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- 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.
- Valorization of Solanum melongena L. crop by-products: Phenolic composition and in vitro antioxidant, antidiabetic, anti-inflammatory, cytotoxic, and antimicrobial propertiesPublication . Añibarro-Ortega, Mikel; Dias, Maria Inês; Petrović, Jovana; Núñez, Sonia; Calhelha, Ricardo C.; Costa, Eduardo M.; Machado, M.; Pintado, Manuela; Soković, Marina; López, Víctor; Barros, Lillian; Pinela, JoséThis study explored the valorization of post-harvest eggplant aerial parts as a sustainable source of value-added ingredients by investigating their phenolic composition and in vitro bioactive properties. HPLC-DAD-ESI/MSn analysis identified chlorogenic acid derivatives as the predominant phenolic compounds (53 % of the phenolic fraction), followed by O-glycosylated kaempferol and quercetin. The extract displayed antioxidant activity in physiologically relevant cell-based assays and significant α-glucosidase inhibitory capacity that far exceeded that of the standard drug acarbose. It also inhibited the formation of advanced glycation end-products (AGEs), suggesting its potential to mitigate diabetes-related complications. Furthermore, the extract showed a modest pancreatic lipase inhibitory effect and capacity to suppress interleukin 6 production. Selective cytotoxicity against human gastric and colon adenocarcinoma cell lines and strong antimicrobial activity against foodborne pathogens were observed. Given the growing demand for natural alternatives to synthetic drugs, these findings position eggplant crop biomass as a promising, sustainable source of active compounds with potential applications in food, nutraceutical, and pharmaceutical formulations for managing type 2 diabetes and other oxidative stress-mediated conditions. This study not only contributes to the valorization of agricultural waste but also expands the research on by-products of Solanaceae crops, offering a pathway for sustainable resource utilization.
- The Effect of Cropping System and Irrigation Regime on the Plant Growth and Biochemical Profile of Cichorium spinosumPublication . Paschoalinotto, Beatriz H.; Polyzos, Nikolaos; Liava, Vasiliki; Mandim, Filipa; Pires, Tânia C.S.P.; Añibarro-Ortega, Mikel; Dias, Maria Inês; Barros, Lillian; Petropoulos, Spyridon A.This study evaluated the effects of three irrigation treatments (control (rain-fed plants), deficit irrigation (DI: 50% of maximum field capacity), full irrigation (FI: 100% of maximum field capacity)), and two crop-management treatments (with or without crop rotation with bean, CR, and NCR, respectively) on the plant growth and chemical composition of C. spinosum. The results indicated that deficit irrigation combined with crop rotation increased the weight of leaves per plant, followed by rain-fed plants for the same crop-management treatment. Additionally, these two factors significantly influenced the nutritional profile, free sugars, and organic acid content in a variable manner. Moreover, the control treatment and deficit irrigation increased the content of K, Na, and Mg, which are highly mobile nutrients, whereas the levels of moderately mobile nutrients such as Fe, Mn, Cu, and Zn decreased. Deficit irrigation without crop rotation significantly increased the content of total tocopherols, followed by deficit irrigation with crop rotation and full irrigation without crop rotation. The main fatty acids were α-linolenic acid (C18:3n3), followed by palmitic acid (C16:0) and linoleic acid (C18:2n6), while the control and deficit irrigation treatments combined with crop rotation increased PUFA and decreased SFA content. Furthermore, deficit irrigation and crop rotation induced the accumulation of phenolic compounds, flavonoids, and phenolic acids, especially the content of the major compounds (e.g., chicoric acid, quercetin-O-hexurunoside, and luteolin-O-hexurunoside). The leaf extracts exhibited varied antioxidant activity (assessed by TBARS and OxHLIA assays), and antimicrobial activity. On the other hand, no antifungal, antiproliferative (except for AGS cell line), hepatotoxic, or anti-inflammatory effects were recorded. In conclusion, the combination of deficit irrigation and crop rotation with bean positively affected the quality traits and the fresh weight of leaves, thus suggesting that such eco-friendly practices could have beneficial effects in the cultivation of C. spinosum plants within the context of climate-change mitigation strategies.
- Plantas de pimento dulce (Capsicum annuum L.) em poscesacha como fuente de ingredientes antioxidantes y antidiabéticosPublication . Añibarro-Ortega, Mikel; López, Víctor; Dias, Maria Inês; Mandim, Filipa; Barros, Lillian; Pinela, JoséEl estrés oxidativo desempeña un papel fundamental en el desarrollo de enfermedades crónicas y metabólicas como la diabetes, enfermedades cardiovasculares y neurodegenerativas, obesidad y cáncer. Esto se debe a la producción excesiva de especies reactivas de oxígeno que dañan los lípidos, proteínas y ADN de los organismos, comprometiendo procesos celulares vitales [1]. Los antioxidantes naturales, como los polifenoles, pueden prevenir estos daños y hoy en día existe un creciente interés por el uso de extractos ricos en estos compuestos como ingredientes funcionales naturales. En el caso del cultivo de pimiento dulce (Capsicum annuum L.), tras la cosecha se genera una grande cantidad de biomasa vegetal infravalorada, que podría contener metabolitos secundarios de alto valor comercial. Así, el objetivo de este estudio fue valorizar estos subproductos agrícolas en línea con los objetivos de la Agenda 2030 de Desarrollo Sostenible de las Naciones Unidas. Para ello, los subproductos de la planta fueron empleados para preparar extractos hidroetanólicos [2], los cuales se caracterizaron con respecto a su perfil de compuestos fenólicos mediante HPLC-DAD-ESI/MSn y sus propiedades antioxidante, antidiabética, anti-obesidad, antiinflamatoria y citotóxica con base en ensayos celulares y enzimáticos in vitro. El análisis cromatográfico reveló un perfil fenólico con predominio de ácidos fenólicos, como el ácido clorogénico, y flavonas O-glicosiladas, concretamente luteolina y apigenina. Aunque los extractos presentaron cierta actividad en todos los ensayos realizados, donde más se destacaron fue en la actividad antioxidante, antidiabética y citotóxica en ciertas líneas celulares tumorales. Como conclusión, se puede afirmar que los subproductos de plantas de pimiento dulce son una prometedora fuente de compuestos bioactivos de alto valor añadido. Estos pueden tener propiedades beneficiosas para la salud del consumidor al ser incorporados en productos alimentarios. Además, fomentar esta práctica de circularidad será importante para la producción sostenible de ingredientes alimentarios naturales.
- Thymus species from romanian spontaneous flora as promising source of phenolic secondary metabolites with health-related benefitsPublication . Babotă, Mihai; Frumuzachi, Oleg; Mafra, Isabel; Nicolescu, Alexandru; Dias, Maria Inês; Pinela, José; Barros, Lillian; Añibarro-Ortega, Mikel; Stojković, Dejan; Carevic, Tamara; Mocan, Andrei; López, Víctor; Crisan, GianinaWild thyme aerial parts (Serpylli herba) are recognized as a valuable herbal product with antioxidant, anti-inflammatory, and antibacterial effects. Although pharmacopoeial regulations allow its collection exclusively from Thymus serpyllum, substitution with other species is frequent in current practice. This study analyzed the phenolic composition, antioxidant, and enzyme-inhibitory and antimicrobial activity of the hydroethanolic extracts obtained from five Romanian wild thyme species (Thymus alpestris, T. glabrescens, T. panonicus, T. pulcherimus and T. pulegioides). The analysis of individual phenolic constituents was performed through LC-ESI-DAD/MS2, while for the in vitro evaluation of antioxidant potential, TEAC, FRAP, DPPH, TBARS and OxHLIA assays were employed. The anti-enzymatic potential was tested in vitro against tyrosinase, alpha-glucosidase and acetylcholinesterase. High rosmarinic acid contents were quantified in all species (20.06 +/- 0.32-80.49 +/- 0.001 mg/g dry extract); phenolic acids derivatives (including salvianolic acids) were confirmed as the principal metabolites of T. alpestris and T. glabrescens, while eriodictyol-O-di-hexoside was found exclusively in T. alpestris. All species showed strong antioxidant potential and moderate anti-enzymatic effect against alpha-glucosidase and acetylcholinesterase, showing no anti-tyrosinase activity. This is the first detailed report on the chemical and biological profile of T. alpestris collected from Romanian spontaneous flora.
- Flowers of allium cepa l. as nutraceuticals: phenolic composition and anti-obesity and antioxidant effects in caenorhabditis elegansPublication . Moliner, Cristina; Núñez, Sonia; Casedas, Guillermo; Valero, Marta Sofia; Dias, Maria Inês; Barros, Lillian; López, Víctor; Gomez-Rincon, CarlotaAllium cepa L., commonly known as onion, is one of the most-consumed vegetables. The benefits of the intake of its bulb are well studied and are related to its high polyphenol content. The flowers of onions are also edible; however, there are no studies about their biological properties. Our aim was to determine the polyphenolic profile and assess the antioxidant and anti-obesity capacity of an ethanolic extract from fresh flowers of A. cepa. The phenolic constituents were identified through LC-DAD-ESI/MSn. For the anti-obesity potential, the inhibitory activity against digestive enzymes was measured. Several in vitro assays were carried out to determine the antioxidant capacity. A Caenorhabditis elegans model was used to evaluate the effect of the extract on stress resistance and fat accumulation. For the first time, kaempferol and isorhamnetin glucosides were identified in the flowers. The extract reduced fat accumulation in the nematode and had a high lipase and alpha- glucosidase inhibitory activity. Regarding the antioxidant activity, the extract increased the survival rate of C. elegans exposed to lethal oxidative stress. Moreover, the activities of superoxide dismutase and catalase were enhanced by the extract. Our results demonstrate, for the first time, the antioxidant and anti-obesity activity of onion flowers and their potential use as functional foods and nutraceuticals.
- Mycorrhization and micropropation of chestnut (Castanea sativa Mill.) seedlings as tools to obtain high added-value phenolic compoundsPublication . Dias, Maria Inês; Pinela, José; Pereira, Carla; Ferreira, Patrícia; Oliveira, Maria de Fátima; Martins, Anabela; Afonso, Andreia; Barros, LillianMiChestnut3 is a project of the company DEIFIL whose main objective is to produce more resistant and productive hybrid chestnut (Castanea sativa Mill.) seedlings through micropropagation and mycorrhization techniques. In this work, in addition to the agronomic traits of the micropropagated mycorrhizal chestnut seedlings, it was also important to evaluate the changes induced by mycorrhization in the phenolic profile of these plants. Phenolic compounds are plant secondary metabolites involved in plant-microbe interactions/symbiosis and act as signaling molecules in the establishment of arbuscular mycorrhizal symbioses, as well as in plant defense mechanisms [1]. According to the literature, considerable increases in phenolic compounds in host plants as a result of arbuscular mycorrhizal fungus inoculation have been reported during the progression of the infection [1]. Therefore, this work was carried out to study the impact of the type of fungal inoculum and the period of mycorrhization (before or after potting) on the qualitative and quantitative profile of phenolic compounds of the roots and leaves of the chestnut seedlings produced by DEIFIL. After collection and lyophilization of the plant material, hydroethanolic extracts were prepared and the phenolic compounds were characterized by HPLC-DAD-ESI/MS [2]. Ellagic acid derivatives and Oglycosylated flavonoids were the major phenolic compounds in both plant roots and leaves, which agreed with previous reports [2,3]. A statistical analysis showed that the type of inoculum and period of mycorrhization significantly (p<0.05) affected the phenolic profile of the chestnut hybrids. In general, the mycorrhizal seedlings with the fungi Amanita caesarea and Boletus edulis were those that presented the highest levels of phenolic compounds. Relationships between the levels of these signaling compounds and the agronomic performance of chestnut seedlings were also found. The results bring new perspectives into the future production of a hybrid chestnut tree resistant to ink disease in the main traditional Portuguese varieties
- Chemical characterization, antioxidant and antimicrobial activities of winemaking industry by-productsPublication . Duarte, Cristina Nogueira; Dias, Maria Inês; Heleno, Sandrina A.; Santos-Buelga, Celestino; Dias, Rolando; Barros, Lillian; Amaral, Joana S.The exploitation of bioactive compounds trom agri-food by-products has been attracting an increasing interest from different industries within a circular economy context since several of compounds may have health properties or be used as natural colorants or preservatives. So far, several studies have been developed on the characterization of red wine grape pomace and its components (seeds, skins, and stems). However, less attention has been paid to other by-products, such as grape pomace from white wine, the residues after distillation, wine lees, and diatomaceous earth, the latter used in the filtration of wine and corresponding to about 250 tons/year of waste in Portugal. In this work, samples of these by-products were evaluated for their chemical composition by HPLC-DAD-ESI-Msn and bioactivities (antioxidant activity by TBARS, DPPH, and reducing power assays, and antimicrobial activity against eight bacteria and two fungi). Fifteen non-anthocyanin phenolic compounds were found, including 5 phenolic acids, 4 flavan-3-0Is, 2 0- glycosylated flavanols, 3 flavanol aglycones, and one unknown. In the red wine byproducts 12 anthocyanins were detected, most being malvidin derivatives. Wine lees and white grape pomace before distillation presented the highest contents of phenolic compounds. Ali samples showed antibacterial and antifunga! activity against most of the tested microorganisms, with red and white grape pomace and diatomaceous earth having the best bacteriostatic activity, while the lees stood out against fungi. Ali samples showed promising antioxidant capacity, with very good results obtained on TBARS for the white pomace after distillation and diatomaceous earth. Overall, the results show that besides grape pomace other wine industry byproducts are also good sources of bioactive compounds with high potential for exploitation.
- 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.
- Blueberry juice as a nutritious and bioactive beverage to be included in novel food productsPublication . Backes, Emanueli; Saldanha, Ana Luísa; Molina, Adriana K.; Pereira, Carla; Dias, Maria Inês; Ferreira, Isabel C.F.R.; Barros, LillianBlueberry ( Vaccinium myrtillus L.) is a very popular fruit, native from the northern hemisphere and consumed worldwide. It has been widely studied for being a rich source of bioactive compounds with recognized beneficial properties for Human health [1]. Therefore, several industrialized products, such as juices and derivatives, have been developed from blueberry fruit, aiming at most practical forms of consumption. In this sense, the present work aimed to analyse the blueberry juice in terms of nutritional value, following AOAC procedures, and chemical compositions, namely in free sugars, by HPLC-RI, organic acids, by UFLC-PDA, and phenolic compounds, by HPLC-DAD/ESI-MS. Moreover, the antioxidant capacity, by TBARS and OxHLIA assays, was also assessed to validate its bioactive properties. Regarding the nutritional value, carbohydrates were the major macronutrients found in blueberry juice, followed by ash and protein. In terms of free sugars, fructose and glucose were detected, with a higher concentration of fructose. Four organic acids were also identified in the juice, namely oxalic, quinic, malic, and shikimic acids, being quinic acid the most abundant one. In what concerns the phenolic composition, four phenolic acids (quinic acid, caffeic acid hexoside, cis 5- O- caffeoylquinic acid, and trans 5- O -caffeoylquinic acid) and six anthocyanins (cyanidin-3- O -glucoside, cyanidin-3- O -pentoside, peonidin-3- O glucoside, peonidin-3- O -pentoside, malvidin-3- O -glucoside, and malvidin-3- O -pentoside) were detected. As expected, considering its chemical composition, the juice also presented a strong antioxidant capacity, being able of inhibiting the lipid peroxidation and the oxidative hemolysis. The results obtained in the present study validate the nutritional and bioactive quality of the juice obtained from Vaccinium myrtillus L., justifying its application in the development of novel foodstuff.