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- Insights into the phenolic composition and in vitro bioactivity of cardoon capitulum: a nutraceutical-oriented valorization studyPublication . Mandim, Filipa; Pinela, José; Marcelino, Sandra Andreia; Dias, Maria Inês; Barracosa, Paulo; Ivanov, Marija; Soković, Marina; Santos-Buelga, Celestino; Barros, LillianThe capitulum constituents (stigma, corolla, bracts, pappus, and receptacle) of seven cardoon cultivars were studied for their polyphenolic composition and bioactive properties. Fifteen phenolic compounds were identified, secoiridoid, secoxyloganin, and apigenin-O-hexuronoside in higher concentrations. The bracts had the highest concentration of phenolic compounds, and the corolla had the greatest variety. The corolla of F4-1-4 and F1-34-1 cultivars, and bracts of F1-1-1, F1-19-4, and F4-37-1 cultivars revealed the greatest capacity to inhibit the thiobarbituric acid reactive substances formation (IC50 of 38 and 40.4 μg/mL, respectively). The corolla of F1-34-1 cultivar showed higher antihaemolytic activity than the positive control Trolox (IC50 5.5 vs. 20 μg/mL). Bracts of the F4-25-2 cultivar showed higher cytotoxic activity. No hepatotoxicity or anti-inflammatory effects were presented by the studied samples. These bioactivities exhibit a significative correlation with phenolic compounds, in particular flavonoids. Antimicrobial capacity was also observed, with all samples presenting higher antifungal potential than positive controls.
- Genetic information influence on phenolic composition and bioactivities of Ceratonia siliqua L. seedsPublication . Marcelino, Sandra Andreia; Mandim, Filipa; Oludemi, Taofiq; Dias, Maria Inês; Barracosa, Paulo; Prieto Lage, Miguel A.; Barros, LillianCeratonia siliqua L., commonly known as the carob tree is native to the Mediterranean countries and is widely known and consumed. Due to its chemical constituents, flavoring properties, and nutritional benefits, it has the potential to be of significant importance to the food industry1. The carob bean is made up of 90% pulp and 10% seeds. Those seeds are widely used in the food industry as a thickening agent (E-410). However, knowledge about phenolic composition and its relation to biological properties is scarce. This study used seeds of thirteen carob varieties harvested in Algarve, Portugal. The phenolic composition of the hydroethanolic extracts was analyzed by HPLC-DAD-ESI/MS. The antioxidant, anti-inflammatory, cytotoxic and antibacterial properties of the extracts were also analysed. The phenolic composition was analyzed by HPLC-DAD-ESI/MS. Cytotoxic activity was evaluated by the colourimetric method of sulforhodamine B. Anti-inflammatory activity was determined by inhibition of NO production in murine macrophages. The antibacterial activity was evaluated through the method of successive microdilutions and the antioxidant activity through the TBARS and CAA assays. Seventeen phenolics compounds were tentatively identified, being (Epi)catechin dimer type dimer and apigenin-O-hexosyl-pentoside the most abundant ones (3.08 – 11.67 mg/mL). All the varieties studied exhibited the capacity to inhibit TBARS formation. The extract obtained from the Gasparinha variety was the only one that inhibited the reactive oxygen species formation in the CAA assay. For the cytotoxic activity only Cavi, Cardeira and Pé Comprido varieties demonstrated the ability to inhibit the proliferation of the tumor cell lines tested, without showing a hepatotoxic effect. All extracts presented a broad-spectrum microbial growth inhibition without an efficient bactericidal power. These findings highlight Carob seed as a rich source of structurally diverse biomolecules with potential application as additives in food formulation development. However, further studies are needed to understand the correlation between phenolic compounds and the bioactive properties associated with carob seed tissues.
- Occlusal changes with clear aligners and the case complexity influence: a longitudinal cohort clinical studyPublication . Marcelino, Vanessa; Baptista, Sofia; Marcelino, Sandra Andreia; Paco, Maria; Rocha, Duarte; Gonçalves, Maria dos Prazeres; Azevedo, Rui; Guimarães, Antonio Sergio; Fernandes, Gustavo Vicentis Oliveira; Pinho, TeresaBackground: Clear aligners (CA) are used 22 h daily, creating a bite-block effect. This work aims to (i) analyze occlusal changes before the beginning of treatment, after the first set of CA and after the use of additional aligners; (ii) compare planned occlusal contacts with the ones obtained after the first set of CA; (iii) analyze the occlusal changes occurred after reaching the orthodontic goals after 3 months of using CA only at night; (iv) evaluate and characterize which tooth movements did not allow the treatment to be completed at the end of the first set of aligners, and finally (v) verify the possible relation between the changes in occlusal contact and areas and parameters such as case complexity and facial biotype. Materials and Methods: A quantitative, comparative, and observational longitudinal cohort study design was implemented to evaluate the clinical data and the complexity levels of cases receiving CA. A non-probabilistic and convenience sample of 82 individuals was recruited. The orthodontic malocclusion traits were classified as simple, moderate, or complex corrections based on the basis of the Align((R)) recommendations with the Invisalign((R)) evaluation tool. According to the Invisalign((R)) criteria, patients need only one complex problem for their case to be classified as complex. Meshlab((R)) v. 2022.02, ClinCheck((R)) version Pro 6.0, My-Itero((R)) version 2.7.9.601 5d plus, and IBM (R) SPSS Statistics software (Statistical Program for Social Sciences), version 27.0 for Windows were the software((R)) used. Results: A statistically significant decrease in area and occlusal contacts number were observed from before the start of orthodontic treatment (T0) to the end of treatment (T1). The changes in the occlusal area (from T0 to T1) were statistically different between hyperdivergent (28.24 [15.51-40.91]) and hypodivergent (16.23 [8.11-24.97]) biotypes (p = 0.031). A significant difference between the hyperdivergent (4.0 [2.0-5.0]) and normodivergent (5.5 [4.0-8.0]) group was found in T1 for the anterior contacts (p = 0.044). Anterior contacts obtained were significantly higher than the planned (p = 0.037) Between T1 and T2 statistically significant increases of occlusal areas, posterior and total contacts were observed. Conclusions: Occlusal contact and area were decreased, either at the end of the first set or after the use of additional aligners. Anterior occlusal contacts obtained were higher than planned as opposed to posterior occlusal contacts obtained. The hardest tooth movements to achieve to complete the treatment were distalization, rotation, and posterior extrusion. After completing orthodontic treatment (T1) to 3 months after (T2) using additional aligners only at night, posterior occlusal contacts were significantly increased, which could be due to the natural settling of the teeth in this period.
- Masticatory function in individuals with temporomandibular disorders: a systematic review and meta-analysisPublication . Marcelino, Vanessa; Rovere, Solene de; Paco, Maria; Gonçalves, Maria dos Prazeres; Marcelino, Sandra Andreia; Guimarães, Antonio Sergio; Pinho, TeresaThe literature search was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) protocol in the PubMed, Cochrane Library, LILACS, EBSCO, Scielo, between 2012 and 2022. The methodological quality was assessed by using the Newcastle-Ottawa Study Quality Assessment Scale. Mean differences and 95% confidence intervals were calculated and combined in meta-analyses. A total of 1202 participants were included in this systematic review (690 with TMD; 512 without TMD), with 22 articles being included in the qualitative analysis. Only three studies enabled the comparative analysis of the results. Ten articles showed a high methodological quality and a low risk of bias, and twelve had a low methodological quality and an increased risk of bias. The meta-analysis showed that the differences between the intervention and control groups were not statistically significant for the percentage overlapping coefficient of the anterior temporal muscle, for the masseter, and for the torque coefficient. The parameters analyzed with the compound technique for chewing showed altered mandibular functions in individuals with TMD. With the EMG method, it was possible to suggest that TMD in adult individuals causes compensatory muscle behaviors, and several changes in the masticatory function were found.
- How do extraction methodologies influence the biological properties of pomegranate leaves?Publication . Marcelino, Sandra Andreia; Oludemi, Taofiq; Mandim, Filipa; Finimundy, Tiane C.; Prieto Lage, Miguel A.; Barros, LillianOne of the oldest known plants is the pomegranate tree (Punica granutum L.), native to the Mediterranean region. This plant's edible and non-edible portions have been shown to provide several health advantages. It has long been recognized as a source of bioactive chemicals, such as phenolic acids, flavonoids, tannins, alkaloids and amino acids. Large amounts of industrial waste are generated during the industrial processing of pomegranates, and 40 to 50% of the whole fruit is typically discarded 1. In this context, the present work was conducted to determine the chemical composition and bioactive properties of pomegranate leaves to support their potential utilisation as functional ingredients. Three extraction methodologies were evaluated: maceration, microwave and ultrasound-assisted extractions. The chemical composition of the different extracts was determined using HPLC-DAD-ESI/MS. The antioxidant potential was assessed using two cell-based assays: TBARS and CAA. Sulforhodamine B colourimetric assay was used to assess the antiproliferative capacity using several tumour cell lines and the primary culture of non-tumour cells (PLP2). The anti-inflammatory activity was measured through the extract's capacity to inhibit nitric oxide production. Finally, antimicrobial activity was evaluated using the microdilution method. The results showed that the main phenolic compounds found in the three extracts were gallic and caffeic acid derivatives, and flavonoids such as luteolin, apigenin, quercetin, and kaempferol derivatives. All the extracts exhibited capacity to inhibit tumor cell lines proliferation (GI50 of 19 - 76 μg/mL). Gastric adenocarcinoma (AGS) showed a higher sensitivity to all three leaf extraction methodologies. All three extracts presented lower IC50 values in the TBARS antioxidant assay (0.83 - 1.70 μg/mL) than the positive control Trolox (IC50 = 9.1 ± 0.3 μg/mL). The extracts presented a broad-spectrum antimicrobial inhibition with. K. pneumoniae showing the highest susceptibility to the extracts, (MIC values 0.6 mg/mL). These results suggest that pomegranate leaves can be sustainably exploited as a source of health-promoting biomolecules, to be used as functional ingredients in some biobased applications.
- Valorization of Punica granatum l. leaves extracts as a source of bioactive moleculesPublication . Marcelino, Sandra Andreia; Mandim, Filipa; Taofiq, Oludemi; Pires, Tânia C.S.P.; Finimundy, Tiane C.; Prieto Lage, Miguel A.; Barros, LillianDue to a lack of innovative valorization strategies, pomegranate processing generates a significant amount of residues with a negative environmental footprint. These by-products are a rich source of bioactive compounds with functional and medicinal benefits. This study reports the valorization of pomegranate leaves as a source of bioactive ingredients using maceration, ultrasound, and microwave-assisted extraction techniques. The phenolic composition of the leaf extracts was analyzed using an HPLC-DAD-ESI/MSn system. The extracts' antioxidant, antimicrobial, cytotoxic, anti-inflammatory, and skin-beneficial properties were determined using validated in vitro methodologies. The results showed that gallic acid, (-)-epicatechin, and granatin B were the most abundant compounds in the three hydroethanolic extracts (between 0.95 and 1.45, 0.7 and 2.4, and 0.133 and 3.0 mg/g, respectively). The leaf extracts revealed broad-spectrum antimicrobial effects against clinical and food pathogens. They also presented antioxidant potential and cytotoxic effects against all tested cancer cell lines. In addition, tyrosinase activity was also verified. The tested concentrations (50-400 mu g/mL) ensured a cellular viability higher than 70% in both keratinocyte and fibroblast skin cell lines. The obtained results indicate that the pomegranate leaves could be used as a low-cost source of value-added functional ingredients for potential nutraceutical and cosmeceutical applications.
- Pomeganate leaf extracts characterization and their potential use as a functional ingredientPublication . Marcelino, Sandra Andreia; Mandim, Filipa; Oludemi, Taofiq; Heleno, Sandrina A.; Finimundy, Tiane C.; Prieto Lage, Miguel A.; Barros, LillianThe pomegranate tree (Punica granatum L.) is indigenous to the Mediterranean region and is one of the oldest known plants. Studies have demonstrated their many biological and pharmaceutical benefits, which have been correlated to their bioactive content, namely secondary metabolites such as phenolic compounds [1]. However, 40 to 50% of the whole fruit is discarded, such as peel and leaves, originating valuable biowaste [2]. To support a circular economy and the prospective use of pomegranate biowaste as functional ingredients, the present research sought to ascertain the chemical composition and bioactive characteristics of pomegranate leaves. Maceration (ME), microwave (MAE), and ultrasound-assisted (UAE) extractions were the three extraction procedures chosen. HPLC-DAD-ESI/MS was used to identify and quantify the chemical composition of the various extracts. Two cell-based procedures were used to evaluate the antioxidant potential, TBARS and CAA. The antiproliferative potential was studied through sulforhodamine B colourimetric assays, using several tumour and a non-tumour cell lines. The extract's ability to reduce nitric oxide generation was used to determine its anti-inflammatory effects. Lastly, the microdilution technique was used to assess antimicrobial activity and cell viability assay conducted in two skin cell lines to support their possible use in topical formulation. According to the findings, the highest content of phenolic compounds was obtained in the MAE extract. Gallic acid, epicatechin, and granatin B were the most abundant compounds detected in all three extraction methodologies studied. All three extracts demonstrated lower IC50 values (0.83 - 1.70 g/mL) than the positive control Trolox (IC50 = 9.1 0.3 mg/mL), being the best results obtained for MAE and ME extracts. All extracts showed the capacity to halt the growth of tumour cell lines (GI50 between 19 and 76 g/mL), being gastric adenocarcinoma (AGS), the cell line with the highest sensitivity. All three extracts demonstrated broad-spectrum antibacterial inhibition, in which K. pneumoniae displayed the highest sensitivity to the extracts (MIC values of 0.6 mg/mL). In the tested skin cell lines, none of the studied extracts had any observable adverse effects, and more than 50% of viability was maintained after exposure to the highest tested concentrations of each extract (400 μg/mL). According to our findings, pomegranate leaves appear to be a source of health-promoting biomolecules. Those results sustain their exploitation and possible use by food and pharmaceutical industries in some biobased products, enhancing the circular economy and zero biowastes.
- Valorização de ingredientes naturais na inclusão de uma dieta saudávelPublication . Marcelino, Sandra Andreia; Barros, Lillian; Heleno, Sandrina A.Os consumidores, conscientes da utilização generalizada de aditivos artificiais, procuram cada vez mais por uma alimentação mais natural e, portanto, mais saudável. Esta mudança de comportamento está a forçar a indústria a responder a estes novos desafios, estando a indústria alimentar a trabalhar arduamente para se adaptar melhor a esta nova realidade. Neste trabalho foram usados três corantes de origem natural, um pigmento extraído de Amaranthus Caudatus L., a curcumina e paprica. Estes corantes foram incorporados em três produtos alimentares: cheesecake, gelado e manteiga. Os efeitos da incorporação destes corantes naturais foram avaliados através da análise dos parâmetros físicos em fresco (cor, pH, atividade da água e humidade), assim como pela sua composição centesimal, ao longo do tempo de prateleira de cada um dos alimentos. Globalmente, estes corantes naturais mostraram ter poder corante suficiente para cumprirem o seu propósito, sendo que o extrato de Amaranthus caudatus L., apresenta maior poder corante. Demonstraram também não alterar significativamente a composição nutricional das amostras, sendo por isso uma alternativa interessante pois espera-se que a utilização de corantes, sejam eles naturais ou artificiais, não afetem a composição do alimento em questão, tendo se verificado ligeiras alterações nas formulações de manteiga. Para verificar a aceitação por parte dos consumidores de alimentos corados com diferentes corantes alimentares foi lhes pedido para preencher um questionário depois da prova de amostras, e foi realizado à posteriori o estudo e o tratamento estatístico dos dados. Foi possível verificar que mais de 61% compraria um dos produtos corados com os corantes naturais em estudo, e que a cor laranja (atribuída à paprica) destaca-se pela persistência do sabor e pelo seu odor mais intenso nos três tipos de alimentos analisados. Em conclusão, estes corantes alimentares, foram estudados em três amostras alimentares de base láctea, tendo revelado potencial de aplicação na indústria alimentar devido à elevada estabilidade observada ao longo do tempo de prateleira destes alimentos, abrindo-se assim a oportunidade para o estudo mais aprofundado sobre a estabilidade destes ingredientes quando aplicados noutros produtos alimentares.