Cyclodextrin Concentration Research Articles

Simultaneous enantioseparation of citrus flavonoids in chiral microemulsion electrokinetic chromatography using deep eutectic solvents

A novel, deep eutectic solvent (DES)-assisted sodium cholate (SC)-chiral microemulsion electrokinetic chromatography method is presented for separation of five flavonoid glycoside enantiomers, namely hesperidin, naringin, narirutin, eriocitrin, and neoeriocitrin. Herein, we develop a novel, green DES, whose addition considerably enhances the separation performance through multiple synergistic effects. A series of factors notably affecting chiral separation are systematically optimized: type and concentration of cyclodextrin, DES, and SC, oil phase type, and pH. The microemulsion comprises 0.5 % (v/v) n-octanol, 0.6 % (v/v) DES prepared using betaine and citric acid, 20 mM 2-HP-β-CD, 100 mM scdium cholate hydrate, and 15 mM borax buffer under the optimized conditions. Enantioseparations of the five enantiomers are achieved within 7 min, with resolutions ranging from 1.29 to 2.88. The method developed provides an innovative and effective way for chiral separation of enantiomers in complex food matrices.

Complexation of telmisartan with cyclodextrins as a tool of solubility enhancement: A comparative analysis of influence of the cyclodextrin type and method of solid dispersion preparation

Cardiovascular telmisartan (TMS) was complexed with modified cyclodextrins (CD) aiming at improvement of its poor solubility which results into low bioavailability. Inclusion complexes with 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) and sulfobutylether-β-cyclodextrin (SBE-β-CD) in liquid and solid state was evaluated for first time. Phase solubility isotherm diagrams of TMS versus the CD concentration were plotted in a buffer pH 7.4. It was found that TMS/CD complexes are formed in solution in an equimolar stoichiometric ratio of 1:1. The aqueous solubility of TMS with the addition of 0.05 mol·L−1 HP-β-CD increased by 23 times and was 38.9·10−5 mol·L−1, and the presence of the same amount of SBE-β-CD in solution increased the solubility of the drug by 7 times, reaching a value of 12.6·10−5 mol·L−1.The binding constants were found to 468 and 114 L·mol−1 for HP-β-CD and SBE-β-CD, respectively. Novel solid forms of TMS with both CDs were prepared using the grinding and evaporation approaches. Successful formation of supramolecular TMS/HP-β-CD and TMS/SBE-β-CD solid complexes was confirmed by FTIR, PXRD, DSC, TGA and SEM. The residual crystallinity of all obtained solid dispersions of TMS/CD did not exceed 10 %. Supersaturation was observed during the dissolution of all the solid complexes. The grinding technique was found to be more advantageous as it ensured the biggest improvement of the drug maximum concentration in the aqueous solution: the TMS solubility in the TMS/HP-β-CD(gr) and TMS/SBE-β-CD(gr) solid complexes increased by 27 and 33 times, respectively. Besides, polyvinyl pyrrolidone (PVP) and polyethylene glycol (PEG) were tested as crystallization inhibitors. Addition of 1 wt% PVP as a stabilizer is a promising approach for the preparation of TMS pharmaceutical formulations with improved bioavailability.

The role of cyclodextrins in the acceleration of the reaction rate in a biphasic hydroformylations

Water based biphasic hydroformylation assisted by cyclodextrins is known for enhanced reaction rates in comparison to the non-assisted biphasic hydroformylation. In this paper, the liquid–liquid interface of the hydroformylation of 1–octene, enhanced by randomly methylated-β cyclodextrins (RAME-β cyclodextrins), was examined for the first time using an in-situ image-based boroscopic technology. This technique enables the observation of the liquid–liquid interfacial area (aLL) during the reaction and under reaction conditions, facilitating an explicit analysis of the droplet population.In this study defined mixtures of the reaction components with RAME-β cyclodextrins increased the aLL by 84 %. A kinetic study showed that RAME-β cyclodextrins reduce the rate limiting effect of the nonanal concentration in the two phasic water/octene system on the interphase. The kinetic expression shows consequently higher reaction rates for this three-phase conversion, leading to a 387 % increase in aldehyde yield. When comparing four different amounts of RAME-β cyclodextrins (nCD:nRh(acac)(CO)2 = 0 / 12:1 / 24:1 / 50:1), we observed a decreasing dependency of the reaction rates on the conversion progress and therefore on the nonanal formation for higher cyclodextrin concentrations. A recovery test of the aqueous phase involving nonanal and 1-octene reveals a drastically increased concentration of 1029 % 1-octene (coctene = 1.08 10-3 mg mg−1) and 13676 % nonanal (cnonanal = 14.41 10–3 mg mg−1) after the addition of 0.002 mol RAME–β cyclodextrins. These findings affirm that RAME-β cyclodextrins primarily remove nonanal from the interfacial region while facilitating an increased availability of 1-octene for the surface bound reaction.

QbD Assisted Systematic Review for Optimizing the Selection of PVP as a Ternary Substance in Enhancing the Complexation Efficiency of Cyclodextrins: a Pilot Study.

Inclusion complexes require higher concentration of Beta cyclodextrins (βCD) resulting in increased formulation bulk, toxicity, and production costs. This systematic review offers a comprehensive analysis using Quality by design (QbD) as a tool to predict potential applications of Polyvinylpyrrolidone (PVP) as a ternary substance to address issues of inclusion complexes. We reviewed 623 documents from 2013 to 2023 and Eighteen (18) research papers were selected for statistical and meta-analysis using the QbD concept to identify the most critical factors for selecting drugs and effect of PVP on inclusion complexes. The QbD analysis revealed that Molecular weight (MW), Partition coefficient (Log P), and the auxiliary substance ratio directly affected complexation efficiency (CE), thermodynamic stability in terms of Gibbs free energy (ΔG), and percent drug release. However, Stability constant (Ks) remained unaffected by any of these parameters. The results showed that low MW (250), median Log P (6), and a βCD: PVP ratio of 2:3 would result in higher CE, lower G, and improved drug release. PVP improves drug solubility, enhances delivery and therapeutic outcomes, and counteracts increased drug ionization due to decreased pH. In certain cases, its bulky nature and hydrogen bonding with CD molecules can form non-inclusion complexes. The findings of the study shows that there is potential molecular interaction between PVP and β-cyclodextrins, which possibly enhances the stability of inclusion complexes for drug with low MW and log P values less than 9. The systematic review shows a comprehensive methodology based on QbD offers a replicable template for future investigations into drug formulation research.

Evaluation of the performance of Hydroxypropyl-β-cyclodextrin for the consolidation of vegetable-tanned leather artifacts

The fragility and weakness of historical, archaeological, and heritage leather artifacts in museums, and libraries due to unsuitable environmental conditions are significant challenges. This study aims to assess the effectiveness of Hydroxypropyl Beta Cyclodextrin, a novel material, in consolidating leather artifacts by examining their physical, chemical, and mechanical properties. Vegetable-tanned leather samples were treated with different concentrations of Hydroxypropyl Beta Cyclodextrin and subjected to artificial heat aging. Evaluation methods included digital and scanning electron microscopy, contact angle measurements, mechanical testing, color analysis, pH measurement, and Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy. Results showed that Hydroxypropyl Beta Cyclodextrin at 1% and 2% concentrations improved chemical stability, surface morphology, color retention, and mechanical properties of the leather samples. The third concentration yielded less favorable outcomes. This study recommends using Hydroxypropyl Beta Cyclodextrin at 1% and 2% concentrations for consolidating historical leathers.

Characterization of behavior of the inclusion complex between Sulfobutylether-β-cyclodextrin loaded alginate/chitosan nanoparticles and indomethacin as a potential approach for drug delivery

Indomethacin (IND) is a nonsteroidal anti-inflammatory drug with low aqueous solubility and irritating gastrointestinal side effects. By complexing this drug inside the cavity of cyclodextrin (CD) and loading it into polymer nanoparticles such as Chitosan (CS) and Alginate (Alg), in addition to increasing its solubility, a targeted drug delivery platform can be created to release this drug. In this study, initially, the complex of IND with Sulfobutylether-β-cyclodextrin (SBE-β-CD) was created using the freeze-drying method. Then, by ionic gelation method, the complex was loaded into CS nanoparticles, and finally, these nanoparticles were coated with Alg to enable drug release in the intestinal environment. Complexes and nanoparticles formation were investigated by phase solubility, FTIR, HNMR, XRD, and SEM methods. Moreover, the Zeta potential method was used to measure the stability of nanoparticles coated with Alg. According to phase solubility studies, drug solubility increased linearly with increasing CD concentration, indicating an AL-type system. In addition, according to the phase solubility diagram, the stability constant and complexation efficiency were obtained 177 M−1 and 0.388, respectively. In the following, the formation of chitosan nanoparticles was confirmed through spectral shifts observed in FTIR analysis as well as morphological changes revealed in SEM analysis. Also, the zeta potential of Alg coated nanoparticles was 110.067 mV, which indicates the good stability of these nanoparticles. At the end, in vitro release studies were conducted to investigate the pH-sensitive capability of these nanoparticles at pH = 1.5 and pH = 7. The formed nanosystem successfully released 78.51 % of the drug at pH = 7 and prevented its release in acidic pH.

Effect of cholesterol-loaded cyclodextrin treatment on boar sperm cryopreservation.

This study investigated the efficacy of different concentrations of cholesterolloaded cyclodextrin (CLC) on cryopreservation in boar sperm quality. In this study, we treated boar sperm with different concentrations of CLC before freezing and analyzed the sperm cholesterol concentration, plasma membrane, acrosome integrity rate and total motility rate before and after freeze-thawing. We also investigated the levels of reactive oxygen species (ROS), malondialdehyde (MDA), adenosine triphosphate (ATP), and structural- and oxidative-damage related proteins in all groups after thawing. The results revealed that the cholesterol concentration of the CLC-treated groups was higher than that of the control group, both before freezing and after thawing (p<0.05). The plasma membrane integrity rate, acrosome integrity rate, and total motility rate of sperm were also enhanced after thawing in the CLC-treated group (all p<0.05). Moreover, ROS and MDA production and ATP loss were reduced in CLC-treated sperm during freezing and thawing (p<0.05). Finally, CLC pretreatment partially prevented the consumption of various proteins involved in metabolism including capping actin protein of muscle Z-line subunit beta (CAPZB), heat shock protein 90 alpha family class A member 1 (HSP90AA1) and phosphoglycerate mutase 2 (PGAM2) (p<0.05). The CLC treatment increased cholesterol concentration and decreased structural injury and oxidative damage during boar sperm freezing and thawing, improving the efficacy of sperm cryopreservation in boar.

Addition of low concentration of cholesterol-loaded cyclodextrin (CLC) has a positive effect on cryopreserved canine spermatozoa evaluated by andrological and biophysical methods

BackgroundThis study was conducted to find the best concentration of cholesterol-loaded cyclodextrin (CLC) which has a positive impact on canine post thaw semen quality. Three different concentrations of CLC (0.83 mg/ml; 1.66 mg/ml; 3.32 mg/ml) and 2-hydroxylpropyl-beta-cyclodextrin (HBCD) (1.66 mg/ml) were used in addition to cryopreservation extender and compared with the control after thawing. Samples were assessed using computer-assisted semen analyzer (CASA), flow cytometry, fluorimeter by measuring the fluorescence anisotropy (ANISO) and determining the generalized membrane polarization (GP).ResultsAn addition of 0.83 mg/ml CLC significantly increased the percentage of progressive motile (PROG) and rapid spermatozoa (RAP) (P < 0.05). 1.66 mg/ml HBCD decreased progressive motility of spermatozoa and population with rapid movement relative to the control (P < 0.05). Furthermore, the groups with an addition of 1.66 mg/ml and 3.32 mg/ml of CLC, as well as the group with only cyclodextrin, increased percentage of dead spermatozoa without lipid peroxidation and decreased percentage of viable spermatozoa without LPO which was lower in these groups than in the control (P < 0.05). Other sperm parameters assessed on flow cytometer were not significantly different. The addition of CLC at 0.83 mg/ml and 3.32 mg/ml concentrations and 1.66 mg/ml of HBCD caused an increase in ANISO measured at 23 ºC (P < 0.05).ConclusionsIn conclusion, the results suggest that increasing cholesterol in the plasma membrane of canine spermatozoa can improve their freezability. However, only low concentrations of CLC may improve semen quality after thawing without adversely affecting other parameters.

Granular polyrotaxane microgels as injectable hydrogels for corneal tissue regeneration.

Corneal diseases, a leading cause of global vision impairment, present challenges in treatment due to corneal tissue donor scarcity and transplant rejection. Hydrogel biomaterials in the form of corneal implants for tissue regeneration, while promising, have faced obstacles related to cellular and tissue integration. This study develops and investigates the potential of granular polyrotaxane (GPR) hydrogels as a scaffold for corneal keratocyte growth and transparent tissue generation. Employing host-guest driven supramolecular interactions, we developed injectable, cytocompatible hydrogels. By optimizing cyclodextrin (CD) concentrations in thiol-ene crosslinked PEG microgels, we observed improved mechanical properties and thermoresponsiveness while preserving injectability. These microgels, adaptable for precise defect filling, 3D printing or tissue culture facilitate enhanced cellular integration with corneal keratocytes and exhibit tissue-like structures in culture. Our findings demonstrate the promise of GPR hydrogels as a minimally invasive avenue for corneal tissue regeneration. These results have the potential to address transplantation challenges, enhance clinical outcomes, and restore vision.

Advantages of Induced Circular Dichroism Spectroscopy for Qualitative and Quantitative Analysis of Solution-Phase Cyclodextrin Host-Guest Complexes.

The presence of a chiral or chirally perturbed chromophore in the molecule under investigation is a fundamental requirement for the appearance of a circular dichroism (CD) spectrum. For native and for most of the substituted cyclodextrins, this condition is not applicable, because although chiral, cyclodextrins lack a chromophore group and therefore have no characteristic CD spectra over 220 nm. The reason this method can be used is that if the guest molecule has a chromophore group and this is in the right proximity to the cyclodextrin, it becomes chirally perturbed. As a result, the complex will now provide a CD signal, and this phenomenon is called induced circular dichroism (ICD). The appearance of the ICD spectrum is clear evidence of the formation of the complex, and the spectral sign and intensity is a good predictor of the structure of the complex. By varying the concentration of cyclodextrin, the ICD signal changes, resulting in a saturation curve, and from these data, the stability constant can be calculated for a 1:1 complex. This article compares ICD and NMR spectroscopic and molecular modeling results of cyclodextrin complexes of four model compounds: nimesulide, fenbufen, fenoprofen, and bifonazole. The results obtained by the different methods show good agreement, and the structures estimated from the ICD spectra are supported by NMR data and molecular modeling.

Chiral Separation of Vildagliptin by Capillary Electrophoresis—The Study of Enantiomeric Complexation

Vildagliptin (VIL) is a dipeptidyl peptidase-4 inhibitor used in the treatment of type 2 diabetes mellitus; in therapy, it is available as the enantiomerically pure S-VIL, the other enantiomer R-VIL being considered as an enantiomeric impurity. A systematic screening of 16 cyclodextrin (CD) derivatives as chiral selectors was performed at three pH levels using phosphate (pH 2.5, pH 7.0) and acetate (pH 4.5) buffers. Method optimization employed an experimental design approach, systematically investigating the effect of buffer and CD concentration, buffer pH, capillary temperature, and applied voltage on the chiral resolution and analysis time. The method’s analytical performance was thoroughly assessed and subsequently employed for determining the enantiomeric purity of VIL in a pharmaceutical formulation. The properties of the inclusion complexes, such as stoichiometry and atomic level intermolecular host–guest interactions were studied by NMR measurements and molecular modeling. Native α-CD at acidic pH has demonstrated its exceptional suitability for the separation of VIL enantiomers with a favorable migration order (R-VIL followed by S-VIL). The optimized analytical conditions (75 mM acetate buffer, pH 4.5, containing 50 mM α-CD, 18 kV applied voltage, and 15 °C capillary temperature) provided a baseline separation of VIL enantiomers within 9 min. The developed method represents a cost-effective alternative to the enantiomeric impurity control of VIL. Symmetry is often a fundamental aspect of molecular structures and interactions, and our detailed analysis of the chiral recognition process contributes to the understanding of symmetry-related aspects in molecular systems. This developed method not only offers a cost-effective alternative for the enantiomeric impurity control of VIL but also provides valuable information regarding the mechanism of the chiral recognition process, aligning with the broader themes of symmetry in molecular sciences.

Cyclodextrins permeabilize DPPC liposome membranes: a focus on cholesterol content, cyclodextrin type, and concentration.

Cyclodextrins (CDs) are known for their ability to extract lipid components from synthetic and biological membranes and therefore to induce an increase of membrane permeability. However, the effect of cholesterol (CHOL) content in the membrane on the CD permeabilizing effect was not considered yet. Given that an increase in CHOL content reduces the membrane permeability, the aim of this work was to reveal how CHOL would modulate the CDs effect on the membrane. Hence, liposomes made of dipalmitoyl phosphatidylcholine (DPPC) and various CHOL contents (DPPC/CHOL 100:10, 100:25, 100:50, and 100:100) encapsulating the hydrophilic fluorophore, sulforhodamine B (SRB), were prepared and exposed to the native CDs (α-CD, β-CD, γ-CD) and four β-CD derivatives: the randomly methylated-β-CD (RAMEB), the low methylated-β-CD (CRYSMEB), the hydroxypropyl-β-CD (HP-β-CD) and the sulfobutyl ether-β-CD (SBE-β-CD) at different CD/DPPC molar ratios (1:1, 10:1, and 100:1). The membrane permeability was monitored following the release of SRB with time. The results demonstrated that the CDs effect on the membrane depends on the CD type, CD concentration, and membrane CHOL content. The investigated CDs exhibited an instantaneous permeabilizing effect promoting vesicle leakage of SRB from the various membranes; this effect increased with CDs concentration. Among the studied CDs, α-CD, β-CD, and RAMEB were the most permeabilizing CDs on the different membranes. Similar modifications of SRB release from the various liposomal formulations were obtained with HP-β-CD, CRYSMEB, and SBE-β-CD. γ-CD was the less potent CD in affecting the membrane permeability. The CDs effect also depended on the CHOL content: at the CD/DPPC molar ratio (100:1), RAMEB and β-CD considerably permeabilized the membrane of high CHOL content (50%, 100%) while the remaining CDs showed a decreasing permeabilizing effect upon CHOL content membrane increase.

Ejected microcrystals probe jammed states of droplets in cyclodextrin-based emulsions

The cyclodextrin (CD)-based emulsions exhibit complex instability behaviors such as rapid flocculation and creaming, and how to capture droplet dispersion states of the emulsions remains a great challenge. Here we prepare the CD-based emulsions with different oil-water volume ratios and CD concentrations by using high-pressure homogenization, and characterize the emulsion droplets by using optical microscopy and confocal laser scanning microscopy. We evaluate the effects of homogenization pressure on the stability of the emulsions, identify armored droplets with different surface features, measure interfacial concentrations of adsorbed ICs microcrystals, and observe ejection of the oil/CD inclusion complexes (ICs) microcrystals from the droplet surface. The droplet dispersion states are sensitive to the dynamic buildup and evolving morphologies of the interfacial microcrystals, and there are clear correlations between the properties of the ejected microcrystals and the characteristics of the emulsions. We ascribe the subsequent ejection of ICs microcrystals from the droplet surface to consolidation and deformation of the films formed between neighboring droplets. The ejection of the ICs microcrystals affords a simple method to detect the droplet-droplet interactions and phase transitions in the CD-based emulsions, which might be a generic feature in the broader context of the creaming processes of emulsions.

Cyclodextrin's Effect on Permeability and Partition of Nortriptyline Hydrochloride.

Cyclodextrin-based delivery systems have been intensively used to improve the bioavailability of drugs through the modification of their pharmaceutically relevant properties, such as solubility, distribution and membrane permeation. The present work aimed to disclose the influence of HP-β-CD and SBE-β-CD on the distribution and permeability of nortriptyline hydrochloride (NTT•HCl), a tricyclic antidepressant drug. To this end, the distribution coefficients in the 1-octanol/buffer and n-hexane/buffer model systems and the coefficients of permeability through the cellulose membrane and lipophilic PermeaPad barrier were determined at several cyclodextrin concentrations. The results demonstrated a dramatic decrease in both the distribution and the permeability coefficients as the cyclodextrin concentration rose, with the decrease being more pronounced in SBE-β-CD due to the charge-charge attraction and electrostatic interactions between NTT and SBE-β-CD. It is these interactions that were shown to be responsible for the greater value of the constant of NTT's association with SBE-β-CD than that with HP-β-CD. The findings of this study revealed similar trends in the 1-octanol/buffer 6.8 pH distribution and permeability through the PermeaPad barrier in the presence of CDs. These results were attributed to the determinative role of the distribution coefficient (serving as a descriptor) in permeation through the PermeaPad barrier modeling the lipophilic nature of biological barriers.

Screening of Cyclodextrins in the Processing of Buserelin Dry Powders for Inhalation Prepared by Spray Freeze-Drying.

In this study, we prepared inhalable buserelin microparticles using the spray freeze-drying (SFD) method for pulmonary drug delivery. Raffinose as a cryoprotectant carrier was combined with two levels of five different cyclodextrins (CDs) and then processed by SFD. Dry powder diameters were evaluated by laser light scattering and morphology was determined by scanning electron microscopy (SEM). Differential scanning calorimetry (DSC) and X-ray diffraction (XRD) analysis were utilized for the determination of crystalline structures. The aerodynamic properties of the spray freeze-dried powders were evaluated by twin stage impinger (TSI) and the stability of prepared samples was assessed under normal and accelerated conditions. The prepared powders were mostly porous spheres and the size of microparticles ranged from 9.08 to 13.53 μm, which are suitable as spray-freeze dried particles. All formulations showed amorphous structure confirmed by DSC and XRD. The aerosolization performance of the formulation containing buserelin, raffinose and 5% beta-cyclodextrin (β-CD), was the highest and its fine particle fraction (FPF) was 69.38%. The more circular and separated structures were observed in higher concentrations of CDs, which were compatible with FPFs. The highest stability was obtained in the formulation containing hydroxypropyl beta-cyclodextrin (HP-β-16. CD) 5%. On the contrary, sulfobutylether beta-cyclodextrin (SBE-β-CD) 5% bearing particles showed the least stability. By adjusting the type and ratio of CDs in the presence of raffinose, the prepared formulations could effectively enhance the aerosolization and stability of buserelin. Therefore, they can be proposed as a suitable career for lung drug delivery.

Effect of FertTalp addition to stallion semen on kinetic parameters and production of intracellular hydrogen peroxide after thawing

Motility stimulants increase sperm energy production and their addition after semen thawing will increase motility (Stephens et al., J. Equine Vet. Sci. 2013;33:615-621), and stimulate metabolic pathways responsible for sperm capacitation (Guasti et al., Anim. Reprod. Sci. 2017;179:27-34). Semen of 12 Mangalarga Marchador stallions was cryopreserved with addition of different concentrations of cholesterol-loaded cyclodextrin (CLC): G1 (control – without cholesterol), G2 (1mg of cholesterol), G3 (1.5mg of cholesterol) and G4 (2mg of cholesterol). To run the experiment, two straws from each stallion per experimental group were thawed and divided into two aliquots of 400µL. This was supplemented with 100µL of Fert Talp, a Tyrode's albumin lactate pyruvate medium with (FT) or without (No-FT) heparin, a motility stimulant. This subdivided the four groups into eight. Sperm kinetics were performed analyzed by CASA (Sperm Class Analyzer, SCA® v.4.0 Microptic®). Sperm longevity was analyzed by using a thermoresistance test at 37°C (TTR). Statistical analysis was done by ANOVA F-test. Complementary to sperm kinetics, the production of intracellular hydrogen peroxide was estimated by flow cytometry using Dichlorodihydrofluorescein diacetate (DCFDA®). Results were classified as: Q1 [Plasma membrane (PM) injured without intracellular hydrogen peroxide]; Q2 (PM injured with intracellularhydrogen peroxide); Q3 (intact MP with intracellular hydrogen peroxide); Q4 (intact PM without intracellular hydrogen peroxide) and Tukey test was performed for statistical analysis. Immediately after thawing, there was no difference between FT and No-FT (p>0.05). After incubation at 37°C, the addition of FT showed higher values for RAP (percentage of rapid sperm, %), VAP (average path velocity, μm/s) and LIN (linearity of the curvilinear trajectory, %) after 80 minutes of incubation and also superior rates of RAP, VSL (straight line velocity, μm/s), VAP and LIN after 120 minutes of incubation (p<0.05), demonstrating satisfactory results both with regard to sperm kinetics and longevity. It can be concluded that the use of FT was beneficial for sperm velocity. These findings suggest that the addition of this motility stimulant after thawing may improve fertility because sperm velocity – VCL (curvilinear velocity, μm/s), VSL and VAP are the most reliable parameters to assess sperm quality (Verstegen et al. Theriogenology, 2002;57:149-179). Significant differences were also observed for Q4 (intact PM, without hydrogen peroxide), where the G2 FT group demonstrated superiority when compared to all groups not supplemented with FT (p>0.05), providing satisfactory plasmatic membrane results against oxidative stress.

Post-thaw addition of pentoxifylline to equine semen previously cryopreserved with different concentrations of cholesterol-loaded cyclodextrin

Cholesterol-loaded cyclodextrin (CLC) grants resistance to sperm during cryopreservation but may compromise sperm capacitation and fertility (Zahn et al. Theriogenology, 2002;58(2):237:240). Stimulants of sperm kinetics like pentoxifyllin (PTX) may increase sperm energy production and stimulate metabolic pathways responsible for sperm capacitation (Guasti et al. Anim.Reprod.Sci. 2017;179:27-34). The present study aimed to stimulate semen motility by adding PTX after thawing of semen previously cryopreserved with different concentrations of CLC. Mangalarga Marchador stallions (n=12; 4-14 years) were assigned to four groups: (G1) untreated control, (G2) inclusion of cholesterol-loaded cyclodextrin (CLC) at concentrations of 1mg, (G3) 1.5mg and (G4) 2mg. Two straws of each stallion per group were thawed and divided into two aliquots, pentoxifylline (2 mg/mL, 7.18 mM) homogenized in BotuCrio®, Botupharma, Brazil (PTX: G1, G2, G3 e G4) was added to one and not to other aliquot (No-PTX: G1, G2, G3 and G4). Sperm kinetics was analyzed by CASA (Sperm Class Analyzer, SCA® v.4.0 Microptic®) immediately after thawing (T0) or at 40 minutes (T40), 80 minutes (T80) and 120 minutes (T120) after thawing after incubation at 37°C. For statistical analysis, ANOVA F-test was performed.Intracellular hydrogen peroxide production was estimated by flow cytometry with dichlorodihydrofluorescein diacetate. Fertility was tested by deep horn insemination 0 to 6h after ovulation: No-PTX G1, No-PTX G3 and PTX G3. PTX reduced sperm velocity parameters at all times (p<0.05) compared to the groups without addition of pentoxifylline (No-PTX G1, G2, G3 and G4). Egg yolk based semen extenders may interfere with PTX cell penetration as they reduce permeability of the cell membrane. Regarding the integrity of the sperm plasma membrane and production of hydrogen peroxide, the use of pentoxifylline conferred an increase in sperm possibly under oxidative stress in the control group, with injured plasma membrane and with intracellular hydrogen peroxide (G1 PTX). When PTX was added to the group with highest concentration of cholesterol (G4 PTX), the intactness of the plasma membrane was improved, but with intracellular hydrogen peroxide. Mitochondrial potential and plasma membrane destabilization were also evaluated (n.s.). Pregnancy rates were 50% (2/4) for No-PTX G1, 67% (2/3) for No-PTX G3 and 67% (2/3) for PTX G3. In conclusion, addition of PTX did not interfere with mitochondrial potential, membrane destabilization and conception rate but was detrimental to cell longevity and to sperm kinetics.

Chiral separation by capillary electrokinetic chromatography with hydrophobic deep eutectic solvents as pseudo-stationary phases

This study demonstrated for the first time that hydrophobic deep eutectic solvents (HDESs) can be used in capillary electrophoresis (CE) for chiral separations. We found that the an HDES methyltrioctylammonium chloride:octanoic acid (N8881Cl:OctA) can exist in the form of nano-sized microdroplets in CE background electrolyte solutions, and show hydrophobic effects as a new type of pseudo-stationary phase (PSP) during CE separation. When used in combination with various cyclodextrin (CD)-type chiral selectors, the presence of N8881Cl:OctA significantly improved the enantioresolutions of several model drugs. Moreover, the migration time of the enantiomers can also be reduced when an anionic CD (e.g., carboxymethyl-β-cyclodextrin (CM-β-CD)) was used. Critical factors influencing the chiral separations were systematically investigated including the HDES concentration, hydrogen-bond acceptor (HBA)/hydrogen-bond donor (HBD) ratio, CD concentration, buffer pH, and applied voltage, etc. An insight into chiral recognition mechanism with HDES is provided for reference. A comparison of the chiral CE performance of HDESs with traditional surfactants was also performed to demonstrate their superiority as a new type of PSP.

Chiral Separation of Apremilast by Capillary Electrophoresis Using Succinyl-β-Cyclodextrin-Reversal of Enantiomer Elution Order by Cationic Capillary Coating.

A stereospecific capillary electrophoresis method was developed for the separation of the novel, antipsoriatic agent, apremilast (APR). Six anionic cyclodextrin (CD) derivatives were screened for their ability to discriminate between the uncharged enantiomers. Only succinyl-β-CD (Succ-β-CD) presented chiral interactions; however, the enantiomer migration order (EMO) was unfavorable, and the eutomer, S-APR, migrated faster. Despite the optimization of all possible parameters (pH, cyclodextrin concentration, temperature, and degree of substitution of CD), the method was unsuccessful for purity control due to the low resolution and the unfavorable enantiomer migration order. Changing the direction of electroosmotic flow (EOF) by the dynamic coating of the inner surface of the capillary with poly(diallyldimethylammonium) chloride or polybrene resulted in EMO reversal, and the developed method could be applied for the determination of R-APR as the enantiomeric purity. Thus, the application of the dynamic capillary coating offers a general opportunity for enantiomeric migration order reversal in particular cases when the chiral selector is a weak acid.

Complexation between the Antioxidant Pterostilbene and Derivatized Cyclodextrins in the Solid State and in Aqueous Solution.

Inadequate aqueous solubilities of bioactive compounds hinder their ability to be developed for medicinal applications. The potent antioxidant pterostilbene (PTB) is a case in point. The aim of this study was to use a series of modified water-soluble cyclodextrins (CDs), namely, hydroxypropyl β-CD (HPβCD), dimethylated β-CD (DIMEB), randomly methylated β-CD (RAMEB), and sulfobutyl ether β-CD sodium salt (SBECD) to prepare inclusion complexes of PTB via various solid, semi-solid, and solution-based treatments. Putative CD-PTB products generated by solid-state co-grinding, kneading, irradiation with microwaves, and the evaporative treatment of CD-PTB solutions were considered to have potential for future applications. Primary analytical methods for examining CD-PTB products included differential scanning calorimetry and Fourier transform infrared spectroscopy to detect the occurrence of binary complex formation. Phase solubility analysis was used to probe CD-PTB complexation in an aqueous solution. Complexation was evident in both the solid-state and in solution. Complex association constants (K1:1) in an aqueous solution spanned the approximate range of 15,000 to 55,000 M-1; the values increased with the CDs in the order HPβCD < DIMEB < RAMEB < SBECD. Significant PTB solubility enhancement factors were recorded at 100 mM CD concentrations, the most accurately determined values being in the range 700-fold to 1250-fold.