Non-Lamellar Liquid Crystalline Nanocarriers for Thymoquinone Encapsulation

Owing to their unique structural features, non-lamellar liquid crystalline nanoparticles comprising cubosomes and hexosomes are attracting increasing attention as versatile investigative drug carriers. Background: Depending on their physiochemical characteristics, drug molecules on entrapment can modulate and reorganize structural features of cubosomes and hexosomes. Therefore, it is important to assess the effect of guest molecules on broader biophysical characteristics of non-lamellar liquid crystalline nanoparticles, since drug-induced architectural, morphological, and size modifications can affect the biological performance of cubosomes and hexosomes. Methods: We report on alterations in morphological, structural, and size characteristics of nanodispersions composed from binary mixtures of glycerol monooleate and vitamin E on thymoquinone (a molecule with wide therapeutic potentials) loading. Results: Thymoquinone loading was associated with a slight increase in the mean hydrodynamic nanoparticle size and led to structural transitions from an internal biphasic feature of coexisting inverse cubic Fd3m and hexagonal (H2) phases to an internal inverse cubic Fd3m phase (micellar cubosomes) or an internal inverse micellar (L2) phase (emulsified microemulsions, EMEs). We further report on the presence of “flower-like” vesicular populations in both native and drug-loaded nanodispersions. Conclusions: These nanodispersions have the potential to accommodate thymoquinone and may be considered as promising platforms for the development of thymoquinone nanomedicines

Internal lamellar and inverse hexagonal liquid crystalline phases during the digestion of krill and astaxanthin oil-in-water emulsions

Krill oil represents an important alternative natural source of omega-3 (ω-3) polyunsaturated fatty acids (PUFAs). Considering the beneficial health effects of these essential fatty acids, particularly in various disorders including cancer, cardiovascular, and inflammation diseases, it is of paramount importance to gain insight into the digestibility of krill oil. In this work, we study the fate of krill oil-in-water emulsion, stabilized by sodium caseinate, during lipolysis by coupling time-resolved synchrotron small-angle X-ray scattering (SAXS) to flow-through lipolysis model. For gaining further insight into the effect of ω-3 PUFA-containing oil type on the dynamic structural features occurring during lipolysis, two additional astaxanthin oil-in-water emulsions, stabilized using either sodium caseinate or citrem, were subjected to lipolysis under identical experimental conditions. In addition to the difference in lipid composition in both oils, ω-3 PUFAs in astaxanthin oil, similar to fish oil, exist in the form of triacylglycerols; whereas most of those in krill oil are bound to phospholipids. SAXS showed the formation of highly ordered nanostructures on exposure of these food emulsions to the lipolysis medium: the detection of a biphasic feature of coexisting inverse hexagonal (H2) and lamellar (Lα) liquid crystalline phases in the digested krill oil droplets' interiors, as compared to a neat Lα phase in the digested astaxanthin oil droplets. We discuss the dynamic phase behavior and describe the suggested important role of these phases in facilitating the delivery of nutrients throughout the body. In addition, the potential implication in the development of food and drug nanocarriers is briefly described

pH-responsive nano-self-assemblies of the anticancer drug 2-hydroxyoleic acid

pH-responsive lipid nanocarriers have the potential to selectively target the acidic extracellular pH environment of cancer tissues and may further improve the efficacy of chemotherapeutics by minimizing their toxic side-effects. Here, we present the design and characterization of pH-sensitive nano-self-assemblies of the poorly water-soluble anticancer drug 2-hydroxyoleic acid (2OHOA) with glycerol monooleate (GMO). pH- triggered nanostructural transformations from 2OHOA/GMO nanoparticles with an internal inverse hexagonal structure (hexosomes) at pH around 2.0–3.0, via nanocarriers with an internal inverse bicontinuous cubic structure (cubosomes) at pH 2.0–4.5, to vesicles at pH 4.5–7.4 were observed with synchrotron small-angle X-ray scattering, and cryogenic transmission electron microscopy. ζ-potential measurements highlight that the pH-driven deprotonation of the carboxylic group of 2OHOA, and the resulting charge-repulsions at the lipid–water interface account for these nanostructural alterations. The study provides detailed insight into the pH-dependent self-assembly of 2OHOA with GMO in excess buffer at physiologically relevant pH values, and discusses the effects of pH alterations on modulating their nanostructure. The results may guide the further development of pH-responsive anticancer nanocarriers for the targeted delivery of chemotherapeutics to the local microenvironment of tumor cells

Direct monitoring of calcium-triggered phase transitions in cubosomes using small-angle X-ray scattering combined with microfluidics

This article introduces a simple microfluidic device that can be combined with synchrotron small-angle X-ray scattering (SAXS) for monitoring dynamic structural transitions. The microfluidic device is a thiol-ene-based system equipped with 125 μm-thick polystyrene windows, which are suitable for X-ray experiments. The device was prepared by soft lithography using elastomeric molds followed by a simple UV-initiated curing step to polymerize the chip material and simultaneously seal the device with the polystyrene windows. The microfluidic device was successfully used to explore the dynamics of the structural transitions of phytantriol/dioleoylphosphatidylglycerol-based cubosomes on exposure to a buffer containing calcium ions. The resulting SAXS data were resolved in the time frame between 0.5 and 5.5 s, and a calcium-triggered structural transition from an internal inverted-type cubic phase of symmetry Im3m to an internal inverted-type cubic phase of symmetry Pn3m was detected. The combination of microfluidics with X-ray techniques opens the door to the investigation of early dynamic structural transitions, which is not possible with conventional techniques such as glass flow cells. The combination of microfluidics with X-ray techniques can be used for investigating protein unfolding, for monitoring the formation of nanoparticles in real time, and for other biomedical and pharmaceutical investigations. A combination of microfluidics with X-ray techniques has been used to perform dynamic structural studies on nanoparticulate formulations

Self-Assembly in Monoelaidin Aqueous Dispersions: Direct Vesicles to Cubosomes Transition

Background: In the present study, synchrotron small-angle X-ray scattering (SAXS) and Cryo-TEM were used to characterize the temperature-induced structural transitions of monoelaidin (ME) aqueous dispersion in the presence of the polymeric stabilizer F127. We prove that the direct transition from vesicles to cubosomes by heating this dispersion is possible. The obtained results were compared with the fully hydrated bulk ME phase. Methodology/principal findings: Our results indicate the formation of ME dispersion, which is less stable than that based on the congener monoolein (MO). In addition, the temperature-dependence behavior significantly differs from the fully hydrated bulk phase. SAXS findings indicate a direct L(alpha)-V(2) internal transition in the dispersion. While the transition temperature is conserved in the dispersion, the formed cubosomes with internal Im3m symmetry clearly contain more water and this ordered interior is retained over a wider temperature range as compared to its fully hydrated bulk system. At 25 degrees C, Cryo-TEM observations reveal the formation of most likely closely packed onion-like vesicles. Above the lamellar to non-lamellar phase transition at 65 degrees C, flattened cubosomes with an internal nanostructure are observed. However, they have only arbitrary shapes and thus, their morphology is significantly different from that of the well-shaped analogous MO cubosome and hexosome particles. Conclusions/significance: Our study reveals a direct liposomes-cubosomes transition in ME dispersion. The obtained results suggest that the polymeric stabilizer F127 especially plays a significant role in the membrane fusion processes. F127 incorporates in considerable amount into the internal nanostructure and leads to the formation of a highly swollen Im3m phase

Calcium Triggered Lα-H2 Phase Transition Monitored by Combined Rapid Mixing and Time-Resolved Synchrotron SAXS

BACKGROUND: Awad et al. reported on the Ca(2+)-induced transitions of dioleoyl-phosphatidylglycerol (DOPG)/monoolein (MO) vesicles to bicontinuous cubic phases at equilibrium conditions. In the present study, the combination of rapid mixing and time-resolved synchrotron small-angle X-ray scattering (SAXS) was applied for the in-situ investigations of fast structural transitions of diluted DOPG/MO vesicles into well-ordered nanostructures by the addition of low concentrated Ca(2+) solutions. METHODOLOGY/PRINCIPAL FINDINGS: Under static conditions and the in absence of the divalent cations, the DOPG/MO system forms large vesicles composed of weakly correlated bilayers with a d-spacing of approximately 140 A (L(alpha)-phase). The utilization of a stopped-flow apparatus allowed mixing these DOPG/MO vesicles with a solution of Ca(2+) ions within 10 milliseconds (ms). In such a way the dynamics of negatively charged PG to divalent cation interactions, and the kinetics of the induced structural transitions were studied. Ca(2+) ions have a very strong impact on the lipidic nanostructures. Intriguingly, already at low salt concentrations (DOPG/Ca(2+)>2), Ca(2+) ions trigger the transformation from bilayers to monolayer nanotubes (inverted hexagonal phase, H(2)). Our results reveal that a binding ratio of 1 Ca(2+) per 8 DOPG is sufficient for the formation of the H(2) phase. At 50 degrees C a direct transition from the vesicles to the H(2) phase was observed, whereas at ambient temperature (20 degrees C) a short lived intermediate phase (possibly the cubic Pn3m phase) coexisting with the H(2) phase was detected. CONCLUSIONS/SIGNIFICANCE: The strong binding of the divalent cations to the negatively charged DOPG molecules enhances the negative spontaneous curvature of the monolayers and causes a rapid collapsing of the vesicles. The rapid loss of the bilayer stability and the reorganization of the lipid molecules within ms support the argument that the transition mechanism is based on a leaky fusion of the vesicles

Tuning Curvature and Stability of Monoolein Bilayers by Designer Lipid-Like Peptide Surfactants

This study reports the effect of loading four different charged designer lipid-like short anionic and cationic peptide surfactants on the fully hydrated monoolein (MO)-based Pn3m phase (Q224). The studied peptide surfactants comprise seven amino acid residues, namely A6D, DA6, A6K, and KA6. D (aspartic acid) bears two negative charges, K (lysine) bears one positive charge, and A (alanine) constitutes the hydrophobic tail. To elucidate the impact of these peptide surfactants, the ternary MO/peptide/water system has been investigated using small-angle X-ray scattering (SAXS), within a certain range of peptide concentrations (R≤0.2) and temperatures (25 to 70°C). We demonstrate that the bilayer curvature and the stability are modulated by: i) the peptide/lipid molar ratio, ii) the peptide molecular structure (the degree of hydrophobicity, the type of the hydrophilic amino acid, and the headgroup location), and iii) the temperature. The anionic peptide surfactants, A6D and DA6, exhibit the strongest surface activity. At low peptide concentrations (R = 0.01), the Pn3m structure is still preserved, but its lattice increases due to the strong electrostatic repulsion between the negatively charged peptide molecules, which are incorporated into the interface. This means that the anionic peptides have the effect of enlarging the water channels and thus they serve to enhance the accommodation of positively charged water-soluble active molecules in the Pn3m phase. At higher peptide concentration (R = 0.10), the lipid bilayers are destabilized and the structural transition from the Pn3m to the inverted hexagonal phase (H2) is induced. For the cationic peptides, our study illustrates how even minor modifications, such as changing the location of the headgroup (A6K vs. KA6), affects significantly the peptide's effectiveness. Only KA6 displays a propensity to promote the formation of H2, which suggests that KA6 molecules have a higher degree of incorporation in the interface than those of A6K