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Diatoms are currently considered valuable feedstocks for different biotechnological applications. To deepen the knowledge on the production of these microalgae, the diel pattern of batch growth, photosystem II performance, and accumulation of target metabolites of two commercially relevant diatoms, Phaeodactylum tricornutum and Skeletonema costatum, were followed outdoors in 100-L flat panel photobioreactors. S. costatum presented a higher light-to-biomass conversion resulting in higher growth than P. tricornutum. Both fluorescence data and principal component analysis pointed to temperature as a limiting factor for the growth of P. tricornutum. Higher protein and carbohydrate contents were found in P. tricornutum, whereas S. costatum fatty acids were charac-terized by a higher unsaturation degree. Higher productivities were found at 1 p.m. for protein, lipid, and ash in the case of S. costatum. Overall, S. costatum showed great potential for outdoor cultivation, revealing a broader temperature tolerance and increased biomass productivity than P. tricornutum. info:eu-repo/semantics/publishedVersion

Few studies have focused on the biotechnological potential of the marine cyanobacterium Synechocystis salina, in particular its bioactive metabolites. Hence, an opportunity exists to explore various solvent combinations in attempts to improve extraction of cellular contents of metabolites with antioxidant features as a part of a structured attempt to determine whether S. salina LEGE 06,155 holds a promise for industrial exploitation. Extracts were obtained by single extractions with four GRAS solvents: hexane (H), acetone (A), ethanol (E), and buffer phosphate (PBS) (series A) and successive extractions: H-E-PBS (series B), H-A-PBS (series C), and H-A-E-PBS (series D). The yields, antioxidant capacity, and cytotoxicity of the extracts, as well their content in pigments, lipids, and phenolic compounds were assessed. The combination of solvents of series B produced the highest yield of extraction, of 33.56%. Aqueous extracts for single (series A) and successive extractions (series B and C) presented the highest antioxidant capacity, for ABTS•+ and NO•−, respectively, and had a high content of phycobiliproteins (PBPs). The acetonic (series A) and ethanolic (series B) extracts also showed high antioxidant capacity, in particular for DPPH• and O2•−, respectively. Overall, organic extracts did not show signs of cytotoxicity. Series B extraction enhanced general pigment content, yet single extraction with ethanol (series A) produced the highest content in carotenoids, in particular echinenone and β-carotene, and PBS (series A) improved PBPs content. The lipid content was enhanced in series A (ethanol), while the total phenolic content was improved in series B. Synechocystis has accordingly demonstrated potential for eventual exploitation as biotechnological resource, in terms value-added compounds with antioxidant features.

Algal bioactive compounds, including pigments, polyunsaturated fatty acids, vitamins, proteins and phenolic compounds, have a wide range of increasing applications in the pharmaceutical, nutraceutical, feed, food, and cosmetics sectors. Extraction of such compounds with the purpose of obtaining nutraceutical and food ingredients at large has been under scrutiny over the last years towards greener and more sustainable bio-based processes. The step of the extraction is indeed considered the most expensive in algal-based industrial bioprocess, mainly owing to the high processing costs and environmental impacts. Therefore, the application of a biorefinery strategy (for biomass reutilization, via choice of specific green and generally recognized as safe solvent), will certainly help attain a more feasible and competitive solution. Furthermore, emerging extraction technologies are potentially suitable for application at a large scale, as part of a multi-integrated extraction strategy. This chapter accordingly addresses the most important (micro)algal-compounds possessing bioactivities, and already or likely to be used as functional ingredients; in addition, extraction procedures thereof are discussed, bearing in mind a biorefinery strategy – with major prospects and associated challenges.

A cyanobacterium of the genus Chroococcidiopsis has been recently unveiled to go to Mars, owing to its unique ability to stand harsh environmental conditions. Nevertheless, it has been little characterized in terms of other biotechnological applications. The aim of this study was thus to explore the biotechnological potential of Chroococcidiopsis sp. LEGE 06174, using GRAS (generally recognized as safe) solvents for the combined extraction of bioactive compounds. Due to its mucilaginous sheath, sulfuric acid, NaOH, PBS, DMSO 20%, DMSO 100%, and acetone were tested as preliminary treatment—in attempts to enhance pigment extraction. Extracts were then obtained by several series of successive extractions: phosphate-buffered saline (PBS) (serie A), ethanol-PBS (serie B), acetone-PBS (serie C), methanol-PBS (serie D), and acetone-methanol-PBS (serie E). Such extracts were screened for antioxidant capacity, cytotoxicity, and physicochemical characterization in terms of polysaccharides, pigments, and total phenolic compounds. Pre-treatment with PBS proved the most effective toward overall pigment extraction and dissolution of the said mucilaginous sheath. Chroococcidiopsis sp. extracts have unfolded some interesting bioactive features, particularly upon pre-treatment with PBS—i.e., high antioxidant capacity and high content in polysaccharides, scytonemin, phycobiliproteins, and phenolic compounds. The highest total pigment content was obtained in serie D, particularly carotenoids, whereas the highest level of phenolic compounds was found in serie B. All in all, Chroococcidiopsis sp. appears to constitute a significant biotechnological resource worthy of further exploitation in terms of high added-value compounds.

Microalgae have shown a great potential as a source of bioactive compounds, such lipidic compounds as fatty acids and carotenoids, with recognized benefit for human health. Their synthesis is directly affected by culturing conditions, but, as photosynthetic organisms, microalgae are greatly influenced by light, particularly light quality and irradiance, responsible for growth regulation and activation of biochemical pathways. Therefore, the aim of this work was to maximize biomass and high added-value compound productivity by Scenedesmus obliquus M2-1, using different light irradiances of two previously selected light sources for the production of antioxidant compounds: monochromatic red light (R) and a mixture of 40% of blue and 60% of red light (BR). Four irradiances (50, 100, 150, and 200 μmolphotons m−2 s−1) were accordingly evaluated for both light sources, upon productivity of biomass, fatty acids, carotenoids, and total antioxidant compounds. Changes in light quality and irradiance influenced these aforementioned productivities. Optimal condition for biomass productivity was 150 μmolphotons m−2 s−1, regardless light quality, for both sources. On the other hand, when the production targets carotenoids, polyunsaturated fatty acids, and/or total antioxidant capacity, the optimal condition was found to be at 200 μmolphotons m−2 s−1 under BR light.

Light plays a crucial role in photosynthetic microalgae, and its quality is a key point for biomass production and the variability of the organisms’ biochemical composition, with blue and red, the light bands being the most effectively absorbed by green algae. Recently, light-emitting diodes (LEDs) have emerged as an alternative that allow a better modulation of algal metabolism. The aim of this work was to study the effects of blue and red LEDs on antioxidant capacity, biomass productivity (Px) and carotenoid and fatty acid content over time in a microalga known to be a producer of antioxidant compounds, Scenedesmus obliquus M2-1. Monochromatic blue (B) and red (R) LEDs and two dichromatic combinations of blue and red (BR (40:60) and BR (50:50)) were tested, and fluorescent lamp (FL) was used as a control. Then, supplementation effects of near-infrared radiation (NIR) LEDs were assessed in the same in parameters. Results show that B induced higher PX and higher content in fatty acids when compared with other LED conditions tested. Scenedesmus obliquus M2-1 under R attained a higher antioxidant capacity and under BR (40:60) a higher carotenoid production, particularly lutein and β-carotene, when compared with that of FL. NIR supplementation in BR (40:60) and R treatments enhanced PX and fatty acid content, changing the profile of antioxidant compound production over time, but keeping the maximum concentration. Additionally, NIR supplementation decreased the total carotenoid production. This study reveals that the use of R and BR (40:60) LEDs are a useful tool to enhance the production of carotenoids in S. obliquus M2-1.

A laboratory experiment is described in which students carry out immobilization of the enzyme invertase via entrapment in alginate (Ca-alginate) and, accordingly, estimate values for kinetic consta...

Aim of this study The major aim of this work was to consistently optimize the production of biomass of the dinoflagellate Karlodinium veneficum and evaluate its extracts biotechnological potential application towards food, nutraceutical or/and pharmaceutical industries. Methods and results A successful approach of biomass production of K. veneficum CCMP 2936 was optimized along with the chemical characterization of its metabolite profile. Several temperatures (12, 16, 20, 25, 30°C), L1 nutrient concentrations (0.5×, 2×, 2.5×, 3×) and NaCl concentrations (20, 25, 30, 40 g L−1) were tested. The growth rate was maximum at 16°C, 2× nutrient concentration and 40 g L−1 of NaCl; hence, these conditions were chosen for bulk production of biomass. Methanolic extracts were prepared, and pigments, lipids and phenolic compounds were assessed; complemented by antioxidant and anti-inflammatory capacities, and cytotoxicity. Fucoxanthin and derivatives accounted for 0.06% of dry weight, and up to 60% (w/w) of all quantified metabolites were lipids. Said extracts displayed high antioxidant capacity, as towards assessed via the NO•− and ABTS•+ assays (IC50 = 109.09 ± 6.73 and 266.46 ± 2.25 µgE ml−1, respectively), unlike observed via the O2•− assay (IC25 reaching 56.06 ± 5.56 µgE ml−1). No signs of cytotoxicity were observed. Conclusions Karlodinium veneficum biomass production was consistently optimized in terms of temperature, L1 nutrient concentrations and NaCl concentration. In addition, this strain appears promising for eventual biotechnological exploitation. Significance and Impact of the Study This work provides fundamental insights about the growth and potential of value-added compounds of dinoflagellate K. veneficum. Dinoflagellates, as K. veneficum are poorly studied regarding its biomass production and added-value compounds for potential biotechnological exploitation. These organisms are difficult to maintain and grow in the laboratory. Thus, any fundamental contribution is relevant to share with the scientific community.

The application of microalgae in wastewater treatment has recently been at the forefront of interest due to the increasing concern about environmental protection and economic sustainability. This work aimed to study two chlorophyte species, Chlorella vulgaris and Scenedesmus acutus, co-cultured outdoors in centrate of municipal wastewater as a nutrient source. Two different thin-layer units were used in these trials & mdash; thin-layer cascade (TLC) and thin-layer raceway pond (TL-RWP), suitable for this purpose due to their high biomass productivity and better culture transparency when using muddy wastewater. The units were operated in batch, and subsequently in semi-continuous growth regime & mdash; and monitored in terms of photosynthetic performance, growth, nutrient removal rate, and bioactivity. The results showed that the co-cultures grew well in the centrate, achieving the maximum biomass densities of 1.3 and 2.1 g DW L-1 in TLC and TL-RWP, respectively, by the end of the batch regime and 1.9 and 2.0 g DW L-1 by the end of the semi-continuous regime. Although TL-RWP grown cultures showed faster growth, the TLC-one revealed better nutrient removal efficiencies batch wise than the culture grown in TL-RWP & mdash; removing up to 48% of total nitrogen and 43% of total phosphorus. Conversely, the latter was more efficient under the semi-continuous regime (54% and 42% consumption of total nitrogen and phosphorus, respectively). In the harvested biomass, an important antimicrobial activity (specifically antifungal) was detected. In this sense, the in-vitro growth of the oomycete Pythium ultimum was inhibited by up to 45% with regard to the control. However, no biostimulating activity was observed. The present findings confirm the possibility of using these two species for biomass production in municipal wastewater centrate using highly productive thin-layer systems. This technology can be a valuable contribution to circular economy since the produced biomass can be re-applied for agricultural purposes. SFRH/BD/129952/2017; UIDB/00511/2020; DINOSSAUR-PTDC/BBB-EBB/1374/2014-POCI-01-0145-FEDER-016640; info:eu-repo/semantics/publishedVersion

Tisochrysis lutea is a marine haptophyte rich in omega-3 polyunsaturated fatty acids (e.g., docosahexaenoic acid (DHA)) and carotenoids (e.g., fucoxanthin). Because of the nutraceutical applications of these compounds, this microalga is being used in aquaculture to feed oyster and shrimp larvae. In our earlier report, T. lutea organic crude extracts exhibited in vitro cytotoxic activity against human hepatocarcinoma (HepG2) cells. However, so far, the compound(s) accountable for the observed bioactivity have not been identified. Therefore, the aim of this study was to isolate and identify the chemical component(s) responsible for the bioactivity observed. Bioassay-guided fractionation through a combination of silica-gel column chromatography, followed by preparative thin layer chromatography (PTLC), led to the isolation of two diastereomers of a monoterpenoid lactone, namely, loliolide (1) and epi-loliolide (2), isolated for the first time in this species. The structural elucidation of both compounds was carried out by GC-MS and 1D (1H and 13C APT) and 2D (COSY, HMBC, HSQC-ed, and NOESY) NMR analysis. Both compounds significantly reduced the viability of HepG2 cells and were considerably less toxic towards a non-tumoral murine stromal (S17) cell line, although epi-loliolide was found to be more active than loliolide. FCT, Portugal: UID/Multi/04326/2019, UID/MAR/04292/2020 (MARE), UID/Multi/04046/2020 (BioISI) and from the grant of 0055 ALGARED +5E—INTERREG V-A; SFRH/BD/116604/2016; SFRH/BD/105541/2014 info:eu-repo/semantics/publishedVersion

Since temperature is an important factor affecting microalgal growth, photosynthetic rate and biomass composition, this study has accordingly focused on its effects on biomass yield and nighttime biomass loss, as well as photochemical changes, using Nannochloropsis oceanica as model species, grown in two outdoor 50-L tubular photobioreactors (PBR). In two independent trials, cultures were subjected to a diurnal light:dark cycle, under a constant temperature of 28 degrees C and, on the second trial, at 18 degrees C. Changes in culture performance were assessed by measuring growth, lipid and fatty acid composition of the biomass in both morning and evening. Our results revealed that N. oceanica shows a wide temperature tolerance with relevant nighttime biomass loss, that decreased at lower temperatures, at the expenses of its daily productivity. Fluorescence measurements revealed reversible damage to photosystem II in cells growing in the PBR under optimal thermal conditions, whereas microalgae grown at suboptimal ones exhibited an overall lower photosynthetic activity. Lipids were partially consumed overnight to support cell division and provide maintenance energy. Eicosapentaenoic acid (EPA) catabolism reached a maximum after the dark period, as opposed to their saturated counterparts; whereas lower temperatures led to higher EPA content which reached the maximum in the morning. These findings are relevant for the production of Nannochloropsis at industrial scale. European Cooperation in Science and Technology (COST) Action: European network for algal-bio-products (EUALGAE) [ES1408] Portuguese national funds from the Foundation for Science and Technology (FCT) [SFRH/BD/129952/2017] Laboratory for Process Engineering, Environment, Biotechnology and Energy -LEPABE -by the FCT/MCTES (PIDDAC) [UIDB/00511/2020] project: "LEPABE-2-ECO-INNOVATION" - North Portugal Regional Operational Program (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (ERDF) [NORTE-01-0145-FEDER-000005] project: "DINOSSAUR" - ERDF through Programa Operacional Competitividade e Internacionalizacao (COMPETE2020) [PTDC/BBB-EBB/1374/2014-POCI-01-0145-FEDER-016640] project: "SABANA"- European Union [727874] [UID/Multi/04326/2019] info:eu-repo/semantics/publishedVersion

Spontaneous fermentation by native strains of lactic acid bacteria has traditionally been used to preserve table olives, by taking advantage of a proactive and competitive reduction of growth of unwanted microorganisms. Starter cultures have proven effective in stabilizing fermentation processes at large. Despite the many starter cultures available commercially, few have originated on the native microflora – and table olives have seldom been processed via deliberate inoculation with them. This study screened variables relevant for the growth of Lactobacillus plantarum AFS13 originally obtained from brines in Alto Douro (Portugal); and for preservation thereof as the starter, should it prove appropriate. A 2 I V 6 − 2 factorial design was used to assess six parameters (and their interactions): 19 fermentation batches were carried out to ascertain growth rates. Temperature and aeration degree were the most significant processing parameters. Maximum growth rate, 0.356 h−1, was attained under anaerobic conditions, at 37 °C, and in the absence of NaCl; ca. 9 log10 CFU mL−1 was reached by 6 h. Glucose concentration, pH, and stirring rate proved not significant, so economic considerations should fix their optimum levels. The starter culture could be preserved for up to 4 mo after freeze-drying in sucrose, with an acceptable reduction in cell viability.

Spontaneous fermentation constitutes the basis of the chief natural method of processing of table olives, where autochthonous strains of lactic acid bacteria (LAB) play a dominant role. A thorough literature search has unfolded 197 reports worldwide, published in the last two decades, that indicate an increasing interest in table olive-borne LAB, especially in Mediterranean countries. This review attempted to extract extra information from such a large body of work, namely, in terms of correlations between LAB strains isolated, manufacture processes, olive types, and geographical regions. Spain produces mostly green olives by Spanish-style treatment, whereas Italy and Greece produce mainly green and black olives, respectively, by both natural and Spanish-style. More than 40 species belonging to nine genera of LAB have been described; the genus most often cited is Lactobacillus, with L. plantarum and L. pentosus as most frequent species-irrespective of country, processing method, or olive type. Certain LAB species are typically associated with cultivar, e.g., Lactobacillus parafarraginis with Spanish Manzanilla, or L. paraplantarum with Greek Kalamata and Conservolea, Portuguese Galega, and Italian Tonda di Cagliari. Despite the potential of native LAB to serve as starter cultures, extensive research and development efforts are still needed before this becomes a commercial reality in table olive fermentation. info:eu-repo/semantics/publishedVersion

This research effort aimed at a better understanding of microbial phenomena taking place during time in spontaneous sourdough fermentation for broa, a traditional Portuguese bread. Unlike most microbiological studies of sourdough, viable counts obtained were not limited to Lactobacillus and yeasts, but encompassed also molds, Gram-negative rods, endospore-(non)forming Gram-positive rods and catalase-positive/negative Gram-positive cocci. This less conventional approach unveiled the ubiquitous Bacillus genus throughout spontaneous broa sourdough fermentation. Presumptive yeasts, Lactobacillus and Bacillus were found to low levels after kneading, but became dominant by the end of regular (no aeration) fermentation. They apparently inhibit undesired rods and Staphylococcus, which were found to relatively high numbers after kneading. By 24 h of fermentation, lactic acid cocci accounted for an important fraction of biodiversity, and pH decreased significantly reaching about 4.1. Aeration accelerated the microbial dynamics. In terms of total viable counts, such a long-term fermentation appeared to reach a quasi-stationary state. Practical Applications Microbial studies encompassing a large set of culture media and incubation conditions were previously undertaken in samples of maize and rye flours, and sourdoughs for broa obtained from 14 artisanal producers and in two seasons. Microbial counts and identification unfolded a complex wild microbiota, and fermentation played a major effect upon all microbial groups. Flours and mother-dough microbiota were characterized during storage afterward. A similar approach was followed here to ascertain the microbiological profile of a wide range of microorganisms throughout long-term sourdough fermentation, in attempts to answer the questions: how microorganisms evolve, and how they respond to increasing aeration. These studies provide an innovative way to look at sourdough systems and microbial dynamics therein. Results showed the importance of increasing fermentation time to take full advantage of ecological competition against undesirable microorganisms, and emphasized the need for further studies to reveal the role of Bacillus in sourdough fermentations.

In recent decades, the world has faced increasing demand for energy, namely from fossil sources, which entails great environmental damage. Thus, green fuels with low environmental impact are urgently required, and production of biofuels from organic molecules of macro- and microalgae, such as carbohydrates, proteins, and lipids, is one alternative. However, a problem still remains as what to do with the spent biomass. In this regard, this chapter mainly focuses on how algae can be used as an efficient and economically viable biorefinery feedstock. To reinforce the algal biorefinery concept, a proper connection needs be established between the various input and output streams, as well as services to be provided by the industries involved; because a biorefinery using algae can only be effective through integration with other industries. This chapter is accordingly organized on three pillars: food/feed and nutraceutical applications, biofuel production, and environmental improvement via upgrading of spent biomass.

The marine environment accounts for most of the biodiversity on our planet, while offering a huge potential for the benefit and wellbeing of mankind. Its extensive resources already constitute the basis of many economic activities – but many more are expected in coming years. This book covers current knowledge on uses of marine algae to obtain bulk and fine chemicals, coupled with optimization of the underlying production and purification processes. Major gaps and potential opportunities in this field are discussed in a critical manner. The currrent trends pertaining to marine macro- and microalgae are explained in a simple and understandable writing style. This book covers a wide variety of topics, and as such it will be appropriate as both student text and reference for advances researchers in the field.

Recent years have witnessed a growing interest in microalgae, namely toward production of lipids for biodiesel, upgrade of effluents, and synthesis of high added-value compounds. However, industrial production remains a major issue – chiefly due to the constraints posed by classical cultivation systems. Common system designs for microalga cultivation exhibit major bottlenecks, with regard to such specific processing parameters as light, shear stress, gas exchange, and biofouling. Non-conventional photobioreactor arrangements have meanwhile arisen in response to those constraints – yet most of them raise challenges for practical use, despite a number of advantages. A detailed description and discussion of such non-conventional photobioreactor configurations is provided, including a qualitative comparison on their performance – after a brief introduction on parameters that affect photobioreactor performance, and the most common designs suitable for microalga cultivation.

Several members of the Regional Directorate of Agriculture of Entre-Douro-e-Minho (DRAEDM, Portugal) and several local farmers are hereby gratefully acknowledged for their cooperation, on a volunteer basis, in supplying samples for analysis. Financial support for author J. M. R. was provided by a Ph.D. fellowship within the program PRAXIS XXI – ref. PRAXIS XXI/BD/16060/98, administered by Fundacao para a Ciencia e a Tecnologia (Portugal) and supervised by author F. X. M. Partial financial support was received within program PAMAF – IED, through research grant “Pao de milho: caracterizacao do processo tradicional de producao e melhoramento tecnologico” – ref. PAMAF 1022, administered by Ministerio da Agricultura, Desenvolvimento Rural e Pescas (Portugal) and coordinated by author F. X. M.

Oscar L. Ramos, Ricardo N. Pereira, and Joana T. Martins gratefully acknowledge their post-doctoral grants (SFRH/BPD/80766/2011, SFRH/BPD/81887/2011, and SFRH/BPD/89992/2012, respectively) to the Fundacao para a Ciencia e Tecnologia (FCT, Portugal). The authors thank the FCT Strategic Project of UID/BIO/04469/2013 unit, COMPETE 2020 (POCI-01-0145-FEDER-006684), and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020—Programa Operacional Regional do Norte, as well as the project POCI-01-0145-FEDER-006939 (Laboratory for Process Engineering, Environment, Biotechnology and Energy—LEPABE) funded by FEDER funds through COMPETE2020—Programa Operacional Competitividade e Internacionalizacao (POCI)—and by national funds through FCT.

Oscar Leandro da Silva Ramos, Ricardo Nuno Correia Pereira, and Joana T. Martins gratefully acknowledge their postdoctoral grants (SFRH/BPD/80766/2011, SFRH/ BPD/81887/2011, and SFRH/BPD/89992/2012, respectively) from Fundacao para a Ciencia e Tecnologia (FCT, Portugal). All authors thank the FCT Strategic Project PEst-OE/EQB/LA0023/2013 and the Project “BioEnv—Biotechnology and Bioengineering for a sustainable world,” REF. NORTE-07-0124-FEDER-000048, cofunded by Programa Operacional Regional do Norte (ON.2—O Novo Norte), QREN, FEDER.

The development of bio-based nanostructures as nanocarriers of bioactive compounds to specific body sites has been presented as a hot topic in food, pharmaceutical and nanotechnology fields. Food and pharmaceutical industries seek to explore the huge potential of these nanostructures, once they can be entirely composed of biocompatible and non-toxic materials. At the same time, they allow the incorporation of lipophilic and hydrophilic bioactive compounds protecting them against degradation, maintaining its active and functional performance. Nevertheless, the physicochemical properties of such structures (e.g., size and charge) could change significantly their behavior in the gastrointestinal (GI) tract. The main challenges in the development of these nanostructures are the proper characterization and understanding of the processes occurring at their surface, when in contact with living systems. This is crucial to understand their delivery and absorption behavior as well as to recognize potential toxicological effects. This review will provide an insight into the recent innovations and challenges in the field of delivery via GI tract using bio-based nanostructures. Also, an overview of the approaches followed to ensure an effective deliver (e.g., avoiding physiological barriers) and to enhance stability and absorptive intestinal uptake of bioactive compounds will be provided. Information about nanostructures potential toxicity and a concise description of the in vitro and in vivo toxicity studies will also be given., Joana T. Martins, Oscar L. Ramos, Ana C. Pinheiro, Ana I. Bourbon, Helder D. Silva and Miguel A. Cerqueira (SFRH/BPD/89992/2012, SFRH/BPD/80766/2011, SFRH/BPD/101181/2014, SFRH/BD/73178/2010, SFRH/BD/81288/2011, and SFRH/BPD/72753/2010, respectively) are the recipients of a fellowship from the Fundacao para a Ciencia e Tecnologia (FCT, POPH-QREN and FSE, Portugal). The authors thank the FCT Strategic Project PEst-OE/EQB/LA0023/2013 and the project "BioInd-Biotechnology and Bioengineering for improved Industrial and Agro-Food processes," REF.NORTE-07-0124-FEDER-000028, co-funded by the Programa Operacional Regional do Norte (ON.2-O Novo Norte), QREN, FEDER. We also thank to the European Commission: BIOCAPS (316265, FP7/REGPOT-2012-2013.1) and Xunta de Galicia: Agrupamento INBIOMED (2012/273) and Grupo con potencial de crecimiento. The support of EU Cost Action FA1001 is gratefully acknowledged.

This work attempted to assess the effect of high intensity ultrasound (HIUS) upon development of bio-based delivery systems, from β-lactoglobulin (β-Lg) gelled microparticles, for encapsulation of a bioactive peptide concentrate (PepC). Solutions of 150 g L−1 of commercial β-Lg and 30 g L−1 PepC, at various pH values (3.0, 4.0 and 5.5), were accordingly subjected to gelation for 30 min using a dry bath kept at 80 °C. The gelled systems were then exposed to HIUS at 0–4 °C, and the effect of processing time (2.5–20.0 min) was ascertained. Laser light scattering and confocal microscopy were used to characterize the particle size distribution, prior to and immediately after HIUS treatment. Gels obtained at pH 5.5 and 4.0 were harder than those obtained at pH 3.0. Ultrasound treatment of gels produced an important reduction in particle mean diameter as sonication time elapsed. Confocal microscopy indicated that application of HIUS led to almost round and monodispersed particles, at both pH 5.5 and 4.0. The peptide encapsulation efficiency was assessed by chromatography and accompanied by assay for bioactivity, after precipitation of the encapsulated material and analysis of the soluble peptides therein., Tânia Tavares and Oscar L. Ramos gratefully acknowledge Post-Doctoral grants (SFRH/BPD/89360/2012 and SFRH/BPD/80766/2011, respectively) by Fundação para a Ciência e Tecnologia (FCT, Portugal), under the supervision of author F. Xavier Malcata. Partial financial support for the research expenses was provided by CYTED Program. The authors are grateful to FCT Strategic Project PEst-OE/EQB/LA0023/2013 and Project BioEnv—Biotechnology and Bioengineering for a sustainable world, Ref. NORTE-07-0124-FEDER-000048, co-funded by Programa Operacional Regional do Norte (ON.2—O Novo Norte), QREN, FEDER; and Project POCI-01-0145-FEDER-006939—Laboratório de Engenharia de Processos, Ambiente, Biotecnologia e Energia—LEPABE and NORTE-01-0145-FEDER-000005—LEPABE-2-ECO-INNOVATION, funded by Fundo Europeu de Desenvolvimento Regional (FEDER), through COMPETE2020—Programa Operacional Competitividade e Internacionalização (POCI) and Programa Operacional Regional do Norte (NORTE2020), and by national funds through FCT. The authors are also indebted to Nerina Camino and Ana Pilosof, from Departamento de Industrias, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, for generous and stimulating support, and for their relevant contributions to this field of research., info:eu-repo/semantics/publishedVersion

The influence that ohmic heating technology and its associated moderate electric fields (MEF) have upon production of whey protein isolate cold-set gels mediated by iron addition was investigated. Results have shown that combining heating treatments (90 °C, 5 min) with different MEF intensities let hydrogels with distinctive micro and macro properties i.e. particle size distribution, physical stability, rheological behavior and microstructure. Resulting hydrogels were characterized (at nano-scale) by an intensity-weighted mean particle diameter of 145 nm, a volume mean of 240 nm. Optimal conditions for production of stable whey protein gels were attained when ohmic heating treatment at a MEF of 3 V cm 1 was combined with a cold gelation step using 33 mmol L 1 of Fe2 +. The consistency index of hydrogels correlated negatively to MEF intensity, but a shear thickening behavior was observed when MEF intensity was increased up to 10 V cm 1. According to transmission electron microscopy, ohmic heating gave rise to a more homogenous and compact fine-stranded whey protein-iron microstructure. Ohmic heating appears to be a promising technique, suitable to tailor properties of whey protein gels and with potential for development of innovative functional foods., This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER006684) and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte. Rui M. Rodrigues, Oscar L. Ramos and Ricardo N. Pereira and, gratefully acknowledge to FCT their financial grants with references SFRH/BD/ 110723/2015, SFRH/BPD/80766/2011 and SFRH/BPD/81887/2011, respectively., info:eu-repo/semantics/publishedVersion

Whey proteins are widely used as nutritional and functional ingredients in formulated foods because they are relative inexpensive, generally recognized as safe (GRAS) ingredient and possess important biological, physical and chemical functionalities. Denaturation and aggregation behavior of these proteins is of particular relevance toward manufacture of novel nanostructures with a number of potential uses. When these processes are properly engineered and controlled, whey proteins may be formed into nanohydrogels, nanofibrils or nanotubes and be used as carrier of bioactive compounds. This review intends to discuss the latest understandings of nanoscale phenomena of whey protein denaturation and aggregation that may contribute for the design of protein nanostructures. Whey protein aggregation and gelation pathways under different processing and environmental conditions such as microwave heating, high voltage and moderate electrical fields, high pressure, temperature, pH and ionic strength were critically assessed. Moreover, several potential applications of nanohydrogels, nanofibrils and nanotubes for controlled release of nutraceutical compounds (e.g. probiotics, vitamins, antioxidants and peptides) were also included. Controlling the size of protein networks at nanoscale through application of different processing and environmental conditions can open perspectives for development of nanostructures with new or improved functionalities for incorporation and release of nutraceuticals in food matrices., Oscar L. Ramos, Ricardo N. Pereira and Clara Fuci~nos gratefully acknowledge their Post-Doctoral grants (SFRH/BPD/80766/2011, SFRH/BPD/ 81887/2011, and SFRH/BPD/87910/2012, respectively) to the Fundação para a Ciência e Tecnologia (FCT, Portugal). All authors thank the FCT Strategic Project PEst-OE/EQB/LA0023/2013 and the Project “BioEnv— Biotechnology and Bioengineering for a sustainable world”, REF. NORTE07-0124-FEDER-000048, co-funded by Programa Operacional Regional do Norte (ON.2–O Novo Norte), QREN, FEDER.