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- Nutritional and bioactive oils from salmon (Salmo salar) side streams obtained by Soxhlet and optimized microwave-assisted extractionPublication . Pinela, José; Mandim, Filipa; Heleno, Sandrina A.; Ferreira, Isabel C.F.R.; Barba, Francisco Jose; Berrada, Houda; Caleja, Cristina; Barros, Lillian; Fuente, Beatriz de laThe efficiency of the microwave-assisted extraction (MAE) technique on recovering nutritional and bioactive oils from salmon (Salmo salar) side streams was evaluated and compared to Soxhlet extraction. The response surface methodology (RSM) coupled with a central composite rotatable design was used to optimize time, microwave power, and solid/liquid ratio of the MAE process in terms of oil yield. The optimal MAE conditions were 14.6 min, 291.9 W, 80.1 g/L for backbones, 10.8 min, 50.0 W, 80.0 g/L for heads, and 14.3 min, 960.6 W, 99.5 g/L for viscera, which resulted in a recovery of 69% of the total lipid content for backbones and heads and 92% for viscera. The oils obtained under optimal MAE conditions showed a healthy lipid profile as well as cytotoxic, antioxidant, anti-inflammatory, or antimicrobial properties. These results highlight that oils from underutilized salmon by-products could be exploited by different industrial sectors under the circular economy approach.
- Upcycling fish by-products into bioactive fish oil: the suitability of microwave-assisted extractionPublication . Pinela, José; Rodrigues, Matilde; Pires, Tânia C.S.P.; Mandim, Filipa; Almeida, André; Dias, Maria Inês; Caleja, Cristina; Barros, LillianThe seafood industry is often left out of the food waste discussion, but this sector is no exception, as it generates large amounts of various by-products. This study aimed to explore the potential of the microwave-assisted extraction (MAE) technique to obtain high-quality oil from fish by-products. The independent variables, which were time (1-30 min), microwave power (50-1000 W), and solid/liquid ratio (70-120 g/L) were combined in a 20-run experimental design coupled with the response surface methodology (RSM) for process optimization. The obtained oil yield values were fitted to a quadratic equation to build the theoretical models, which were statistically validated based on statistical criteria and used to predict the optimal MAE condition. The oil yields were significantly affected by the three independent variables through linear, quadratic, and/or interactive effects. Compared to a conventional Soxhlet extraction (SE), the optimal MAE conditions allowed between 60 and 100% of oil to be recovered in less than 19 min and with less solvent consumption. The fatty acid profiles of the oils obtained through SE and optimized MAE were characterized by gas chromatography with flame ionizing detection (GC-FID) after a derivatization process. These oils were constituted mainly of health, beneficial unsaturated fatty acids, such as oleic, docosahexaenoic (DHA), linoleic, and eicosapentaenoic (EPA) acids, which were not affected (p > 0.05) by the extraction methods. Interestingly, the oils obtained through MAE showed the best microbial growth inhibition results may have been due to thermolabile compounds, preserved via this unconventional non-thermal method. The oils also exhibited anti-inflammatory effects via nitric oxide production inhibition and cytotoxic potential especially, against breast and gastric adenocarcinoma cells. However, the threshold of toxicity should be further investigated. Overall, this work emerges as a future-oriented approach to upcycling fish by-products into high-quality oils that can be used in the formulation of pet food and other products.
- Sea bass (Dicentrarchus labrax) and sea bream (Sparus aurata) head oils recovered by microwave-assisted extraction: nutritional quality and biological propertiesPublication . Fuente, Beatriz de la; Pinela, José; Calhelha, Ricardo C.; Heleno, Sandrina A.; Ferreira, Isabel C.F.R.; Barba, Francisco Jose; Berrada, Houda; Caleja, Cristina; Barros, LillianMicrowave-assisted extraction (MAE) and Soxhlet extraction (SE) were used to obtain oil from European sea bass (Dicentrarchus labrax) and gilthead sea bream (Sparus aurata) heads. The MAE technique allowed the recovery of more than 50% of the total lipid con- tent for both fish by-products in less than 11 min extraction. Based on their fatty acid composition, all fish head oils presented a healthy lipid profile and were found to be a good source of docosahexaenoic acid (DHA, 11–14%). Different lipid quality indices also revealed their cardiovascular protective potential. Oils obtained by MAE showed higher antibacterial and antifungal effects against food pathogens than those extracted by SE. Cellular antioxidant activity (29–35% oxidation inhibition) and anti-inflammatory poten- tial via NO production inhibition (IC50 = 14–21 μg/mL) were evaluated using murine macrophages cells (RAW 264.7). The highest cytotoxic effect (GI50 = 38–46 μg/mL) of fish head oils was observed against breast cancer cells (MCF-7). These results showed that sea bass and gilthead sea bream heads could be exploited for the production of oil with nu- tritional and bioactive properties in line with circular bioeconomy concepts.
- A Comprehensive Review of Fish Protein Hydrolysates Targeting Pet Food FormulationsPublication . Ribeiro, Tânia Isabel Bragança; Maia, Margarida R.G.; Fonseca, António J.M.; Marques, Bianca; Caleja, Cristina; Rosa, Ana; Martins, Rui; Almeida, André; Mota, Maria J.; Aires, Tiago; Rocha, Cristina M.R.; Teixeira, José António; Cabrita, Ana R.J.; Barros, Lillian; Pintado, ManuelaThe fish industry generates significant amounts of fish by- and co-products (FBCPs) annually, projected to reach 160.8 million tonnes by 2030. This growth highlights the urgent need for sustainable FBCP management and an opportunity to improve the sector’s environmental sustainability. Fish protein hydrolysates (FBCPHs) and bioactive peptides (BPs) derived from these FBCPs are gaining recognition in the pet food sector for their nutritional and bioactives benefits. FBCPHs, primarily sourced from category 3 by-products unsuitable for human consumption, could significantly enhance the economic viability of both industries. This review analyzes production processes, highlighting the benefits and challenges of enzymatic hydrolysis and reviewed emerging technologies such as subcritical water hydrolysis (SWH), which are promising sustainable alternatives by enhancing extraction efficiency and reducing energy consumption. The review explores FBCPHs’ applications in pet food, focusing on beneficial biological activities (e.g. antioxidant, prebiotic, neuroprotective). Findings show FBCPHs have significant potential in pet food formulations, providing palatability, hypoallergenic benefits, and addressing health concerns like gastrointestinal disorders and stress-related behaviors. However, further research is required to optimize production processes, scale industrial application, and ensure regulatory compliance. In conclusion, FBCPHs present a valuable solution for promoting sustainability, improving pet nutrition, and supporting the circular economy.
- Red seaweeds as a source of nutrients and bioactive compounds: Optimization of the extractionPublication . Carpena Rodríguez, María; Caleja, Cristina; Pereira, Eliana; Pereira, Carla; Ćirić, Ana; Soković, Marina; Soria-Lopez, Anton; Fraga-Corral, Maria; Simal-Gandara, Jesus; Ferreira, Isabel C.F.R.; Barros, Lillian; Prieto Lage, Miguel A.The present work aimed to determine the nutritional composition (ash, protein, fat, carbohydrate content and energy value), phenolic compounds, pigments and organic acids content of three typical red algae from the Northwest of Spain: Chondrus crispus, Mastocarpus stellatus, and Gigartina pistillata; as well as their antioxidant and antimicrobial activities. Furthermore, the present work compared two extraction techniques: conventional heat assisted extraction (HAE) and high pressure assisted extraction (HPAE) to maximize the yield and the concentration of target compounds. Different independent variables were considered for the response study. Time (t) and percentage of ethanol of the solvent (S) were chosen for both techniques and temperature (T) and pressure (P) were used for HAE and HPAE, respectively. The experiments were designed following a response surface methodology (RSM) approach. The obtained results showed a similar nutritional composition between algae samples: low-fat content and high content of proteins, carbohydrates and energy. All tested algae showed good antioxidant and antimicrobial properties. Finally, HEA demonstrated to be the most efficient extraction technique. This study confirms the potential of red algae to be part of the human diet as a source of non-animal protein, due to its nutritional content, phenolic profile, pigments concentration and bioactive properties, which proves that HAE is the optimum technique for the extraction maximization.
- Matricaría recutita L. (chamomile) decoction as a source of phenolic compoundsPublication . Caleja, Cristina; Dias, Maria Inês; Barros, Lillian; Oliveira, Beatriz; Santos-Buelga, Celestino; Ferreira, Isabel C.F.R.Aromatic herbs have been used to prepare several infüsions and decoctions with claimed beneficiai health effects, many of them attributed to the presence of phenolic compounds (hydrophilic molecules highly abundant in those aqueous preparations) (Guimarães et al., 2013). In fact antioxidant, antimicrobial and antitumor properties, among others, of phenolic compounds have been extensively reported (Kahkonen et al., 1999; Rauha et al., 2000). In the present work, Matricaria recutita L. (chamomile), one of the species mostly used to prepare herbal drinks was characterized in terms of phenolic profile. Américo Duarte Paixão Lda. (ADP) provided the samples, which were fúrther extracted by decoction, lyophilized, re-dissolved in water (5 mg/mL) and analysed by HPLC-DADESI/ MS. Nineteen phenolic compounds were identified, among which were nine flavonoids and ten phenolic acids. Phenolic acids were the major group present in M. recutita decoctions (23.66 ± 0.27 mg/g lyophilized decoction), in which four compounds were tentatively assigned as caffeoyl-2, 7-anhydro-3-deoxy-2-octulopyranosonic acids (CDOA) derivatives, being a di-CDOA (6. 83 ± 0.05 mg/g) the main phenolic acid. Among the flavonoids identified (17.89 ± 0.91 mg/g), luteolin-0-glucuronide (4.80 ± 0.54 mg/g) was the major compound. To the best ofour knowledge, such type ofcompounds hás not been previously reported in M. recutita flowers. Nevertheless, luteolin glycosides are commonly found in relevant amounts in this matrix and in its iníüsions. Due to the wellestablished bioactive properties ofphenolic compounds, M. recutita flowers could be used in arder to obtain extracts with health-promoting properties or be incorporated into foods as natural preservatives.
- The compositional aspects of edible flowers as an emerging horticultural productPublication . Pires Junior, Eleomar de O.; Di Gioia, Francesco; Rouphael, Youssef; Ferreira, Isabel C.F.R.; Caleja, Cristina; Barros, Lillian; Petropoulos, Spyridon Α.Edible flowers are becoming very popular, as consumers are seeking healthier and more attractive food products that can improve their diet aesthetics and diversify their dietary sources of micronutrients. The great variety of flowers that can be eaten is also associated with high variability in chemical composition, especially in bioactive compounds content that may significantly contribute to human health. The advanced analytical techniques allowed us to reveal the chemical composition of edible flowers and identify new compounds and effects that were not known until recently. Considering the numerous species of edible flowers, the present review aims to categorize the various species depending on their chemical composition and also to present the main groups of compounds that are usually present in the species that are most commonly used for culinary purposes. Moreover, special attention is given to those species that contain potentially toxic or poisonous compounds as their integration in human diets should be carefully considered. In conclusion, the present review provides useful information regarding the chemical composition and the main groups of chemical compounds that are present in the flowers of the most common species.
- Extraction optimization of bioactive compounds from Thymus ulgaris L.: comparison of heat-assisted and ultrasound-assisted extractionsPublication . Caleja, Cristina; Pereira, Eliana; Sprea, Rafael M.; Finimundy, Tiane C.; Carocho, Márcio; Amaral, Joana S.; Prieto Lage, Miguel A.; Barros, LillianPlants have been used since ancient times due to their biological action associated mainly with several phytochemicals, including phenolic compounds1. Thymus vulgaris L. is a species described as a potential source of natural molecules with applications in the food industry, but also in the cosmetic and pharmaceutical industry due to its antibacterial, antifungal, and anti-inflammatory effects2. In this sense, the present study aimed to compare extraction systems in terms of phenolic compounds from T. vulgaris through the heat-assisted (HAE) and ultrasound-assisted extraction (UAE) techniques, aiming to improve its potential for industrial applications. For this propose, an extraction optimization was studied using three independent variables, time (t, minutes), solvent (S, % of ethanol), and temperature (T, ºC) or power (P, W), in a three-level Box-Behnken design, analyzing the surface methodology (RSM). The extraction yield and the content in phenolic compounds identified by HPLC-DAD-ESI/MS were the experimental responses applied. The polynomial models were successfully fitted to the experimental data and used to determine the optimal HAE and UAE conditions. Nineteen phenolic compounds were identified, nine of which were phenolic acids (mainly caffeic acid derivatives) and ten flavonoids: flavan-3-ols (catechin derivatives), flavones (quercetin derivatives), flavonols (apigenin and luteolin derivatives) and flavanones (eriodictyol derivatives). The conditions obtained for the optimal extraction of phenolic compounds, that allowed the experimental validation of the predictive model are 3% ethanol (v/v) for 89 minutes at a temperature of 98º C for HAE and 77% ethanol (v/v) for 17 minutes at 207 W for UAE. Extraction optimization is important from an industrial point of view, as it allows obtaining extracts with a high content of bioactive compounds. Considering that the phenolic composition appears in literature as a direct influence on bioactive properties, the optimization processes will allow a better exploitation of extracts, ensuring the greatest cost reduction for the industry.
- Caracterização dos resíduos de Brassica oleracea L. para obtenção de um potencial ingrediente naturalPublication . Pepinelli, Ana Luísa; Oliveira, Tatiane C.G.; Junior, Eleomar Pires; Caleja, Cristina; Dias, Maria Inês; Kostić, Marina; Soković, Marina; Calhelha, Ricardo C.; Mandim, Filipa; Gonçalves, Odinei Hess; Oliveira, Beatriz; Pereira, Eliana; Barros, LillianA comunidade cientifica, em parceria com a indústria alimentar, tem vindo a estudar diferentes matrizes naturais, a fim de obter ingredientes funcionais com potencial aplicação neste setor industrial. Nesse sentido, este trabalho pretendeu fazer a caracterização de Brassica oleracea var. Acephala (popularmente conhecida como couve galega) quanto ao seu perfil fenólico e ação biológica, com o objetivo de identificar compostos fenólicos com potencial conservante e/ou bioativo para incorporação na indústria alimentar. O perfil fenólico individual foi obtido através de um sistema de HPLC-DAD/ESI-MSn e o potencial bioativo foi avaliado através de ensaios in vitro. A atividade citotóxica foi avaliada em linhas celulares tumorais humanas (AGS, CaCo2, MCF-7, NCI-H460) e a toxicidade em uma linha celular não tumoral (VERO), através do método colorimétrico da sulforodamina B. A atividade anti-inflamatória foi estudada em macrófagos de rato (RAW 264.7); a atividade antimicrobiana foi testada usando o método de microdiluição; e a atividade antioxidante foi avaliada através dos ensaios de TBARS (inibição da peroxidação lipidica) e CAA (atividade antioxidante celular). O perfil fenólico dos bioresíduos de couve galega revelou a presença de moléculas com elevado interesse em concentrações promissoras. Em relação ao potencial bioativo da amostra, foi possível observar uma interessante atividade antioxidante pela análise de TBARS, onde na concentração de 19 μg/mL foi possível oferecer 50% da atividade, porém pela análise de CAA os resultados foram moderados, com uma concentração de 2000 μg/mL com uma inibição de 40%. A atividade antimicrobiana revelou resultados muito satisfatórios, destacando-se as bactérias Bacillus cereus e Enterobacter cloacae e os fungos Aspergillus fumigatus e Penicillium fumiculosum como as estirpes mais suscetíveis ao efeito do extrato da couve. Em relação à atividade citotóxica e antiinflamatória, o extrato da couve não apresentou capacidade antiproliferativa na concentração máxima testada (400 μg/mL), revelando a ausência de potencial antitumoral e anti-inflamatório, mas também, provando a ausência de toxicidade do extrato. Considerando esses resultados, os bioresíduos da couve apresentam uma composição rica em compostos de elevado interesse, nomeadamente com potencial antioxidante, o que pode ser interessante para o desenvolvimento de ingredientes funcionais com possíveis aplicação na indústria. Para além disso, a reutilização de subprodutos permitirá não só diminuir o desperdício alimentar, como também criar soluções estratégicas no setor agroindustrial de crescimento económico e sustentabilidade.
- Caracterização do perfil fenólico do extrato aquoso e hidroetanólico de Rosmarinus officinalis L.Publication . Ribeiro, Andreia; Caleja, Cristina; Barros, Lillian; Santos-Buelga, Celestino; Barreiro, M.F.; Ferreira, Isabel C.F.R.Desde a antiguidade, as plantas têm suscitado grande interesse pelos benefícios que trazem para a saúde humana. Em particular, algumas espécies de plantas, para além das suas características aromáticas, constituem uma fonte natural de diversos compostos bioativos nomeadamente compostos fenólicos, que têm sido associados a propriedades antioxidantes, anti -inflamatórias, entre outras. Neste trabalho, estudaram -se amostras de Rosmarinus officinalis L. fornecidas pelo Cantinho das Aromáticas Lda. (Vila Nova de Gaia). Prepararam -se duas tipologias de extratos, aquoso e hidroetanólico, que foram subsequentemente caracterizados por HPLC -DAD -ESI/MS. Relativamente ao extrato hidroetanólico, o extrato aquoso apresentou um teor de compostos fenólicos total superior, assim como concentrações dos compostos individuais superiores. O ácido rosmarínico foi o composto mais abundante em ambos os extratos.