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- Ethosomes: an approach for bioactive plant extract preservation envisaging cosmetic applicationsPublication . Plasencia, Paula; Santamaria-Echart, Arantzazu; Heleno, S.A.; Colucci, Giovana; Garcia, Pablo; Barros, Lillian; Barreiro, FilomenaThe present work is focused on upgrading the commercial potential of berry crop by-products by encapsulating them into liposomes to preserve their bioactivity. The extracts have been obtained with an ethanol-water mixture using ultrasound-assisted extraction, and the most promising ones were encapsulated in ethosome system. To achieve this goal, ethosomes were prepared using the cold method. Ethosomal suspensions were characterized concerning particle size distribution by laser dispersion, differential scanning calorimetry, infrared Fourier-transform spectroscopy, encapsulation efficiency, and morphological analysis using optical, scanning electron, and transmission electron microscopy. These results indicate that ethosomes are an appropriate method to encapsulate hydroethanolic bioactive plant bioresidue extracts and a good option to preserve them for further use in industrial applications, such as cosmetics. Future work will include optimizing the process and proof of concept by developing a cosmetic application.
- Lignin from aldehyde-assisted fractionation can provide light-colored Pickering emulsions through colloidal particles formed using alkaline antisolventPublication . Colucci, Giovana; Ribeiro, Andreia; Figueirêdo, Monique Bernardes; Charmillot, Justine; Santamaria-Echart, Arantzazu; Rodrigues, Alírio; Barreiro, FilomenaColloidal lignin particles (CLPs) are gaining attention as eco-friendly stabilizers for Pickering emulsions. Still, conventional lignin sources, like kraft lignin, are often limited by their dark color and strong odor. This study explores, for the first time, the use of a light-colored lignin derived from an aldehyde-assisted fractionation with glyoxylic acid (GA-lignin) for producing CLPs and derived Pickering emulsions. CLPs were produced by anti- solvent precipitation with water (CLPs-W, pH 6) and alkaline buffer (CLPs-B, pH 8) as the antisolvents. The results revealed that the selected antisolvent significantly influenced the CLPs’ properties. CLPs-W were larger, uniform in size, and hydrophobic, whereas CLPs-B were smaller, agglomerated into clusters, and exhibited greater hydrophilicity. Despite both CLPs’ effectiveness in stabilizing oil-in-water emulsions, the stabilization mechanisms differed markedly; CLPs-W formed a robust membrane barrier at the oil-water interface, while CLPs- B facilitated oil droplet bridging. Overall, this work demonstrates that GA-lignin’s light color nature offers ad- vantages for Pickering emulsions design, surpassing a lignin typical limitation. This advancement highlights the versatility of GA-lignin-derived CLPs and supports the development of sustainable lignin-based products with significant commercial prospects
- Development of functional finishes based on natural products for leather applicationsPublication . Colucci, Giovana; Barreiro, M.F.; Fernandes, Isabel P.; Sipoli, Caroline CasagrandeNanoemulsions are biphasic systems characterized by having droplets with nanometer size range, which confers them several advantages, such as high hydration capacity and increased bio-efficacy of the functionalities incorporated therein. In this context, the objective of this work was to develop a water-in-oil (W/O) nanoemulsion system based on natural compounds, like sweet almond oil and Cinnamomum zeylanicum aqueous extract, in order to produce a moisturizing and antimicrobial spray for leather application. Firstly, a systematic study was performed by testing different base formulations to establish the procedure for nanoemulsion preparation using a high-pressure homogenizer (HPH). A total of 16 base formulations were prepared by varying the W/O ratio and emulsifier composition. These samples were then analyzed by microscopic and visual analysis in order to evaluate the droplets size and morphology together with stability along the time. Based on these results, the best formulation revealed to be the system S4, corresponding to a W/O ratio of 40/60 (v/v), and Span 80/Tween 80 mixture (54/46 ratio, v/v) as emulsifier. Using this formulation, nanoemulsions functionalized with cinnamon at contents of 1.25%; 2.5%; 3.75%; 5% (w/v; water base) were prepared. Afterwards, these products, together with the base system, were characterized by Confocal Laser Scanning Microscopic (CLSM), color parameters, evaluated concerning the antimicrobial and antioxidant bioactivities, and stability. As results, CLSM confirmed the nature of the water-in-oil system, revealing also a change in the nanoemulsions microstructure due to the increase of cinnamon extract concentration. The color measurement evidenced a luminosity diminishing and a color variation to yellow and red as the extract content was increased. In addition, the formulations containing the cinnamon extract evidenced antimicrobial and antioxidant activity. The absence of bioactivity for the base nanoemulsion corroborated the effect of the cinnamon extract in the detected antimicrobial and antioxidant. Furthermore, for the antimicrobial activity, the formulations containing the extract presented a prolonged effect against E. coli and S. aureus bacteria, being detected similar inhibition zones for all the tested extract concentrations. Regarding the antioxidant activity, the rise in the extract concentration resulted in a relevant increase of this property. Finally, and according to the performed accelerated stability tests, a high stability against centrifugation for the base nanoemulsion was detected, evidencing that the presence of the extract contributes to the formulation instability. Beyond this, all the formulations showed high stability against thermal stress. From a general perspective, the results obtained are of great importance for the product design and development having in view the application as leather preservative and moisturizer, since the formulations presented antioxidant and antimicrobial activity, parameters that contribute to the protection and preservation of the shoes. In addition, their stability points for a good shelf life of these systems, being suitable for storage under commercialization conditions.
- Spirulina (Arthrospira platensis) protein-rich extract as a natural emulsifier for oil-in-water emulsions: optimization through a sequential experimental design strategyPublication . Silva, Samara Cristina; Almeida, Thayná Domingues; Colucci, Giovana; Santamaria-Echart, Arantzazu; Manrique, Yaidelin A.; Dias, Madalena M.; Barros, Lillian; Fernandes, Ângela; Colla, Eliane; Barreiro, M.F.Spirulina (Arthrospira platensis) proteins have been proven to present emulsifying properties. In this work, a Spirulina protein-rich extract obtained by ultrasound extraction (SpE) was tested to stabilize oil-in-water (O/W) emulsions. For this purpose, a sequential experimental design strategy (Fractional Factorial Design (FFD) 24–1 followed by a Central Composite Rotatable Design (CCRD) 22)) was applied. The effect of four variables, SpE concentration, O/W weight ratio, pH and storage time, on emulsions’ zeta potential and number-mean droplet diameter was considered for the FFD 24–1, indicating SpE concentration and storage time as the relevant variables for the CCRD 22. According to zeta potential and number-mean droplet diameter evaluation, for the studied SpE concentration range (2–5 wt%), quite stable emulsions were obtained along the tested 30-days period. Even so, for 5%, visual inspection revealed extract segregation after 20-days. The optimal solution comprised 4 wt% of SpE, for an O/W weight ratio of 30/70 and a pH of 7.0 (number-mean droplet diameter of 55.66 nm and zeta potential of -43.83 mV). Overall, SpE has proven to be an excellent emulsifier, offering the potential to substitute animal-based proteins and synthetic emulsifiers. In addition, no signs of contamination by microorganisms were observed, suggesting that the SpE may also act as an antimicrobial agent.
- Development of water-in-oil emulsions as delivery vehicles and testing with a natural antimicrobial extractPublication . Colucci, Giovana; Santamaria-Echart, Arantzazu; Silva, Samara Cristina; Fernandes, Isabel P.; Sipoli, Caroline C.; Barreiro, M.F.Water-in-oil (W/O) emulsions have high potential for several industrial areas as delivery systems of hydrophilic compounds. In general, they are less studied than oil-in-water (O/W) systems, namely in what concerns the so-called fluid systems, partly due to problems of instability. In this context, this work aimed to produce stable W/O emulsions from a natural oil, sweet almond oil, to be further tested as vehicles of natural hydrophilic extracts, here exemplified with an aqueous cinnamon extract. Firstly, a baseW/O emulsion using a high-water content (40/60, v/v) was developed by testing di erent mixtures of emulsifiers, namely Tween 80 combined with Span 80 or Span 85 at di erent contents. Among the tested systems, the one using a 54/46 (v/v) Span 80/Tween 80 mixture, and subjected to 12 high-pressure homogenizer (HPH) cycles, revealed to be stable up to 6 months, being chosen for the subsequent functionalization tests with cinnamon extract (1.25–5%; w/v; water-basis). The presence of cinnamon extract leaded to changes in the microstructure as well as in the stability. The antimicrobial and antioxidant analysis were evidenced, and a sustained behavior compatible with an extract distribution within the two phases, oil and water, in particular for the higher extract concentration, was observed.
- New trends in natural emulsifiers and emulsion technology for the food industryPublication . Santamaria-Echart, Arantzazu; Fernandes, Isabel P.; Silva, Samara Cristina; Rezende, Stephany; Colucci, Giovana; Dias, Madalena M.; Barreiro, M.F.The food industry depends on using different additives, which increases the search for effective natural or natural-derived solutions, to the detriment of the synthetic counterparts, a priority in a biobased and circular economy scenario. In this context, different natural emulsifiers are being studied to create a new generation of emulsion-based products. Among them, phospholipids, saponins, proteins, polysaccharides, biosurfactants (e.g., compounds derived from microbial fermentation), and organic-based solid particles (Pickering stabilizers) are being used or start to gather interest from the food industry. This chapter includes the basic theoretical fundamentals of emulsions technology, stabilization mechanisms, and stability. The preparation of oil-in-water (O/W) and water-in-oil (W/O) emulsions, the potential of double emulsions, and the re-emerging Pickering emulsions are discussed. Moreover, the most relevant natural-derived emulsifier families (e.g., origin, stabilization mechanism, and applications) focusing food applications are presented. The document is grounded in a bibliographic review mainly centered on the last 10-years, and bibliometric data was rationalized and used to better establish the hot topics in the proposed thematic.
- Development of chitosan microspheres through a green dual crosslinking strategy based on tripolyphosphate and vanillinPublication . Correa, Rodolpho Fagundes; Colucci, Giovana; Halla, Noureddine; Pinto, João A.; Santamaria-Echart, Arantzazu; Blanco, Silvia Priscila; Fernandes, Isabel P.; Barreiro, M.F.Microencapsulation procedures have recently focused attention on designing novel mi-crospheres via green synthesis strategies. The use of chitosan (CS) as an encapsulating material has increased interest due to its unique bioactive properties and the various crosslinking possibilities offered by their functional groups. The consolidation of the microspheres by physical crosslinking using sodium tripolyphosphate (TPP) combined with chemical crosslinking using vanillin (VA) open new opportunities in the framework of green dual crosslinking strategies. The developed strategy, a straightforward technique based on an aqueous medium avoiding complex separa-tion/washing steps, offers advantages over the processes based on VA, mostly using water-in-oil emulsion approaches. Thus, in this work, the combination of TPP crosslinking (3, 5, and 10 wt.%) via spray-coagulation technique with two VA crosslinking methods (in situ and post-treatment using 1 wt.% VA) were employed in the preparation of microspheres. The microspheres were characterized concerning morphology, particle size, physicochemical properties, thermal stability, and swelling behavior. Results revealed that the combination of 5 wt.% TPP with in situ VA crosslinking led to microspheres with promising properties, being an attractive alternative for natural bioactives encapsulation due to the green connotations associated with the process.
- Evaluation of saponin-rich extracts as natural alternative emulsifiers: A comparative study with pure Quillaja Bark saponinPublication . Schreiner, Tatiana B.; Colucci, Giovana; Santamaria-Echart, Arantzazu; Fernandes, Isabel P.; Dias, Madalena M.; Pinho, Simão; Barreiro, M.F.Due to synthetic surfactants' environmental impact, their replacement by natural alternatives has gained relevance, with saponins emerging as sustainable approaches. In this work, three saponin-rich extracts from different sources (Tribulus terrestris (TT), Trigonella foenum-graecum (FG), and Ruscus aculeatus (RA)) were tested as emulsifiers, and their performance compared with Quillaja Bark saponin (PS). Characterisation comprised FTIR, solubility studies, CMC assays, and emulsifying properties (emulsifying capacity (EC) and foaming capacity (FC)). For all samples, solubility assays indicated high solubility in water and low in apolar solvents (e.g., n-hexane), compatible with their O/W emulsifier character. In general, the saponin content ruled extracts' performance (PS > TT > FG > RA). EC values (without pH adjustment) were found to be 82.5, 55.0, 47.5, 36.3%, respectively. When pH changed for 7 and 9, a shift in FG and RA order was observed. The pseudo-ternary diagrams, constructed to map emulsion's composition zones, indicate the formation of single-phase systems in the region of low oil and high extract content. Except for RA extract, gel samples were formed, which are interesting technological solutions for several applications. Among the studied samples, and in alternative to PS, TT extract showed the best performance.
- Spirulina (Artrhospira platensis) protein extract: techno-functional properties and potential application as a natural emulsifierPublication . Silva, Samara Cristina; Almeida, Thayná Domingues; Colucci, Giovana; Santamaria-Echart, Arantzazu; Manrique, Yaidelin A.; Dias, Madalena M.; Barros, Lillian; Fernandes, Ângela; Colla, Eliane; Barreiro, M.F.Microalgae are valuable sources of proteins (20-70%) and have emerged as great alternatives to animal-based proteins (e.g., whey protein, egg yolk)1. This work aimed to obtain a microalga protein extract (SpE) from Artrhospira platensis (Spirulina) and evaluate its techno-functional properties and emulsion formation capability. The SpE was obtained by ultrasound and conventional mechanical stirring under an alkaline medium (pH 9.0) followed by acid precipitation (pH 3.0)2. In addition, the SpE techno-functional (protein solubility; oil and water absorption capacities) and structural properties (FTIR analysis) were investigated. Moreover, an emulsion (oil/water ratio (30/70); pH 7.0; SpE concentration (3.5%)) was prepared according to two sequential steps: i) high-shear homogenization (UltraTurrax; 11,000 rpm; 3 min) and ii) high-pressure homogenization (100MPa; 6 cycles)3. Then, it was analysed by optical microscopy, zeta potential and particle size. The SpE (66.6%±0.31 protein) presented the lowest solubility at pH 3.0 and the highest at pH 8.0. The protein extract showed higher absorption capacity in oil (21.7±0.10 g oil/ g SPE) than in water (10.3±0.50 g water/ g SPE). From the FTIR analysis (Figure 1a), the SpE showed two peaks at 1635 cm-1 and 1535 cm-1 (red circle in Figure 1a) attributed to the presence of characteristic protein groups, amide I and amide II, respectively. The oil-in-water emulsion (ESp) presented a light blue-green colour due to the presence of SpE (Figure 1a) and remained stable for 30 days (Figure 1b). The SpE presented a zeta potential of -47.20 mV and a particle size of 330 nm (D50, volume distribution) on the 30th storage day. Small and spherical droplets were observed, evidencing the particle size and stability over the storage time (Figure 1c). Thus, the SpE showed the ability to form emulsions with stability for up to 30 days, being a great alternative to replace animal-based and synthetic emulsifiers.
- Saponin-based natural nanoemulsions as alpha-tocopherol delivery systems for dermal applicationsPublication . Schreiner, Tatiana B.; Santamaria-Echart, Arantzazu; Colucci, Giovana; Plasencia, Paula; Costa, Patrícia Santos; Dias, Madalena M.; Pinho, Simão; Barreiro, M.F.Nanoemulsions can be produced using simple methods and compounds from natural sources. They can increase water dispersibility and bioavailability and optimise active ingredient dispersion in particular skin layers. Lipophilic compounds of the vitamin E family (tocopherols and tocotrienols) are well-known for their high antioxidant activity and capacity to protect the skin from oxidative stress. In this context, oil-in-water (o/w) nanoemulsions with and without α-tocopherol (Vitamin E, VE) were formulated with two emulsifier alternatives, Quillaja saponin (QS), and a combination of QS with Tribulus terrestris (QSTT) (50/50, w/w). The emulsions were evaluated concerning stability, microstructure, droplet size, colour attributes, encapsulation efficiency, UV photostability, antioxidant activity, and in vitro permeation studies to assess the delivery potential. Results showed highly stable systems, with round-shape droplets of 80–121 nm size. QS and QSTT samples’ colours were close to white and light brownish, respectively. The topical nano cream had the capacity to entrap VE, producing a protective effect from UV degradation, and very significant antioxidant activity, with IC50 values around 0.01 %wt. The skin permeation profiles showed the efficiency of the formulations in the delivery of VE, with permeabilities between 64 and 74 μg/cm2, while the control sample showed no VE permeation.