Valorização da casca de Ficus carica L. como fonte de pigmentos naturais: estudos de extração, caracterização química e biológica e incorporação em alimentos

Dupla diplomação com a UTFPR - Universidade Tecnológica Federal do ParanáDevido à sua coloração, a casca do figo, infrutescência da figueira (Ficus carica L.), pode ser considerada como fonte potencial de antocianinas. Neste contexto, o objetivo deste estudo consistiu na comparação de diferentes técnicas de extração (assistidas por calor, ultrassons e micro-ondas) para a recuperação de antocianinas. Adicionalmente, e para as condições ótimas de extração, o extrato obtido foi caracterizado quanto à composição química e propriedades bioativas (atividades antioxidante, antimicrobiana, anti-inflamatória e citotóxica). A fim de obter as condições que maximizam a extração de antocianinas, foi utilizada a metodologia de superfície de resposta usando um delineamento composto central circunscrito de três variáveis com cinco níveis. As variáveis relevantes testadas e otimizadas neste estudo foram o tempo, temperatura e composição do solvente (razão etanol/água) para os métodos de extração assistidos por calor e micro-ondas. Para o método de ultrassons a temperatura foi mantida constante enquanto a potência ultrassônica foi a variável estudada, juntamente com o tempo e a composição do solvente. A composição em antocianinas foi determinada por HPLC-DAD-ESI/MS e as respostas utilizadas como critérios foram: quantificação da cianidina-3-rutinósido (C) no resíduo extraído (R; mg C/g R) e na casca do figo (P; mg C/g P dw) e a relação de ambas (g R/g P dw). A extração ultrassónica mostrou ser o método mais eficaz, conduzindo a extração de 7,0 ± 0,2 mg C/g R, 3,11 ± 0,05 mg C/g P dw e 0,19 ± 0,001 g R/g P dw nas condições ótimas globais de extração (21 min e 310 W com 100% de etanol e 180 g/L). Em relação às propriedades bioativas, o extrato otimizado de casca de figo apresentou atividade antioxidante com EC50 de 2,447 mg/mL pelo método de TBARS e EC80, em 60 min (EC80 (60 min)) e em 120 min (EC80 (120 min)) de 0,52 e 1,62 mg/mL, respetivamente pelo método OxHLIA. O extrato revelou, para diferentes bactérias Gram-positivo e Gram-negativo, atividade antimicrobiana, além de não apresentar citotoxicidade para células não tumorais. Porém, não apresentou atividade anti-inflamatória ou citotóxica para células tumorais humanas. O estudo da sua aplicação num produto alimentar foi efetuado tendo como objetivo a preparação de coberturas para donuts (glacê). Em síntese, os resultados obtidos colocam em evidência a potencial aplicação da casca de F. carica como fonte de antocianinas para diferentes setores industriais, nomeadamente alimentar, farmacêutico e cosmético, agregando os benefícios provenientes da sua atividade antioxidante e antimicrobiana ao seu poder corante.Due to its coloration, the fig (Ficus carica L.) peel, a by-product of fruit processing and/or consumption, is a potential source of anthocyanin compounds. In the present study, different extraction techniques (heat, ultrasound, and microwave), aiming to recover the anthocyanin pigments and optimize its extraction, were compared by response surface methodology using a circumscribed central composite design of three variables with five levels. The relevant tested and optimized variables were time, temperature, and solvent composition (ethanol/water ratio) for the heat and microwave extraction methods. For the ultrasound method, the variables were the ultrasonic power, time, and solvent composition. The anthocyanin composition of the extract was determined by HPLC-DAD-ESI/MS, and the used criteria responses were the quantification of cyanidin 3-rutinoside (C) in the extracted residue (R; mg C/g R) and in the dried fig peel (P; mg C/g P dw), and the extraction yield of the obtained residue (g R/g P dw). Ultrasound extraction was the most effective method, yielding 17,0 ± 0,2 mg C/g R, 3,11 ± 0,05 mg C/g P dw e 0,19 ± 0,001 g R/g P dw at the optimal global extraction conditions (21 min and 310 W with 100% of ethanol and 180 g/L). Regarding its bioactive properties, the fig peel optimized extract showed antioxidant activity with EC50 of 2,447 mg/mL by the TBARS method and EC80 in 60 min (EC80 (60 min)) and in 120 min (EC80 (120 min)) of 0,52 and 1,62 mg/mL, respectively by the OxHLIA method. The extract revealed, for different Gram-positive and Gram-negative bacteria, antimicrobial activity, besides not presenting cytotoxic potential against non-tumor cells. However, the extract showed no anti-inflammatory or cytotoxic potential against human tumor cells. Its application in a food product was carried out aiming at the preparation of toppings for donuts. In conclusion, the obtained results highlight the potential application of F. carica peels as source of anthocyanin compounds to be considered in different industries, such as food, pharmaceutical and cosmetic, adding the benefits derived from its antioxidant and antimicrobial activity to the colorant power.Este trabalho é financiado pelo Fundo Europeu de Desenvolvimento Regional (FEDER) através do Programa Operacional Regional Norte 2020, no âmbito do Projeto NORTE-01-0145-FEDER-023289 (DeCodE) e Norte-01-0247-FEDER-024479 (projeto Mobilizador ValorNatural), e pelo programa FEDER-Interreg España-Portugal, no âmbito do projeto 0377_Iberphenol_6_E.Dupla diplomação com a UTFPR - Universidade Tecnológica Federal do Paran

Recovery of bioactive anthocyanin pigments from Ficus carica L. peel by heat, microwave, and ultrasound based extraction techniques

Due to its coloration, the fig (Ficus carica L.) peel, a by-product of fruit processing and/or consumption, is a potential source of anthocyanin compounds. In the present study different extraction techniques (heat, ultrasound, and microwave) were compared aiming to recover the anthocyanin pigments and optimize its extraction conditions. A response surface methodology tool with three factors and five levels for each factor was used according to a circumscribed central composite design. The variables tested for the heat and microwave extraction methods were time, temperature, and solvent proportion (ethanol/water ratio), meanwhile, for the ultrasound method, the variables tested were the ultrasonic power, time, and solvent proportion. The anthocyanin composition of the extract was determined by HPLC-DAD-ESI/MS, and the used criteria responses were: i) quantification of cyanidin 3-rutinoside (C) in the extracted residue (mg C/g R) and in the dried peel (mg C/g P dw), and the extraction yield of the obtained residue (g R/g P dw). Ultrasound extraction was the most effective method, yielding 3.82 mg C/g R at the optimal global extraction conditions (21 min, 310 W, and 100% of ethanol). Additionally, the solid-to-liquid ratio effect was studied at the optimal conditions, using a dose-response format, in view of its plausible transference to industrial level. For the ultrasound method, an increased non-linear relationship was observed for concentrations in the range 5 to 200 g/L, being the optimal solution close to 150 g/L. In brief, the obtained results show the potential of fig peels as a source of anthocyanin pigments, with potential uses in various industrial fields, such as food, pharmaceutical, and cosmetic.The authors are grateful to the Foundation for Science and Technology (FCT, Portugal) and FEDER under Programme PT2020 for financial support to CIMO (UID/AGR/00690/2013); to FCT for L. Barros research contract and C. Pereira grant (SFRH/BPD/122650/2016) under the Programa Operacional Capital Humano (POCH) supported by the European Social Fund and National Funds of MCTES (Ministério da Ciência, Tecnologia, e Ensino Superior); to FEDERInterreg España-Portugal programme for financial support through the project 0377_Iberphenol_6_E.; to European Structural and Investment Funds (FEEI) through the Regional Operational Program North 2020, within the scope of Mobilizador project Norte-01-0247-FEDER-024479: ValorNatural® and Project NORTE-01-0145-FEDER-023289: DeCodE. This work was also financially supported by: Project POCI-01-0145- FEDER-006984 – Associate Laboratory LSRE-LCM funded by FEDER through COMPETE2020 - Programa Operacional Competitividade e Internacionalização (POCI) – and by national funds through FCT - Fundação para a Ciência e a Tecnologia. To Xunta de Galicia for financial support to M.A. Prieto grant.info:eu-repo/semantics/publishedVersio

Ficus carica L. and Prunus spinosa L. extracts as new anthocyanin-based food colorants: a thorough study in confectionery products

The present work describes the evaluation of fig peels and blackthorn fruit extracts as natural purple colorants in doughnuts (icing) and in a typical Brazilian pastry called “beijinho”. The extracts were screened for their antioxidant activity as well as their antibacterial capacity. Nutritionally, the employed extracts did not induce significant changes, contrarily to the observed for the rheological features, mainly the darker purple tone observed when blackthorn extract was used in the icing solution. After 24 h, both prepared formulations showed a decrease in color intensity, with no significant differences between fig and blackthorn extracts. In turn, the firmness and consistency of the doughnuts benefited from using natural colorants in the icing solution, while “beijinhos” became softer and chewier, which are valued attributes. A significant increase in the antioxidant and antimicrobial activities was also observed for both natural extracts. Accordingly, the evaluated extracts are promising candidates as natural food colorants.The authors would like to thank the Foundation for Science and Technology (FCT, Portugal) and FEDER, under Program PT2020 for financing CIMO (Centro de Investigação de Montanha) (UID/AGR/ 00690/2019). This work was further funded by the European Structural and Investment Funds (FEEI) through the Regional Operational Program North 2020 within the scope of the Mobilizador ValorNatural® project, to whom the author M. Carocho thanks for his contract. Further acknowledgments are due to the national funding by FCT, P.I., through the institutional scientific employment program-contract for L. Barros’ contracts, and though the celebration of program-contract foreseen in No. 4, 5 and 6 of article 23° of Decree-Law No. 57/2016, of 29th August, amended by Law No. 57/2017, of 19th July for C. Pereira’s contract.info:eu-repo/semantics/publishedVersio

Activation of pro-BDNF by the pericellular serine protease plasmin

Brain-derived neurotrophic factor (BDNF) is secreted as either a mature furin-processed form or an unprocessed pro-form. Here, we characterise the extracellular processing of pro-BDNF by the serine protease plasmin. Using recombinant BDNF, maintained in the pro-form by site-directed mutagenesis or inhibition of furin, we demonstrate that plasmin (but not related proteases) is a specific and efficient activator of pro-BDNF. The proteolytic cleavage site is identified as Arg125-Val, within the consensus furin-cleavage motif (RVRR), generating an active form that stimulated neurite outgrowth on TrkB-transfected PC12 cells. Furthermore, we demonstrate that this processing can also occur in the pericellular environment by the action of cell-associated plasminogen activators

Chestnut and lemon balm based ingredients as natural preserving agents of the nutritional profile in matured “Serra da Estrela” cheese

Chestnut flowers, lemon balm plants and their decoctions were incorporated into "Serra da Estrela" cheese, to assess their potential to preserve its nutritional properties and provide new foodstuffs. The analyses were carried out after the normal ripening period of 1month and after 6months of storage. The most abundant nutrients were proteins and fats. The most abundant minerals were Ca and Na, while C16:0 and C18:1 were the main fatty acids. Saturated fatty acids were the most abundant, followed by the monounsaturated. Moisture seemed to be lower in the samples with the plants incorporated. The dried plants, when incorporated, seemed to be more efficient as preservers then the decoctions, although these better preserved the proteins. These plants can be regarded as promising natural preservers in foodstuffs cheese, given the preservation of key parameters and the slight impact on the nutritional value.The authors are grateful to Queijos Casa Matias, Lda and Mais Ervas, Lda. For providing the cheese and M. officinalis samples, respectively. The authors also acknowledge PRODER project No. 46577-PlantLact, the Foundation for Science and Technology (FCT, Portugal) for financial support to the CIMO research centre (Pest-OE/AGR/UI0690/2014) and ALIMNOVA research group (UCM-951505/2012), J.C.M. Barreira acknowledges the FCT for his post-doctoral grant (BPD/72802/2010)

Laccases in food processing: Current status, bottlenecks and perspectives

Background: Laccases (benzenediol:oxygen oxidoreductases, EC 1.10.3.2) catalyze the oxidation of a wide variety of organic and inorganic substrates, typically p-diphenols with a concomitant reduction of oxygen (O2) to water. Several molecules naturally occurring in foods and beverages (e.g., phenols, carbohydrates, unsatured fatty acids and thiol-containing proteins) can be modified by laccases. Hence, the interaction between laccase and these molecules can and has been widely explored by the food industry for various technological purposes. Scope and approach: The present work aims at providing a critical review on the current uses of laccases in food processing, at discussing the main bottlenecks for its popularization, and at presenting future perspectives. Both scientific reports and patents, covering preferably the last five years, were considered. Key findings and conclusions: Several traditional uses of laccases in food processing including baking, beverage, and dairy industries were detailed. Special efforts were developed, however, in analyzing future perspectives. The latter includes the application of laccases in the synthesis of new compounds with functional properties, such as antioxidant and antimicrobial activities. No less attention was devoted to the recent developments in the field of crosslinking of polymers, such as proteins and polysaccharides. Scaling up of the production of the laccase itself and especially of the novel products derived from its applications in the food sector will be essential for cost reduction and, consequently, for market expansion.This work was supported by the National Council of Scientific and Technological Development (CNPq, Proc. 404898/2016-5). R.C.G. Corrêa is a research grant recipient of Cesumar Institute of Science Technology and Innovation (ICETI). C.G. Kato (Proc. 151189/2019-6), E. Backes (Proc. 304406/2019-8), R.M. Peralta, R.A. Peralta, R.F. Peralta Muniz Moreira and A. Bracht are research grant recipients from CNPq.info:eu-repo/semantics/publishedVersio

Blueberry bagasse recovery for the development of enriched juices

The blueberry, Vaccinium corymbosum L., is considered one of the most antioxidant fruits and it is consumed worldwide for its properties [1]. The development of different industrialized products, as juices and derivatives, generates a considerable quantity of residues, such as bagasse, which are rarely used by the industry for other applications [2]. However, these residues contain valuable chemical compounds that can provide beneficial properties to human health [3]. The main interest of this study was, therefore, to recover the bagasse produced in blueberry juice production to prepare a rich extract to incorporated in the juice. Thus, juices with and without bagasse extract were developed, analyzed, and compared in order to validate the applicability of the extract in this matrix, to enhance the bioactive properties (antioxidant and antimicrobial), the nutritional value (AOAC methods), and the composition in free sugars (HPLC-RI) and phenolic compounds (HPLC-DAD/ESI-MS). The two juices presented a balanced nutritional value, with carbohydrates as the main macronutrients. The free sugars found in higher concentrations were fructose and glucose. In terms of phenolic composition, in the juice and extract-enriched juice, the same nonanthocyanin compound was detected (3-O-caffeoylquinic acid). Regarding anthocyanins, both samples revealed cyanidin derivatives as main compounds. Regarding bioactive properties, the juice enriched with the bagasse extract showed higher antioxidant capacity than the blueberry juice. None of the two juices showed cytotoxicity, being considered safe for consumption. Thus, this study corroborated the importance of the recovery of food industry waste for the development of novel foodstuff.The authors are grateful to Foundation for Science and Technology (FCT, Portugal) for financial support through national funds FCT/MCTES to CIMO (UIDB/00690/2020). National funding by FCT, P.I., through the institutional scientific employment program-contract for C. Pereira, M.I. Dias, and L. Barros contracts and A.K. Molina PhD grant (2020.06231.BD). To FEDER-Interreg España-Portugal programme for financial support through the project TRANSCoLAB 0612_TRANS_CO_LAB_2_P.info:eu-repo/semantics/publishedVersio

Blueberry (Vaccinium myrtillus L.) juice and bagasse for antioxidant products development

Blueberry, Vaccinium corymbosum L., is a fruit consumed worldwide and considered one of the most antioxidant fruits [1]. Its pulp, seeds, and peel contain different chemical compounds responsible for its beneficial properties for human health [2]. This knowledge has led to the development of different industrialized products, such as juices and derivatives, in search of more practical forms of consumption. However, in the manufacture of these products, a large amount of waste is generated and has no further application, generating an environmental problem and a high cost for the industry [3]. In this context, this study aimed to evaluate the blueberry juice in terms of bioactive properties (antioxidant and antimicrobial), nutritional value (AOAC methods), free sugars (HPLC-RI), organic acids (UFLC-PDA), and phenolic (HPLC-DAD/ESI-MS) composition. Moreover, the hydroethanolic extract of blueberry bagasse, resulting from juice production, was also analyzed regarding its phenolic composition and bioactivity, to assess its applicability as source of added-value molecules to be used in food industry. The blueberry juice presented a balanced nutritional value, with carbohydrates as the main macronutrients. The free sugars found in higher concentrations were fructose and glucose. Four organic acids were also identified, with quinic acid as the most abundant. As for the phenolic composition of the juice, four phenolic acids and six anthocyanins were detected. With respect to bagasse, eight anthocyanin compounds were identified in significant amounts, with cyanidin-3- O-glucoside and malvidin-3-O-glucoside being the most abundant compounds, and four non- anthocyanin compounds, with a prevalence of cis 5-O-caffeoylquinic acid and trans 5-O- caffeoylquinic. Concerning the bioactive properties, as expected considering their chemical composition, both the juice and the bagasse extract revealed a strong antioxidant capacity, being able to inhibit lipid peroxidation and oxidative hemolysis. None of the two showed cytotoxic properties, being considered safe for food consumption. Thus, this study validated the nutritional and bioactive quality of blueberry juice and justified the application of bagasse in the development of novel foods.The authors are grateful to the Foundation for Science and Technology (FCT, Portugal) for financial support through national funds FCT/MCTES to CIMO (UIDB/00690/2020). National funding by FCT, P.I., through the institutional scientific employment program-contract for C. Pereira, M.I. Dias, and L. Barros contracts and A.K. Molina PhD grant (2020.06231.BD). To FEDER-Interreg España-Portugal programme for financial support through the project TRANSCoLAB 0612_TRANS_CO_LAB_2_P.info:eu-repo/semantics/publishedVersio

Mycosterols

Sterols are amphipathic lipids that play essential roles in the physiology of eukaryotic organisms in general. The fungal sterols are collectively known as mycosterols and they exert numerous physiological functions. For humans, the interest on this class of compounds relies heavily on the fact that they can promote health benefits. For this reason, fungal extracts rich in sterols of various forms are valuable and promising ingredients. One of the best-known benefits of mycosterols is their inhibitory actions on cholesterol absorption and biosynthesis, but there are several interesting regulatory and modulatory phenomena that mycosterols can affect and that might eventually be of therapeutic interest. Within this domain, the practical application of mycosterols or mycosterol-enriched fungal extracts presents several challenges. The latter include isolation of novel bioactive mycosterols from still underexploited fungi species, the optimization of existing methodologies for production and recovery, extensive study of their applications and, finally, substantial clinical trials for attesting their health benefits and safety.The authors are grateful to Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação Araucária, and Instituto Cesumar de Ciência, Tecnologia e Inovação (ICETI) for financial support.info:eu-repo/semantics/publishedVersio

Ficus carica L. infructescence exploitation for anthocyanin-rich extracts preparation: optimized extraction, bioactivity, and application as food colorant.

Given its coloration, the peel of the infructescence of Ficus carica L. a by-product of fruit processing and/or consumption, was considered as a potential source of anthocyanin compounds. In the present study, different extraction techniques (heat, ultrasound, and microwave), aiming to recover the anthocyanin pigments and optimize its extraction, were compared by response surface methodology using a circumscribed central composite design of three variables with five levels. The relevant tested and optimized variables were time, temperature, and solvent composition (ethanol/water ratio) for heat and microwave extraction methods. For ultrasound method, the variables were the ultrasonic power, time, and solvent composition. The anthocyanin profile of the extract was determined by HPLC-DAD-ESI/MS, and the used criteria responses were the quantification of cyanidin 3-rutinoside (C) in the extracted residue (R; mg C/g R) and in the dried fig peel (P; mg C/g P dw), and the extraction yield of the obtained residue (g R/g P dw). Ultrasound extraction was the most effective method, yielding 17.0 mg C/g R, 3.11 mg C/g P dw, and 0.19 g R/g P dw at the optimal global extraction conditions (21 min, 310 W, and 100% of ethanol). Regarding its bioactive properties, the fig peel optimized extract showed antioxidant activity with an EC50 value of 2,447 mg/mL in the TBARS method and EC80 values of 52 and 1.62 mg/mL, in 60 and 120 min, respectively, in the OxHLIA assay. The extract also revealed antimicrobial activity in different Gram-positive and Gram-negative bacteria and did not present cytotoxicity against non-tumour cells. Its application in a food product was carried out through the preparation of a donut topping. The obtained results highlight the potential application of F. carica peels as sources of anthocyanins to be considered in food industry, adding the benefits derived from its antioxidant and antimicrobial activity to the colorant power.FCT and FEDER under Programme PT2020 for financial support to CIMO (UID/AGR/00690/2013), LA LSRE-LCM (POCI-01-0145-FEDER-006984), Carla Pereira (SFRH/BPD/122650/2016) grants, and L. Barros contract. Interreg España-Portugal for financial support to project 0377_Iberphenol_6_E. FEEI through the Regional Operational Program North 2020, within the scope of Project NORTE-01-0145-FEDER-023289: DeCodE and Norte-01-0247-FEDER-024479: ValorNatural®. Xunta de Galicia for financial support to M.A. Prieto grant.info:eu-repo/semantics/publishedVersio