy followed by molecular modelling studies. The effect of encapsulation on the Ner photostability was also carried out over time under UVB irradiation. AL-type phase-solubility diagrams were obtained, suggesting the formation of 1:1 CD/Ner inclusion complexes. The solubility of Ner was enhanced by approximately 70-fold in the presence of 10 mM HP-ß-CD. Moreover, high EE % values were obtained for 5:1 and 10:1 HP-ß-CD:Ner molar ratios. NMR and molecular modelling studies revealed the most stable structure for trans-Ner inside the CD cavity with the OH group oriented towards the wider rim of the CD. Finally, CD encapsulation of Ner as pure compound or as main component of the cabreuva EO, protected it from degradation. This effect was more pronounced as the concentration of CD increased. These findings suggested that CDs are promising encapsulating carriers for Ner by enhancing its solubility and stability and thereby its application in food industry.

Interaction of Selected Phenylpropenes with Dipalmitoylphosphatidylcholine Membrane and Their Relevance to Antibacterial Activity.

The effect of structurally closely related phenylpropenes (PPs), estragole, anethole, eugenol, and isoeugenol, on the fluidity of dipalmitoyl phosphatidyl choline (DPPC) liposome membrane was investigated by DSC, Raman, and fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene (DPH). Liposomes were prepared by thin-film hydration method at various DPPC:PP molar ratios. The DPH anisotropy measurements of blank and PP-loaded liposomes were performed at 28, 41, and 50 °C, which correspond, respectively, to gel phase, main transition temperature of DPPC, and liquid phase. The Raman images showed the formation of nano- and micrometric spherical multi-lamellar vesicles. All studied PPs exhibited a membrane fluidizing effect which was reinforced by the presence of phenolic hydroxyl group in eugenol and isoeugenol. The PPs interacted with the choline head group and the alkyl chains of DPPC membrane, wherein isoeugenol and anethole possessing the same C7-C8 position of the double bond in the propenyl side chain, incorporated deeply in the bilayer. Additionally, the PPs were analyzed for antibacterial activity against E. coli by macrobroth dilution method. Anethole and estragole were more efficient in inhibiting the bacterial growth than eugenol and isoeugenol. We conclude that the fluidizing effect of PPs on the membrane is a common mechanism that is not related to the hydrophobicity of the PP molecule. Besides, other target sites may be involved in PP antibacterial activity against Gram-negative bacteria. The greater hydrophobicity of these PPs may contribute to their penetrability through the outer bacterial membrane.

Drug-in-cyclodextrin-in-liposomes as a carrier system for volatile essential oil components: Application to anethole.

A combined approach based on cyclodextrin/drug inclusion complex formation and loading into liposomes was applied to improve the effectiveness of liposome loading with essential oils. Hydroxypropyl-ß-cyclodextrin/ANE (HP-ß-CD/ANE) inclusion complexes were prepared and encapsulated into liposomes (ACL). ANE-double-loaded liposomes (ACL2) were obtained with the HP-ß-CD/ANE complex in the aqueous phase and ANE in the organic phase. Liposomes were prepared from saturated (Phospholipon 90H) or unsaturated (Lipoid S100) phospholipids and characterized for size, polydispersity index, zeta potential, morphology, loading rate (LR) and photo- and storage stabilities. All liposome batches were nanometric oligolamellar-type vesicles. Compared to ANE-loaded liposomes, ACL-90H, ACL2-90H and ACL2-S100 displayed significantly increased ANE LR, with ACL2-S100 exhibiting the highest LR. All formulations provided ANE photoprotection, were physically stable after 15months of storage at 4°C (with the exception of ACL2-S100), and retained more than 25% of the ANE initially present in the liposome suspensions.

Association of preoperative levels of selected blood inflammatory markers with prognosis in gliomas.

BACKGROUND: Red cell distribution width (RDW), neutrophil-lymphocyte ratio (NLR), and platelet count (PLT) routinely tested as part of the complete blood count are indicative of systemic inflammation. The prognostic significance of NLR and PLT in cancer was demonstrated in many studies while the role of RDW has been hardly investigated. The present study aimed to assess the association of RDW, NLR, and PLT with survival and tumor grade in glioma patients. METHODS: Clinical data from 178 patients with primary gliomas treated in a single institution were retrospectively analyzed. Receiver operating characteristic curves for cutoff value determination, Kaplan-Meier survival analysis, various bivariate tests, and univariate and multivariate Cox regression analyses were performed. RESULTS: Patients with high RDW (≥13.95) and NLR (≥4) levels had worse overall survival (OS) (Wilcoxon test, P<0.026 and P<0.003, respectively) while the effect of thrombocytosis (≥400×109/L) on prognosis was not significant. Besides, a strong association between RDW and NLR was found (Spearman's rho =0.230, P<0.02; χ2=8.887, P<0.03; Mann-Whitney U-test, P<0.017). Moreover, RDW and NLR were significantly associated with tumor grade. In univariate Cox analysis, elevated NLR (hazard ratio, HR 1.385; confidence interval, CI 1.020-1.881, P<0.037), older age (HR 0.452, CI 0.329-0.621, P<0), and higher tumor grade (HR 1.624, CI 1.187-2.223, P<0.002) were associated with poor outcomes. In the multivariate analysis, tumor grade, age, and Karnofsky performance score were identified as being independently prognostic for OS. CONCLUSION: Preoperative NLR and RDW values can help to evaluate disease progression and outcomes in patients with gliomas, thereby contributing to patient follow-up optimization.

Preparation of drug-in-cyclodextrin-in-liposomes at a large scale using a membrane contactor: Application to trans-anethole.

The present study aimed to prepare liposomes loaded with cyclodextrin/drug inclusion complexes at a pilot scale based on the ethanol injection technique. Anethole (ANE), a major component of anise and fennel essential oils, was used as a model of a volatile and highly hydrophobic drug. Membrane contactor (600mL) and a pilot plant (3L) were used for liposome production. The liposome preparations obtained were characterized for size, polydispersity index, zeta potential, morphology, stability and ANE release rate. All experimental set-ups were shown to be appropriate for the preparation of small, multilamellar vesicles with narrow size distribution and good stability at 4°C. The drug release study showed that only a small amount of ANE was released from liposome formulations after 21days of storage at 4°C. The loading rate of ANE was higher when ethanol was evaporated directly on the pilot plant compared to a rotary evaporation.

Release studies of trans-anethole from ß-cyclodextrin solid inclusion complexes by Multiple Headspace Extraction.

This study aimed to evaluate the effect of the preparation method, temperature and humidity on the release of aroma from ß-cyclodextrin (ß-CD) solid inclusion complexes (IC). Therefore ß-CD/trans-anethole (ß-CD/AN) IC were prepared by freeze-drying (FD) and co-precipitation coupled to FD (Cop-FD). Release experiments were performed at various temperatures and relative humidities (RH). Multiple headspace extraction-gas chromatography (MHE) was used to determine the loading capacity (LC) and encapsulation efficiency (EE%) and perform release studies. Results underlined that the quantification of encapsulated AN by MHE requires the IC dissolution. The release of AN was accelerated by increases in RH and temperature. However, it was quite negligible below 75% RH. The release behavior of AN was well simulated by Avrami's equation. Cop-FD IC retained more efficiently AN and the release depended on the preparation method and treatment conditions. Thus, the preparation method could be chosen based on the application.

Chemical Composition, Antioxidant and Cytotoxic Activities of Roots and Fruits of Berberis libanotica.

Fourteen compounds belonging to different chemical classes were characterized in the roots and fruits extracts from Berberis libanotica, using the same HPLC-DAD-MS method. Thirteen were reported, for the first time, from the fruits whereas the roots contained mostly alkaloids of which 3 out of 5 are reported for the first time. Their structures were established on the basis of MS data as gallic acid (1), chlorogenic acid (2), delphinidin (3), oxyacanthine (4), rutin (5), hyperoside (6), berbamine (7), isoquercitrin (8), quercitrin (9), jatrorrhizine (10), palmatine (11), berberine (12), quercetin (13) and luteolin (14). Extracts containing compounds 4 and 7 showed significant cytotoxicity against the HT29 cell line with an IC50 of 12.2-26.1 µg/mL. Fruits extracts, due mostly to compounds 1 and 2, showed potent antioxidant activities with an EC50 of 0.0025-0.019 mg/mL.

Development of a Total Organic Carbon method for the quantitative determination of solubility enhancement by cyclodextrins: Application to essential oils.

Formation of inclusion complexes with cyclodextrins (CDs) is known to enhance guest solubility in aqueous medium. Different techniques allow determining the evolution in solubility of individual guest compounds. However, examination of mixtures solubility encapsulated in CDs is still a challenge. This is mainly related to the difference in the response of mixture components to the applied technique or to the fact that most of the conventional methods examine the signal of an individual constituent of the mixture. Thus, applying current techniques may not reflect the behavior of the whole mixture. Here, we used for the first time Total Organic Carbon (TOC) analysis to explore and assess the efficiency of 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD) to enhance the solubility of natural complex mixtures such as essential oils (EOs). Phase solubility studies were performed for eleven EOs with HP-ß-CD. The TOC method has provided good validation parameters for linearity, precision and accuracy. For further validation of the method, phase solubility studies were performed with HP-ß-CD for eugenol, as a model EO component. The eugenol solubility was determined by UV-Visible and TOC analyses in order to compare the results. Data obtained from both methods were similar (p < 0.05), thereby proving the effectiveness of the developed TOC method. Finally, the phase solubility diagrams of EOs showed that the solubilizing potential of CD increased proportionally with the decrease in EO intrinsic solubility. Results proved that TOC could be successfully applied to investigate CD/guest inclusion complexes and is expected to have a broad range of applications in the field of mixtures encapsulation.

Determination of formation constants and structural characterization of cyclodextrin inclusion complexes with two phenolic isomers: carvacrol and thymol.

Carvacrol and thymol have been widely studied for their ability to control food spoilage and to extend shelf-life of food products due to their antimicrobial and antioxidant activities. However, they suffer from poor aqueous solubility and pronounced flavoring ability that limit their application in food systems. These drawbacks could be surpassed by encapsulation in cyclodextrins (CDs). Applications of their inclusion complexes with CDs were reported without investigating the inclusion phenomenon in deep. In this study, inclusion complexes were characterized in terms of formation constants (K f), complexation efficiency (CE), CD:guest molar ratio and increase in bulk formulation by using an UV-visible competitive method, phase solubility studies as well as (1)H and DOSY (1)H NMR titration experiments. For the first time, a new algorithmic treatment that combines the chemical shifts and diffusion coefficients variations for all guest protons was applied to calculate K f. The position of the hydroxy group in carvacrol and thymol did not affect the stoichiometry of the inclusion complexes but led to a different binding stability with CDs. 2D ROESY NMR experiments were also performed to prove the encapsulation and illustrate the stable 3D conformation of the inclusion complexes. The structural investigation was accomplished with molecular modeling studies. Finally, the radical scavenging activity of carvacrol and thymol was evaluated by the ABTS radical scavenging assay. An improvement of this activity was observed upon encapsulation. Taken together, these results evidence that the encapsulation in CDs could be valuable for applications of carvacrol and thymol in food.

Solubility, photostability and antifungal activity of phenylpropanoids encapsulated in cyclodextrins.

Effects of the encapsulation in cyclodextrins (CDs) on the solubility, photostability and antifungal activities of some phenylpropanoids (PPs) were investigated. Solubility experiments were carried out to evaluate the effect of CDs on PPs aqueous solubility. Loading capacities and encapsulation efficiencies of freeze-dried inclusion complexes were determined. Moreover, photostability assays for both inclusion complexes in solution and solid state were performed. Finally, two of the most widespread phytopathogenic fungi, Fusarium oxysporum and Botrytis cinerea, were chosen to examine the antifungal activity of free and encapsulated PPs. Results showed that encapsulation in CDs significantly increased the solubility and photostability of studied PPs (by 2 to 17-fold and 2 to 44-fold, respectively). Free PPs revealed remarkable antifungal properties with isoeugenol showing the lowest half-maximal inhibitory concentration (IC50) values of mycelium growth and spore germination inhibition. Encapsulated PPs, despite their reduced antifungal activity, could be helpful to solve drawbacks such as solubility and stability.

Promising applications of cyclodextrins in food: Improvement of essential oils retention, controlled release and antiradical activity.

Essential oils (EOs) are gaining great interest as alternatives for harmful synthetic food preservatives. Due to their volatile nature, they could be applied in food packaging to improve food quality and extend shelf-life. To provide long-term effects of EOs by increasing their retention and ensuring controlled release of their components, they could be encapsulated in cyclodextrins (CDs). Herein, the ability of six CDs to retain nine EOs and to bind their individual components was investigated. Retention capacities and binding abilities of CDs were assessed by static headspace-gas chromatography (SH-GC) using a new validated "rapid method". The ability of CDs to generate controlled release systems was examined by multiple headspace extraction (MHE). Finally, radical scavenging activity of free and encapsulated EOs was evaluated. The highest retention capacity toward the studied EOs was obtained for ß-CD and its derivatives (69-78%). Also, ß-CD and its derivatives showed, with one exception, the highest Kf values for all the studied guests. In addition, encapsulation in CDs reduced the releasing rate of EO components (from 1.43 to 2.43-fold for ß-CD/Satureja montana EO used as a model). Furthermore, the inclusion complexes showed higher ABTS(+) scavenging capacity than the free EOs. Results confirmed the usefulness of CDs as encapsulant for EOs and should encourage their application in food and as part of active packaging systems.

Liposomes incorporating cyclodextrin-drug inclusion complexes: Current state of knowledge.

Cyclodextrins (CDs) are cyclic oligosaccharides, consisting of glucopyranose units, which are able to form host-guest inclusion complexes with lipophilic molecules. The ability of CD to increase drug solubility may be used to increase drug entrapment in the aqueous compartment of liposomes and liposomes can protect CD/drug inclusion complexes until drug release. Liposomes are phospholipid vesicles composed of lipid bilayers enclosing one or more aqueous compartments. They have been widely used as safe and effective carriers for both hydrophilic and lipophilic drugs. However, lipophilic drugs incorporated in the membrane bilayers can be rapidly released, which limits the effectiveness of this drug delivery system. The coupling of both delivery systems by encapsulating CD/drug inclusion complex into liposomes is proposed to circumvent the drawbacks of each separate system. Here, we review the literature regarding the encapsulation of CD/drug inclusion complex into conventional, deformable and double loaded liposomes. The review highlights the characteristics of these systems and presents the advantages and disadvantages of each one.