Percorrer por autor "Coelho, Natacha"
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- Approaches for the cryopreservation of Plantago algarbiensis, a rare endemic species of the AlgarvePublication . Coelho, Natacha; González-Benito, María Elena; Romano, AnabelaPlantago algarbiensis is an endangered endemic species from the Algarve, Portugal. The main goal of this study was to investigate the viability of cryopreservation procedures in the conservation of seeds and nodal explants from this species. Seeds were directly immersed in liquid nitrogen (LN) for 30 days. Two methods were tested for the cryopreservation of nodal explants, namely droplet-vitrification and encapsulation-dehydration. For both methods, nodal segments were precultured on Murashige and Skoog (MS) medium and recovered on MS supplemented with 0.2 mg l(-1) 6-benzyladenine (BA), after freezing. After 30 days in LN, the germination capacity of seeds was not affected. The regrowth percentages of cryopreserved nodal segments were approximately 60%. With the droplet-vitrification method, a regrowth percentage of 60.0+/-15.2% was obtained after 120 min exposure to PVS2 (plant vitrification solution 2) and with encapsulation-dehydration method the highest percentage, 63.3+/-9.6%, was achieved after 3 h desiccation. Seed cryopreservation and cryopreservation of nodal segments by droplet-vitrification and encapsulation-dehydration are therefore effective approaches for the conservation of P. algarbiensis.
- Comparative analysis of isoprostanoid profiles in Chlorella sorokiniana grown under autotrophic and heterotrophic conditionsPublication . Conde, Tiago; Lopes, Diana; Gros, Valérie; Reversat, Guillaume; Oger, Camille; Galano, Jean-Marie; Vigor, Claire; Rodrigues, Alexandre M. C.; Coelho, Natacha; Cardoso, Helena; Domingues, M. Rosário; Durand, ThierryOxylipins are bioactive lipid mediators derived from polyunsaturated fatty acids (PUFAs), with roles in oxidative stress responses, immunomodulation, and inflammation. While microalgae are recognized as valuable sources of oxylipins, their profiles remain less studied across different species and cultivation conditions. In this study, we characterized the non-enzymatic oxylipin profile of Chlorella sorokiniana grown under autotrophic and heterotrophic conditions to assess the influence of cultivation strategies on their production. A total of 22 isoprostanoids, mainly Phytoprostanes (PhytoP), Phytofurans (PhytoF), Isoprostanes (IsoP), and Neuroprostanes (NeuroP). Autotrophic cultivation resulted in a higher accumulation of isoprostanoids, particularly the alpha-linolenic acid (ALA) derivatives, PhytoP and PhytoF species, likely due to oxidative stress induced by fluctuating light and temperature conditions. In contrast, heterotrophic growth, performed under controlled conditions, yielded lower overall oxylipin levels highlighting the presence of 10(R)-10-F4t-NeuroP which was only present in heterotrophic Chlorella. We observed a correlation between the PUFA composition of Chlorella and its non-enzymatic oxylipin profile. Notably, several oxylipins identified in Chlorella have been associated with antiinflammatory, immunomodulatory, and neuroprotective properties, emphasizing the potential of this micro-alga as a source of high-value bioactive oxylipins. This study paves the way to the utilization of Chlorella as a source of bioactive oxylipins, as well as to develop cultivation strategies to enhance the production of these lipid mediators.
- Cryopreservation as a tool for preserving genetic variability of endangered species endemic from Algarve regionPublication . Coelho, Natacha; Romano, Anabela; González-Benito, María ElenaThis thesis aims the development of complementary conservation strategies based on plant biotechnology to support the preservation of three rare endemic species of the Algarve, Portugal. The species studied were Thymus lotocephalus G. López & R. Morales, Plantago algarbiensis Samp. and Tuberaria major (Willk.) P. Silva & Rozeira. An in vitro propagation protocol was firstly developed for T. lotocephalus using seedlings as explants (Chapter 2.1). The germination conditions and feasibility of cryopreservation was also studied for T. lotocephalus seeds (Chapter 2.2). In Chapter 2.3 a shoot tip cryopreservation protocol was developed for this species and the genetic stability of the cryopreserved material was assessed. For P. algarbiensis, the work was initiated with the cryopreservation of seeds and nodal segments (Chapter 3.1), followed by the study of the genetic diversity of three wild populations (Chapter 3.2). Chapter 4.1 was dedicated to the improvement of the germination and cryopreservation of T. major seeds and the development of a shoot tip cryopreservation protocol. An efficient micropropagation protocol was established for the mass production of T. lotocephalus (Chapter 2.1) and the seeds germination requirements were determined (Chapter 2.2). From the shoots produced in vitro, apices were excised and used in the development of a shoot tip cryopreservation protocol (Chapter 2.2). P. algarbiensis nodal segments were successfully cryopreserved using two different methods (Chapter 3.1). The structure of P. algarbiensis populations was effectively evaluated using molecular markers, revealing high levels of genetic diversity within populations (Chapter 3.2). Cryopreservation proved to be a suitable method for the conservation of seeds from both species (Chapters 2.2 and 3.1). Finally, the germination conditions and cryopreservation of T. major seeds were considerably improved and the cryopreservation of T. major shoot tips was accomplished without major optimizations of the different procedures (Chapter 4.1). In conclusion, during the course of this thesis different ex situ conservation strategies were developed giving a major contribution to the preservation of T. lotocephalus, P. algarbiensis and T. major.
- Cryopreservation of shoot tips from the endangered endemic species Tuberaria majorPublication . Coelho, Natacha; González-Benito, María Elena; Romano, AnabelaTuberaria major is an endangered endemic species from the Algarve, in the south of Portugal. We investigated two techniques for the cryopreservation of T. major shoot tips, namely vitrification and encapsulation-dehydration. Before the cryopreservation trials, shoot tips were precultured for 1 day on liquid Murashige and Skoog (MS) medium containing 0.3 M sucrose. For the vitrification method, shoots tips were exposed for 0, 30, 60, 90 and 120 min to plant vitrification solution 2 (PVS2). As for the encapsulation-dehydration method, shoot tips were dried inside a laminar air flow cabinet for 0, 1, 2, 3, 4, 5 and 6 h at room temperature. The highest regrowth percentages were approximately 60 and 67 % for vitrification and encapsulation-dehydration, respectively. The best times were 60 min exposure to PVS2 for vitrification and 3 h desiccation for encapsulation-dehydration. Though these are preliminary results, the use of the cryopreservation techniques tested here proved to be an important asset in the conservation of this endangered species and will complement the conservation strategies previously developed.
- Culturable yeast diversity associated with industrial cultures of the microalga microchloropsis gaditana and their ability to produce lipids and biosurfactantsPublication . Matos, Madalena; Fernandes, Mónica A.; Costa, Inês; Coelho, Natacha; Santos, Tamara; Rossetto, Veronica; Varela, João; Sá-Correia, IsabelThe marine oleaginous microalga Microchloropsis gaditana (formerly Nannochloropsis gaditana) exhibits a high capacity to thrive in a broad range of environmental conditions, being predominantly utilized as feed in aquaculture. This article reports the characterization of the culturable yeast population present during the scale-up process of M. gaditana cultivation at Necton S.A. facilities, from 5 L flasks until tubular photobioreactors. The 146 yeast isolates obtained, molecularly identified based on D1/D2 and ITS nucleotide sequences, belong to the species Rhodotorula diobovata, R. mucilaginosa, R. taiwanensis, R. sphaerocarpa, Vishniacozyma carnescens, Moesziomyces aphidis, and Meyerozyma guilliermondii. The yeast abundance was found to increase throughout upscaling stages. The yeast populations isolated from microalgal cultures and water samples share phylogenetically close isolates, indicating a possible common source. The impressive high percentage of red yeasts isolated (90%) is consistent with the recognized role of carotenoid pigments in yeast photoprotection. Sixty yeast isolates were tested for lipid (Nile Red staining) and biosurfactant (oil drop dispersion and emulsification index) production. Results revealed that these capacities are common features. Microbial lipids and biosurfactants have promising biotechnological applications. Moreover, biosurfactants can fulfill various physiological roles and provide advantages in natural environments contributing to the promising use of yeasts as probiotics in microalgae production.
- Elicitation improves rosmarinic acid content and antioxidant activity in Thymus lotocephalus shoot culturesPublication . Gonçalves, Sandra; Mansinhos, Inês; Rodríguez Solana, Raquel; Perez-Santin, Efren; Coelho, Natacha; Romano, AnabelaThymus lotocephalus G. Lopez & R. Morales is an aromatic species endemic to the south of Portugal with medicinal properties. The aim of this work was to evaluate the effect of elicitors, cytokinin, and different sucrose concentrations on chemical composition and antioxidant activity of extracts from in vitro regenerated shoots. Elicitors (YE: yeast extract, salicylic acid, and AgNO3) were added directly to the medium. Phytochemical analysis include evaluation of total phenolic (TPC), flavonoid (TFC), and rosmarinic acid (RAC) contents. The TPCs and TFCs were determined by Folin-Ciocalteu and aluminum chloride calorimetric assays, respectively. Rosmarinic acid (RA), the main compound found in the studied extracts, was quantified by liquid chromatography-photodiode array detector. Results showed that the cytokinin benzyladenine has an inhibitory effect on TPC, TFC and RAC, as well as on antioxidant activity. TPC was only significantly improved by the presence of YE (from 95.49 +/- 3.34 to 118.34 +/- 6.24 mg(GAE)/g(extract)) and TFCs were not affected by elicitation. The increase in sucrose concentration, from 2 to 4%, and the three elicitors tested, particularly YE, enhanced RACs (from 48.61 +/- 0.09 in the control medium to 78.57 +/- 0.99 mg/g(extract) in YE-elicited medium). In accordance with these results, the antioxidant activity measured by 2,2-diphenyl-1-picrylhydrazyl, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), and ferric reducing antioxidant power assays was significantly greater in extracts from elicited shoots. Overall, the results obtained suggest that shoots of T. lotocephalus are a good source of antioxidant compounds and showed that the production of RA can be promoted by altering in vitro culture conditions.
- Establishment of an in vitro propagation protocol for Thymus lotocephalus, a rare aromatic species of the Algarve (Portugal)Publication . Coelho, Natacha; Gonçalves, Sandra; González-Benito, María Elena; Romano, AnabelaThe aim of this work was to develop an in vitro propagation protocol for the endangered species Thymus lotocephalus using seedlings as explants. Several macronutrient concentrations of Murashige and Skoog medium (MS), cytokinin types and concentrations, and cytokinin/auxin combinations were tested to assess the shoots’ proliferation capacity. Although the best proliferation results were obtained with 6-benzyladenine, high percentages of hyperhidric shoots were observed. Because high proliferation of healthy shoots was observed in MS medium that was free of plant growth regulators, this medium was chosen for proliferation studies. The best rooting results were achieved in MS medium without auxins (92.00 ± 6.11%, 6.54 ± 0.52 and 1.60 ± 0.10 cm regarding rooting frequency, number of roots per shoot and longest roots, respectively) or supplemented with 0.5 mg l-1 indole-3-acetic acid (98.00 ± 2.11%, 11.14 ± 0.75 and 2.40 ± 0.24 cm, respectively). Plantlets were successfully acclimatised to ex vitro conditions with a survival rate of 93.33%. With the development of this micropropagation protocol, an important contribution has been made to the conservation of the endangered species T. lotocephalus.
- Estimation of genetic diversity in seedlings of Plantago algarbiensis, an endangered species endemic to the south of Portugal in risk of global extinctionPublication . Coelho, Natacha; Martin, Carmen; Gonzalez-Benito, Maria Elena; Romano, AnabelaPlantago algarbiensis samp is an endangered species endemic to the Algarve, in southern Portugal. In the present work, the genetic diversity of three populations was assessed by RAPD markers. Samples were amplified using ten primers that generated 145 markers, 80% of which were polymorphic. Tunes population presented the highest polymorphism percentage (73.68%) and Algoz the lowest (67.67%). In the cluster analysis, two major groups were formed, one including individuals from Gambelas and the other clustered together individuals from the other populations. The highest level of genetic diversity, estimated by both Nei's gene diversity and Shannon's information measure, was found in Tunes and the lowest in Algoz. From the estimates of Shannon's index, the proportion of the diversity within populations was 86.12% and that among populations was 13.88%. Similar results were obtained by AMOVA analysis. A correlation was found between geographic distance and genetic differentiation among populations. The N-m values obtained suggest a high level of gene flow among populations, which was inversely proportional to the distance between populations. RAPDs proved to be a useful tool to determine the genetic diversity of P. algarbiensis populations. The data obtained can be used to develop effective conservation strategies to prevent the decline of populations.
- Evaluation of the antioxidant and antimicrobial properties of in vitro cultured Drosera intermedia extractsPublication . Grevenstuk, Tomás; Gonçalves, Sandra; Almeida, Sara; Coelho, Natacha; Quintas, Célia; Gaspar, Maria Nelma; Romano, AnabelaEvaluation of the antioxidant activity of the methanol, water and n-hexane extracts of Drosera intermedia, determined by the Folin-Ciocalteau (F-C), trolox equivalent antioxidant capacity (TEAC) and oxygen radical antioxidant capacity (ORAC) assays showed that the methanol extract had the highest antioxidant activity (F-C: 378.6 ± 31.5 μmolGAE/mgextract; TEAC: 332.2 ± 29.1 μmolTE/mgextract; ORAC: 64.7 ± 7.8 μmolTE/mgextract). Antimicrobial activity was tested against seven bacterial and eight yeast strains using the agar diffusion assay, followed by the determination of minimum inhibitory concentrations (MIC). All tested D. intermedia extracts demonstrated strong antimicrobial properties with a broad spectrum of activity. However, the n-hexane extract exhibited much greater activity than water and methanol extracts. The most susceptible microorganisms to the n-hexane extract were Staphylococcus epidermidis ATCC 12228 and Candida albicans YP0175, for which a MIC value of 13.0 μg/mL was scored.
- From Lignocellulosic residues to protein sources: insights into biomass pre-treatments and conversionPublication . Vera dos Anjos, Isabela; Coelho, Natacha; Duarte, Hugo; Proença, Diogo Neves; Duarte, Maria F.; Jorge de Barros, Raúl José; Raposo, Sara; Gonçalves, Sandra; Romano, Anabela; Medronho, BrunoWith the global population steadily rising, the demand for sustainable protein sources has become increasingly urgent. Traditional animal- and plant-based proteins face challenges related to scalability, resource efficiency, and environmental impact. In this context, single-cell protein has emerged as a promising alternative. Derived from microorganisms such as algae, bacteria, fungi, and yeast, single-cell protein offers a high nutritional profileincluding all essential amino acids and vitamins—while enabling rapid production, minimal land and water requirements, and no generation of greenhouse gas emissions. A particularly compelling advantage of single-cell protein is its ability to be produced from agro-industrial waste, converting low-cost residues into valuable nutritional resources and contributing to environmental sustainability. Among these waste streams, lignocellulosic biomass from agricultural and forestry residues stands out as a renewable, biodegradable, and abundant feedstock. This review explores the potential of lignocellulosic waste as a substrate for single-cell protein production, emphasizing both its environmental advantages and nutritional value. It highlights the single-cell protein role as a sustainable and scalable alternative to conventional protein sources. The review also identifies key scientific, economic, and regulatory challenges, and recognizes the importance of targeted investments, particularly in policy development, public awareness, and technological innovation, to enable the broader adoption and acceptance of single-cell protein -based products.
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