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- Response surface methodology in the optimization of extraction conditions for Gracilaria gracilis extracts for use in thermoplastic food coatingsPublication . Reboleira, João; Andrade, Mariana A.; Sanches-Silva, Ana; Ganhão, Rui; Mendes, Susana; Vilarinho, Fernanda; Félix, Rafael; Pinheiro, Joaquina; Bernardino, Susana M.Incorporation of antioxidant agents in edible films and packages often relies in the usage of essential oils and other concentrated hydrophobic liquids, with reliable increases in antimicrobial and antioxidant activities of the overall composite, and consequent improvements in product shelf life. These oils are often obtained from plant sources and the usage of organic solvents in their extraction has been criticized as to whether it constitutes a health hazard for consumers [1, 2, 3]. The use of water-soluble antioxidant agents is less common, due to lower activities and extraction yields. As such, using commonly available seaweeds as the source for both the main polymers and the supplementing antioxidant/antimicrobial agents in bioactive films can potentially reduce production costs and create a safer, more sustainable product [1]. Hydroethanolic extracts of commercially available red macroalgae Gracilaria gracilis were evaluated for their antioxidant potential and phenolic content, as part of the preliminary assays for the selection of algal biomass for the enrichment of thermoplastic films. The extracts were obtained through use of solid-liquid extractions, over which yield, DPPH radical reduction capacity, total phenolic content, and FRAP activity assays were measured [4, 5]. Solid to liquid ratio (SLR), extraction time, and ethanol to water ratio were selected as independent variables with experimental ranges and configurations obtained using a Box-Behnken design with three factors, resulting in 15 experimental conditions. Extraction duplicates were used [4]. Response surface methodology was then used to estimate the effect of each extraction condition on the tested bioactivities. Bioactivities were significantly (p<0.05) and positively affected by the presence of ethanol in the extraction solvent, while extraction yield was reduced. Lower solid to liquid ratios (higher solvent proportions) resulted in higher yields, but had no significant (p<0.05) impact on bioactivities. Time was not a relevant factor in any of the measured variables, and as such will be minimized in any further work. 100% ethanol with minimum extraction time (10 minutes) and 1g to 5mL SLR maximize antioxidant activities per gram of dry extract. Maximum ethanolic yields were obtained with a theoretical extraction of 100 minutes and a 1g to 25mL SLR. Aqueous extractions displayed similar results in terms of optimum conditions, but with overall lower bioactivities and higher yields, very likely due to high amounts of soluble polysaccharides. While the high antioxidant activities from the ethanolic extracts encourage the selection of this solvent for extraction of bioactives, low yields may make them later unfeasible. Future studies over the compatibility of these extracts in the polymeric matrix of the films, as well as further optimization of the extraction process will be necessary before the definitive choice for bioactive origin and processing is made.
- A pilot plant scale testing of the application of seaweed-based natural coating and modified atmosphere packaging for shelf-life extension of fresh-cut applePublication . Augusto, Ana; Miranda, Andreia; Costa, Leonor; Pinheiro, Joaquina; Campos, Maria J.; Raimundo, Délio; Pedrosa, Rui; Mitchell, Geoffrey; Niranjan, Keshavan; Silva, Susana F.J.Codium tomentosum hydroethanolic extract was obtained using a pilot solid–liquid extractor to validate the anti-browning functionality of the extract under industrial conditions. Fresh-cut apple slices were coated by immersion in: (1) a seaweed extract solution (0.5% w/v) and (2) a commercial coating, and the two sets of samples were compared with a control (immersion in water). Packaged samples were stored, under ambient and modified atmosphere conditions at 4°C. After 30 days of storage, the samples that were coated with the seaweed extract and packaged under modified atmosphere, demonstrated lower peroxidase activity and polyphenol oxidation when compared with the samples treated with the commercial additive. These results confirm, at pilot scale and under industrial production conditions, the efficacy of the seaweed extract as a bio-based substitute for the synthetic coatings, which are currently used to prevent browning in fresh-cut apples.
- Supercritical CO2 extraction of Aurantiochytrium sp. biomass for the enhanced recovery of omega-3 fatty acids and phenolic compoundsPublication . Melo, M.M.R. de; Sapatinha, M.; Pinheiro, J.; Lemos, M.F.L.; Bandarra, N.M.; Batista, I.; Paulo, M.C.; Coutinho, J.; Saraiva, J.A.; Portugal, I; Silva, C.M.The microalgae Aurantiochytrium sp. is a strong alternative source of ω-3 fatty acids, including ocosahexaenoic acid (DHA). This work encompasses the optimization of SFE conditions to maximize the total extraction yield (ηTotal), DHA content (CDHA), total phenolics content (TPC), and antioxidant capacity (AOC) of the extracts produced from Aurantiochytrium sp. biomass. A full factorial experimental plan was performed, comprising three factors (pressure, temperature, and flow rate) and two levels (200−300 bar, 40−80 °C, and 6–12 gmin−1, respectively). The maximum and minimum experimental results were ηTotal = 2.1 and 13.4 wt.%, CDHA= 27.3 and 39.3 wt.%, TPC =1.19 and 2.24 mgGAE g−extract 1 , and AOC = 0.3 and 1.4 mg g− TEAC extract 1. Under the studied experimental conditions, increasing pressure up to 300 bar is the optimum to rise both ηTotal and CDHA. Temperature increase from 40 to 80 °C leads to opposing effects: it favors the concentration of phenolics in the supercritical extracts at the expenses of decreasing DHA content and total yield. Surface models were adjusted to ηTotal, CDHA and TPC data, and the goodness of the fits ranged from coefficients of determination of 0.752-0.711 (TPC) to 0.997-0.994 (CDHA). Under optimized conditions, supercritical extracts exhibited a DHA content more than 3.5-fold richer than fish oil, and 7.9-fold richer than the best alternative microalgae species (Pavlova lutheri) found in the literature.
- Um guia para uma alimentação sustentável, segura e saudávelPublication . Pinheiro, Joaquina; Gailanes, Alfredo; Farinha, Ana Margarida; Portugal, Artur; Costa, Daniela; Vieira, Inês; Rafael, Célia
- Quality changes during thermal processing of two mixed formulas of fruits and vegetables pulpsPublication . Gonçalves, Elsa M.; Raposo, Isa; Pinheiro, Joaquina; Alegria, Carla; Moldão, Margarida; Abreu, MartaThe present work aimed to evaluate, through thermal degradation kinetics (80 °C to 98 °C in time intervals of 0.5 to 25 min), the effects of different thermal treatments on the biochemical, physicochemical, sensory and microbiological parameters of two mixed fruit and vegetable pulps, a yellow and a red one. The evaluated fruit and vegetable pulps resulted from the mixture of different fruits and vegetables proportions (pineapple, beetroot, strawberry and lemon juice) added to a 50% (p/p) pear-based pulp to maximize their bioactivity, physicochemical stability and sensorial acceptance. Evaluated quality parameters included the determination of peroxidase activity (POD), pH, soluble solids content (SSC), total phenolic content (TPC), CIELab colour, sensory evaluation (colour, taste and aroma) and total mesophilic aerobic counts (TAPC). Regarding heat treatments optimization for both pulps with lower pH, it was concluded that higher temperature treatments (90 to 98 °C) applied over a shorter time (less than 5 min) were more effective to inactivate POD, to reduce the initial microbial load (>2 log10 cycles) and to maximize sensorial attributes. In both mix pulps, total phenolic content (TPC) was not significantly influenced by the different applied time-temperature binomials. From the degradation kinetic models and as an example, it was possible to conclude that POD followed a 1st order kinetic, where the temperature effect was well fitted to the Arrhenius equation. The results allowed to obtain optimized time-temperature binomials for each pulp to simultaneously achieve POD enzyme inactivation, microbial reduction, and maximization of quality parameters relatively to fresh pulps, 90 °C/5 min and 98 °C/2.5 min, for the yellow pulp and red pulp, respectively.
- Effect of airflow conditions on growth and simultaneously DHA and squalene production by Aurantiochytrium sp. PRA-276Publication . Paulo, M.C.; Coutinho, J.; Saraiva, J.A.; Pinto, C.A.; Amaral, R.; Melo, M.M.R.; Pinheiro, J.; Lemos, M.F.L.; Cardoso, C.; Sapatinha, M.; Bandarra, N.M.There has been a growing trend worldwide in the use of squalene and docosahexaenoic acid (DHA), due in part to their recognized health benefits and their range of applications. However, the growth of the industry exploiting these resources has been extremely limited by the scarcity and limitations of raw material, leading to the need to develop more sustainable sources of production. Aurantiochytrium sp. PRA-276, with the ability to produce high concentrations of DHA and squalene, is a promising candidate for the commercial production of these biomolecules. The aim of this study was to determine the cumulative effect of airflow levels and stirring speed of Aurantiochytrium sp. PRA-276 biomass during DHA and squalene production. Different airflow levels (3, 6 and 9 L min-1) and two stirring speeds (150 and 300 rpm) were investigated. The highest biomass production was achieved after 96 hours with 300 rpm stirring speeds and airflow level of 9 L min-1. The maximum total lipids content reached 61.4% of the dried biomass, and the optimal parameters for DHA and squalene production were stirring speed 150 rpm and airflow level 3 L min-1. The maximum yield of DHA production was 4.3 g L-1 at 96 h of fermentation. The highest yield of squalene production was 762.5 mg L-1 at 48 hours of growth. The results show that Aurantiochytrium sp. PRA-276 is a promising resource for microbial DHA and squalene production due to the high yield of DHA and squalene that it generates. Moreover, the capacity for large-scale fermentation of this organism could be useful for the synthesis of biomass related products with high added value.
- Characterization of nutritional, physicochemical, and phytochemical composition and antioxidant capacity of three strawberry “Fragaria X ananassa Duch.” Cultivars (“Primoris”, “Endurance”, and “Portola”) from western region of PortugalPublication . Ganhão, Rui; Pinheiro, Joaquina; Tino, Clara; Faria, Hugo; Gil, Maria MIn this study, nutritional composition (protein, lipids, carbohydrates, ash, and moisture), physicochemical properties (soluble solid content, titratable acidity, texture and instrumental colour on surface, and internal section), phytochemicals (total phenolic content and anthocyanin content), and antioxidant capacity (DPPH—2,2'-diphenyl-1-picrylhydrazyl radical scavenging capacity and ferric-reducing antioxidant power) of three strawberry (Fragaria X ananassa Duch.) cultivars (cv. “Primoris”, cv. “Endurance”, and cv. “Portola”) produced in the western region of Portugal (Caldas da Rainha) were evaluated. From the obtained, results no significant differences (P > 0.05) in nutritional composition were detected in all of the cultivars; with the exception of lower protein content observed in cv. “Portola” (0.57 g/100 g +- 0.04; P < 0.05). Regarding the a* value of whole strawberry fruits, no significant differences (P > 0.05) were found in any of the cultivars, which revealed a similar redness. The cv. “Endurance” revealed the highest bioactivity content compared to the other cultivars. Overall, these results provide important information about the high quality of strawberry produced in the western region of Portugal and may be used as a tool for adding value to a functional food in the Mediterranean diet due to the phytochemical composition and nutritional value of strawberry fruits.
- Effect of heat treatment on smoothie quality by response surface methodologyPublication . Pinheiro, Joaquina; Santos, Diana I.; Gonçalves, Elsa M.; Abreu, Marta; Moldão-Martins, MargaridaSmoothies are a popular and convenient way for to consume bioactive compounds from fruits and vegetables such as total phenolics, carotenoids and flavonoids, with the preservation treatment being an important action to guarantee the safety and extension of shelf-life. The main goal of this study was to evaluate the impact of heat treatment (HT) on smoothie prepared with “Fuji “apple (41%), pineapple (31%), cabbage (8%), pumpkin (10%) and banana (10%), by response surface methodology (RSM), where the temperature (70–100 °C) and treatment time (0.5–10.5 min), were the dependent variables. After optimization of HT conditions, a validation assay was performed to guarantee the minimal changes on color and reduction of 90% of polyphenoloxidase enzyme (PPO). Antioxidant activity (Ferric reducing antioxidant power assay (FRAP), DPPH, ABTS), total phenolics content (TPC), pH and solids soluble content were also analyzed. Predicted models of color parameters (L*, a*, ºh) and PPO enzymatic activity were found to be significant (p < 0.05) with regression coefficients (R2) of 0.84, 0.86, 0.92 and 0.97, respectively. From the RSM-generated model, the HT conditions that ensure a minimal green loss of smoothie and inactivation of PPO enzyme was at 85 °C over 7 min. In the validation study, these conditions were tested and proved to be sufficient to achieve the main goals. In the heat-treated smoothie, increases in TPC (10%) and antioxidant capacity (ABTS: 50%, DPPH: 17%, FRAP: 13%) were attained. This study demonstrated that RSM was efficient to select the optimal conditions of HT and improve the important quality properties that influence the product quality and the potential consumer’s health (TPC and antioxidant capacity).
- Bioactivity screening of Aurantiochytrium sp. for antioxidant activitiesPublication . Pinheiro, Joaquina; Garcia, Adriana; Paulo, Maria; Coutinho, Joana; Baptista, Teresa M.; Bernardino, Susana M.; Tecelão, Carla; Gil, Maria M.; Lemos, Marco F.L.Aurantiochytrium sp. is a heterotrophic microalgae that, in certain growth conditions, besides producing significant amounts of squalene, may potentially produce other bioactive compounds with high value and commercial interest such as docosahexaenoic acid (DHA), carotenoids, among others. Antioxidant compounds, naturally occurring in microalgae sources, have been identified as free radical or active oxygen scavengers, with beneficial effects on oxidation and oxidative damage inhibition in human body and foods. Several in vitro methodologies have been proposed to evaluate the antioxidant capacity, such as free radicals: 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2 -azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS+), and reactive species including hydrogen peroxide (H2O2), singlet oxygen (O2), and superoxide anion (O2-) (Balboa et al., 2013). The main objective of the present study was to evaluate the antioxidant activity of aqueous and ethanolic extracts of Aurantiochytrium sp., by means of different methodologies: reducing and scavenging activity by total phenolics content (TPC, Singleton & Rossi, 1965), DPPH (Custódio et al., 2012), and oxygen radical absorbance capacity (ORAC, Dávalos et al., 2004). The obtained results indicated that the microalgae Aurantiochytrium sp. is rich in antioxidant compounds, by reducing activity through total phenolics and DPPH, as well as scavenging activity by ORAC. Moreover, it is evident the higher selectivity of ethanol as solvent for the extraction of antioxidants compounds. The potential of this microalgae biomass as a good, natural and sustainable source for use as a functional food ingredient is evident, representing an opportunity with additional benefits due to the antioxidant role in diseases prevention and treatment.
- A pilot plant scale testing of the application of seaweed-based natural coating and modified atmosphere packaging for shelf-life extension of fresh-cut applePublication . Augusto, Ana; Miranda, Andreia; Costa, Leonor; Pinheiro, Joaquina; Campos, Maria J.; Raimundo, Délio; Pedrosa, Rui; Mitchell, Geoffrey; Niranjan, Keshavan; Silva, Susana F. J.Codium tomentosum hydroethanolic extract was obtained using a pilot solid–liquid extractor to validate the anti-browning functionality of the extract under industrial conditions. Fresh-cut apple slices were coated by immersion in: (1) a seaweed extract solution (0.5% w/v) and (2) a commercial coating, and the two sets of samples were compared with a control (immersion in water). Packaged samples were stored, under ambient and modified atmosphere conditions at 4°C. After 30 days of storage, the samples that were coated with the seaweed extract and packaged under modified atmosphere, demonstrated lower peroxidase activity and polyphenol oxidation when compared with the samples treated with the commercial additive. These results confirm, at pilot scale and under industrial production conditions, the efficacy of the seaweed extract as a bio-based substitute for the synthetic coatings, which are currently used to prevent browning in fresh-cut apples. Novelty impact statement Fresh-cut fruits are subjected to processing operations leading to a decrease in nutritional and organoleptic properties. It is therefore necessary to adopt strategies to delay the degradative processes. In this study, the efficacy of a pilot-scale production and industrial application of a coating formulated with Codium tomentosum seaweed extract has been established for the first time. This seaweed extract possesses the potential to prevent browning development in fresh-cut apples under industrial operating conditions.