Efficient Development of Green Emulsifier/Emollient-Based Emulsion Vehicles: From RSM Optimal Experimental Design to Abridged In Vivo Assessment.

Since natural-origin, sustainable ingredients are preferred by modern consumers, novel emulsifiers and emollients keep entering the market. This study hypothesizes that a combination of in silico, instrumental tools and simplified sensory studies could be used to efficiently characterize emulsions in a shorter timeframe. A total of 22 rather simple o/w emulsions were prepared by a time/energy-saving emulsification process. A natural mixed emulsifier (Lauryl Glucoside/Myristyl Glucoside/Polyglyceryl-6 Laurate) and two emollients (both with INCI name C15-19 Alkane) were used. The performed D-optimal experimental design within the response surface method (RSM) significantly narrowed down the number of samples about to enter the stage of texture, friction and sensory studies to the samples comprising 30% of a respective Emogreen emollient and 2% or 3% of the emulsifier. The sample comprising 2% emulsifier/30% Emogreen® L15 showed significantly higher firmness (42.12 mN) when compared to the one with 2% emulsifier/30% Emogreen® L19 (33.62 mN), which was somewhat unexpected considering the emollients' inherent viscosity values (4.5 mPa·s for L15 and 9 mPa·s for L19). The sample with 2% emulsifier/30% Emogreen® L19 managed to maintain the lowest friction, while the one with 3% emulsifier/30% Emogreen® L19 released its full lubricating potential in the second part of the measurement (30-60 s). The obtained results revealed the strengths and weaknesses of each formulation, narrowing down their possible applications in the early development stage.

The Impact of the Oil Phase Selection on Physicochemical Properties, Long-Term Stability, In Vitro Performance and Injectability of Curcumin-Loaded PEGylated Nanoemulsions.

A nanotechnology-based approach to drug delivery presents one of the biggest trends in biomedical science that can provide increased active concentration, bioavailability, and safety compared to conventional drug-delivery systems. Nanoemulsions stand out amongst other nanocarriers for being biodegradable, biocompatible, and relatively easy to manufacture. For improved drug-delivery properties, longer circulation for the nanoemulsion droplets should be provided, to allow the active to reach the target site. One of the strategies used for this purpose is PEGylation. The aim of this research was assessing the impact of the oil phase selection, soybean or fish oil mixtures with medium chain triglycerides, on the physicochemical characteristics and injectability of curcumin-loaded PEGylated nanoemulsions. Electron paramagnetic resonance spectroscopy demonstrated the structural impact of the oil phase on the stabilizing layer of nanoemulsions, with a more pronounced stabilizing effect of curcumin observed in the fish oil nanoemulsion compared to the soybean oil one. The design of the experiment study, employed to simultaneously assess the impact of the oil phase, different PEGylated phospholipids and their concentrations, as well as the presence of curcumin, showed that not only the investigated factors alone, but also their interactions, had a significant influence on the critical quality attributes of the PEGylated nanoemulsions. Detailed physicochemical characterization of the NEs found all formulations were appropriate for parenteral administration and remained stable during two years of storage, with the preserved antioxidant activity demonstrated by DPPH and FRAP assays. In vitro release studies showed a more pronounced release of curcumin from the fish oil NEs compared to that from the soybean oil ones. The innovative in vitro injectability assessment, designed to mimic intravenous application, proved that all formulations tested in selected experimental setting could be employed in prospective in vivo studies. Overall, the current study shows the importance of oil phase selection when formulating PEGylated nanoemulsions.

Curcumin Loaded PEGylated Nanoemulsions Designed for Maintained Antioxidant Effects and Improved Bioavailability: A Pilot Study on Rats.

The current study describes the experimental design guided development of PEGylated nanoemulsions as parenteral delivery systems for curcumin, a powerful antioxidant, as well as the evaluation of their physicochemical characteristics and antioxidant activity during the two years of storage. Experimental design setup helped development of nanoemulsion templates with critical quality attributes in line with parenteral application route. Curcumin-loaded nanoemulsions showed mean droplet size about 105 nm, polydispersity index <0.15, zeta potential of -40 mV, and acceptable osmolality of about 550 mOsm/kg. After two years of storage at room temperature, all formulations remained stable. Moreover, antioxidant activity remained intact, as demonstrated by DPPH (IC50 values 0.078-0.075 mg/mL after two years) and FRAPS assays. In vitro release testing proved that PEGylated phospholipids slowed down the curcumin release from nanoemulsions. The nanoemulsion carrier has been proven safe by the MTT test conducted with MRC-5 cell line, and effective on LS cell line. Results from the pharmacokinetic pilot study implied the PEGylated nanoemulsions improved plasma residence of curcumin 20 min after intravenous administration, compared to the non-PEGylated nanoemulsion (two-fold higher) or curcumin solution (three-fold higher). Overall, conclusion suggests that developed PEGylated nanoemulsions present an acceptable delivery system for parenteral administration of curcumin, being effective in preserving its stability and antioxidant capacity at the level highly comparable to the initial findings.

'All-natural' anti-wrinkle emulsion serum with Acmella oleracea extract: A design of experiments (DoE) formulation approach, rheology and in vivo skin performance/efficacy evaluation.

OBJECTIVE: The growing consumers' preferences and concerns regarding healthy ageing, youthful skin appearance, environmental protection and sustainability have triggered an ever-increasing trend towards natural, eco-friendly and ethically sourced anti-ageing products. Accordingly, this paper describes design and evaluation of novel, safe, effective and high-quality emulsion serums, completely based on ingredients of natural origin, intended for improving facial fine lines and wrinkles. METHODS: Model formulations, stabilized by an innovative glycolipid mixed emulsifier (lauryl glucoside/myristyl glucoside/polyglyceryl-6 laurate) and containing Acmella oleracea extract as a model anti-ageing active, were prepared by cold process and fully assessed regarding their rheological behaviour (continuous rotational and oscillatory tests) and physical stability (dynamic-mechanical thermoanalysis - DMTA test). To study and optimize the simultaneous influence of varied formulation factors (emollients and emulsifier concentrations) on critical rheological attributes of the developed serums, a central composite design within 'design of experiments' approach was employed. The general skin performance - preliminary safety and anti-wrinkle efficacy of selected model serum, was evaluated in human volunteers, by employing several objective, non-invasive bioengineering techniques. RESULTS: Rheological characterization revealed favourable shear-thinning flow behaviour with yield point, and dominating elastic character (storage modulus G' > loss modulus G") in both amplitude and frequency sweeps, which together with relatively small structural change obtained in DMTA test indicated overall satisfying rheological and stability properties of formulated serums. From the established design space, and taking into account formulation cost and carbon footprint, promising model serum (desired/optimal apparent viscosity, yield point and loss factor, rather small and constant structural change), containing 15% of emollients and 1% of emulsifier, was chosen for in vivo evaluations. Screening of skin irritation effects revealed the absence of potential irritancy of investigated serum, suggesting overall satisfying skin tolerability/preliminary safety. Silicone skin replica image analysis demonstrated noticeable reduction/improvement in all measured skin wrinkle parameters after only 2 weeks of test serum application in periorbital and perioral areas, indicating its rapid and beneficial effects on the facial expression lines and wrinkles. CONCLUSION: Altogether, the results corroborate the promising potential of the developed Acmella oleracea extract-loaded emulsion serum as safe, effective and non-invasive natural anti-wrinkle product.

OBJECTIF: Les préférences et les préoccupations croissantes des consommateurs concernant le vieillissement sain, l'apparence jeune de la peau, la protection de l'environnement et la durabilité ont déclenché une tendance toujours croissante vers des produits anti-âge naturels, respectueux de l'environnement et éthiques. En conséquence, ce document décrit le plan et l'évaluation de nouveaux sérums d'émulsion sûrs, efficaces et de haute qualité, entièrement basés sur des ingrédients d'origine naturelle, destinés à améliorer les ridules et rides du visage. MÉTHODES: Des formulations modèles stabilisées par un émulsifiant mixte glycolipide innovant (lauryl glucoside/myristyl glucoside/polyglycéryl-6 laurate) et contenant de l'extrait d'Acmella oleracea comme anti-vieillissement actif de modèle, ont été préparées par un procédé à froid et ont été pleinement évaluées en ce qui concerne leur comportement rhéologique (tests de rotation continue et examens oscillatoires) et stabilité physique (analyse thermomécanique dynamique - DMTA). Pour étudier et optimiser l'influence simultanée de facteurs de formulation variés (concentrations d'émollients et d'émulsifiants) sur les attributs rhéologiques critiques des sérums développés, une conception composite centrale dans le cadre d'une approche « conception d'expériences ¼ a été employée. Les performances cutanées générales - sécurité préliminaire et efficacité antirides du sérum du modèle sélectionné ont été évaluées chez des sujets humains volontaires, en utilisant plusieurs techniques de bio-ingénierie objectives et non invasives. RÉSULTATS: La caractérisation rhéologique a révélé un comportement favorable du débit de cisaillement avec une limite de rendement et une domination du caractère élastique (modulus de stockage G' > module de perte G) dans les balayages d'amplitude et de fréquence, qui, avec un changement structurel relativement faible obtenu dans l'analyse DMTA, a indiqué des propriétés rhéologiques et de stabilité satisfaisante globales des sérums. A partir de l'espace de conception établi, et en tenant compte du coût de composition et de l'empreinte carbone, un sérum modèle prometteur (viscosité apparente souhaitée/optimale, seuil de rendement et facteur de perte, changement structurel assez faible et constant), contenant 15 % d'émollients et 1 % d'émulsifiant, a été choisi pour les évaluations in vivo. Le dépistage des effets d'irritation cutanée a révélé l'absence d'irritation potentielle du sérum expérimental, suggérant une tolérance cutanée/une sécurité préliminaire globalement satisfaisante. L'analyse de l'image de la réplique cutanée en silicone a démontré une réduction/amélioration notable de tous les paramètres de rides cutanées mesurés après seulement deux semaines d'application du sérum test dans les zones périorbitaires et péribuccales, indiquant ses effets rapides et bénéfiques sur les lignes d'expression et les rides du visage. CONCLUSION: Au total, les résultats corroborent le potentiel prometteur du sérum d'émulsion à base d'extrait d'Acmella oleracea développé comme un produit anti-rides naturel sûr, efficace et non invasif.

Identification and photostability of N-alkylamides from Acmella oleracea extract.

The identification of N-alkylamides from commercial Acmella oleracea extract, their UV-B photostability in different solvents, and identification of degradation products were the main goals of this study. By UHPLC-DAD-ESI-MS/MS method the presence of nine N-alkylamides was identified. Investigation of UV-B irradiation effect on identified N-alkylamides from Acmella oleracea extract was monitored in various the most commonly used solvents (methanol, ethanol, saline solution, and water) during 120 min. The results obtained indicated that spilanthol and homospilanthol were the most stable N-alkylamides presented in Acmella oleracea extract, while the photostability of identified N-alkylamides in whole in tested extract solutions decreased as follows: methanol>ethanol>saline solution>water. As the main degradation products in all investigated solutions 6,9-dihydroxy-deca-2,7-dienoic acid isobutyl-amide and 8,9-dihydroxy-deca-2,6-dienoic acid isobutyl-amide were identified.

Low-energy nanoemulsions as carriers for red raspberry seed oil: Formulation approach based on Raman spectroscopy and textural analysis, physicochemical properties, stability and in vitro antioxidant/ biological activity.

Considering a growing demand for medicinal/cosmetic products with natural actives, this study focuses on the low-energy nanoemulsions (LE-NEs) prepared via the Phase inversion composition (PIC) method at room temperature as potential carriers for natural oil. Four different red raspberry seed oils (ROs) were tested, as follows: cold-pressed vs. CO2-extracted, organic vs. non-organic, refined vs. unrefined. The oil phase was optimized with Tocopheryl acetate and Isostearyl isostearate, while water phase was adjusted with either glycerol or an antioxidant hydro-glycolic extract. This study has used a combined approach to formulation development, employing both conventional methods (pseudo-ternary phase diagram - PTPD, electrical conductivity, particle size measurements, microscopical analysis, and rheological measurements) and the methods novel to this area, such as textural analysis and Raman spectroscopy. Raman spectroscopy has detected fine differences in chemical composition among ROs, and it detected the interactions within nanoemulsions. It was shown that the cold-pressed, unrefined, organic grade oil (RO2) with 6.62% saturated fatty acids and 92.25% unsaturated fatty acids, was optimal for the LE-NEs. Textural analysis confirmed the existence of cubic gel-like phase as a crucial step in the formation of stable RO2-loaded LE-NEs, with droplets in the narrow nano-range (125 to 135 nm; PDI ≤ 0.1). The DPPH test in methanol and ABTS in aqueous medium have revealed a synergistic free radical scavenging effect between lipophilic and hydrophilic antioxidants in LE-NEs. The nanoemulsion carrier has improved the biological effect of raw materials on HeLa cervical adenocarcinoma cells, while exhibiting good safety profile, as confirmed on MRC-5 normal human lung fibroblasts. Overall, this study has shown that low-energy nanoemulsions present very promising carriers for topical delivery of natural bioactives. Raman spectroscopy and textural analysis have proven to be a useful addition to the arsenal of methods used in the formulation and characterization of nanoemulsion systems.

Dynamic-mechanical thermoanalysis test as a high-performance alternative for accelerated freeze-thaw stability testing: a case study of O/W emulsions.

Objective: The main objective of this work was to evaluate the performance of recently developed dynamic-mechanical thermoanalysis (DMTA) test as a rapid rheological alternative to conventional freeze-thaw cycling for accelerated stability testing of oil-in-water (O/W) emulsions.Significance: The rational for this approach was reducing the time needed for product and process development and optimization, potentially through shortening the time needed for stability evaluation, in order to keep the pace with high formulating turnover imposed by increasing demands for placing products on the market, that is, to facilitate decision making in R&D and QC settings.Methods: Six model O/W emulsions were designed, rheologically characterized (continuous rotational and oscillatory tests), and subjected to stability evaluation through freeze-thaw test in stability chamber and DMTA tests using an air-bearing rheometer.Results: Investigated samples were characterized by favorized shear-thinning flow behavior with yield point. The elastic behavior dominated the viscous one in the LVE region of amplitude sweeps, as well as in the frequency sweeps of used frequency range. Statistical method comparison studies demonstrated that the results obtained by freeze-thaw test, routinely used for accelerated stability testing of emulsions, were in good accordance with those obtained with DMTA tests, whereas the time needed for stability assessment was significantly reduced (2-6 h versus 12 days).Conclusions: In summary, DMTA test proved to be an expeditious alternative for accelerated freeze-thaw stability testing of O/W emulsions, with great promise in new product development and optimization (R&D), as well as in determination of borderline product batches status (QC).

Dynamic-mechanical thermoanalysis test: a rapid alternative for accelerated freeze-thaw stability evaluation of W/O emulsions.

Objective: The aim of this study was to develop a new dynamic-mechanical thermoanalysis (DMTA) test and evaluate its performance as rapid rheological alternative to routinely used freeze-thaw test for accelerated stability testing of water-in-oil (W/O) emulsions.Significance: Due to inherent emulsion instability and versatilities of storage and use conditions, stability assessment of emulsion products still remains complex and challenging task. Recommended stability evaluation protocols are time-consuming, imposing need for alternate test procedures, especially in the early stage of product development, as well as in the quality assurance setting, including quality control.Methods: Five model W/O emulsions were prepared, comprehensively rheologically characterized (continual and oscillatory tests), and subjected to stability evaluation through freeze-thaw test in stability chamber and DMTA tests using an air-bearing rheometer.Results: Analyzed emulsions displayed desired shear-thinning flow behavior with yield point. The storage modulus dominated over the loss modulus in the linear viscoelastic regions of amplitude sweeps, as was the case in frequency sweeps over entire frequency range. Statistical comparison showed good agreement between freeze-thaw test, as a method available and used in daily routine work for accelerated evaluation of the physical stability, and DMTA test, as a rheological simulation of the said routine method. Duration of DMTA test was significantly shorter compared to routine but lengthy freeze-thaw test (3.5 h versus 12 days).Conclusions: According to our results, DMTA test could be a rapid alternative for accelerated freeze-thaw stability evaluation of W/O emulsions, thus enabling high formulating turnover and decision making in R&D and QC departments.

Combined use of biocompatible nanoemulsions and solid microneedles to improve transport of a model NSAID across the skin: In vitro and in vivo studies.

This study aimed to investigate the potential of lecithin-based nanoemulsions costabilized by sucrose esters, with and without skin pretreatment with stainless steel microneedles, to improve delivery of aceclofenac, as a model drug, into/across the skin. The characterization revealed favorable droplet size (about 180 nm), narrow size distribution (<0.15), high surface charge (about -40 mV) and satisfying long-term stability (one year at 4 ±â€¯1 °C) of the formulation costabilized by sucrose palmitate, demonstrating a similar trend observed for the reference stabilized by widely used lecithin/polysorbate 80 combination. In vitro release/permeation testing and differential stripping on the porcine ear proved the superiority of the sucrose ester- over polysorbate-based nanoemulsion. However, in vitro findings were not fully indicative of the in vivo performances - no significant differences were observed between investigated formulations in pharmacokinetic profile and total amount of aceclofenac deposited in the rat skin 24 h after dosing, simultaneously pointing to delayed aceclofenac delivery into the systemic circulation. In addition, the ratio of plasma concentrations of aceclofenac and its major metabolite in rats, diclofenac, was remarkably changed after topical application of tested nanoemulsions compared to intravenous administration of aceclofenac solution. Finally, skin pretreatment with microneedles improved aceclofenac delivery into/across the rat skin from tested formulations, resulting in 1.4-2.1-fold increased bioavailability and 1.2-1.7-fold enhanced level of aceclofenac retained in the skin, as measured 24 h after administration. Moreover, the plasma concentrations of aceclofenac 24 h after application of tested formulations (lecithin/sucrose palmitate vs. lecithin/polysorbate 80) combined with microneedles (173.37 ±â€¯40.50 ng/ml vs. 259.23 ±â€¯73.18 ng/ml) were significantly higher than those obtained through intact skin (105.69 ±â€¯19.53 ng/ml vs. 88.38 ±â€¯14.46 ng/ml). However, obtained results suggest that combination of microneedles and sucrose palmitate-costabilized nanoemulsion could be useful to attain higher skin concentration, while combination of microneedles with polysorbate 80-costabilized nanoemulsion could be a preferable option for enhancing drug delivery into the bloodstream.

Sulfanilamide in solution and liposome vesicles; in vitro release and UV-stability studies.

The main goal of this study was to develop a liposome formulation with sulfanilamide and to investigate the liposomes impact on its release and stability to the UV-A/UV-B and UV-C irradiation. Liposome dispersions with incorporated sulfanilamide were prepared by thin-film hydration method and liposomes role to the sulfanilamide release was investigated by using a dialysis method. Comparatively, sulfanilamide in phosphate buffer solution was subject to release study as well to the UV irradiation providing for the possibilities of kinetics analysis. In vitro drug release study demonstrated that 20% of sulfanilamide was released from liposomes within 1 h that is approximately twice as slower as in the case of dissolved sulfanilamide in phosphate buffer solution. The kinetic release process can be described by Korsmeyer-Peppas model and according to the value of diffusion release exponent it can be concluded that drug release mechanism is based on the phenomenon of diffusion. The sulfanilamide degradation in phosphate buffer solution and liposomes is related to the formation of UV-induced degradation products that are identified by UHPLC/MS analysis as: sulfanilic acid, aniline and benzidine. The UV-induced sulfanilamide degradation in the phosphate buffer solution and liposome vesicles fits the first- order kinetic model. The degradation rate constants are dependent on the involved UV photons energy input as well as sulfanilamide microenvironment. Liposome microenvironment provides better irradiation sulfanilamide stability. The obtained results suggest that liposomes might be promising carriers for delayed sulfanilamide delivery and may serve as a basis for further research.