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Dive into the research topics where Rosa León is active.

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Featured researches published by Rosa León.


Enzyme and Microbial Technology | 1998

Whole-cell biocatalysis in organic media

Rosa León; Pedro Fernandes; H.M. Pinheiro; J. M. S. Cabral

Abstract The use of water-immiscible organic solvents in whole-cell biocatalysis has been exploited for biotransformations involving sparingly water-soluble or toxic compounds. These systems can overcome the problem of low productivity levels in conventional media due to poor substrate solubility, integrate bioconversion and product recovery in a single reactor, and shift chemical equilibria enhancing yields and selectivities; nevertheless, the selection of a solvent combining adequate physicochemical properties with biocompatibility is a difficult task. The cell membrane seems to be the primary target of solvent action and the modification of its characteristics the more relevant cellular adaptation mechanism to organic solvent-caused stress. Correlations between the cellular toxicity or the extractive capacities of different solvents and some of their physical properties have been proposed in order to minimize preliminary, solvent-selection experimental work but also to help in the understanding of the molecular mechanisms of toxicity and extraction. The use of whole cells in organic-media biocatalysis provides a way to regenerate cofactors and carry out bioconversions or fermentations requiring multi-step metabolic pathways; some processes already are commercially exploited. Immobilization can further protect cells from solvent toxicity, and has thus been effectively used in organic solvent-based systems. Several examples of extractive fermentations and other whole-cell bioconversions in organic media are presented.


Marine Drugs | 2011

Enhancement of Lutein Production in Chlorella sorokiniana (Chorophyta) by Improvement of Culture Conditions and Random Mutagenesis

Baldo F. Cordero; Irina Obraztsova; Inmaculada Couso; Rosa León; M.A. Vargas; Herminia Rodríguez

Chlorella sorokiniana has been selected for lutein production, after a screening of thirteen species of microalgae, since it showed both a high content in this carotenoid and a high growth rate. The effects of several nutritional and environmental factors on cell growth and lutein accumulation have been studied. Maximal specific growth rate and lutein content were attained at 690 μmol photons m−2 s−1, 28 °C, 2 mM NaCl, 40 mM nitrate and under mixotrophic conditions. In general, optimal conditions for the growth of this strain also lead to maximal lutein productivity. High lutein yielding mutants of C. sorokiniana have been obtained by random mutagenesis, using N-methyl-N′-nitro-nitrosoguanidine (MNNG) as a mutagen and selecting mutants by their resistance to the inhibitors of the carotenogenic pathway nicotine and norflurazon. Among the mutants resistant to the herbicides, those exhibiting both high content in lutein and high growth rate were chosen. Several mutants exhibited higher contents in this carotenoid than the wild type, showing, in addition, either a similar or higher growth rate than the latter strain. The mutant MR-16 exhibited a 2.0-fold higher volumetric lutein content than that of the wild type, attaining values of 42.0 mg L−1 and mutants DMR-5 and DMR-8 attained a lutein cellular content of 7.0 mg g−1 dry weight. The high lutein yield exhibited by C. sorokiniana makes this microalga an excellent candidate for the production of this commercially interesting pigment.


Applied Microbiology and Biotechnology | 2011

Enhancement of carotenoids biosynthesis in Chlamydomonas reinhardtii by nuclear transformation using a phytoene synthase gene isolated from Chlorella zofingiensis

Baldo F. Cordero; Inmaculada Couso; Rosa León; Herminia Rodríguez; M. Angeles Vargas

The isolation and characterization of the phytoene synthase gene from the green microalga Chlorella zofingiensis (CzPSY), involved in the first step of the carotenoids biosynthetic pathway, have been performed. CzPSY gene encodes a polypeptide of 420 amino acids. A single copy of CzPSY has been found in C. zofingiensis by Southern blot analysis. Heterologous genetic complementation in Escherichia coli showed the ability of the predicted protein to catalyze the condensation of two molecules of geranylgeranyl pyrophosphate (GGPP) to form phytoene. Phylogenetic analysis has shown that the deduced protein forms a cluster with the rest of the phytoene synthases (PSY) of the chlorophycean microalgae studied, being very closely related to PSY of plants. This new isolated gene has been adequately inserted in a vector and expressed in Chlamydomonas reinhardtii. The overexpression of CzPSY in C. reinhardtii, by nuclear transformation, has led to an increase in the corresponding CzPSY transcript level as well as in the content of the carotenoids violaxanthin and lutein which were 2.0- and 2.2-fold higher than in untransformed cells. This is an example of manipulation of the carotenogenic pathway in eukaryotic microalgae, which can open up the possibility of enhancing the productivity of commercial carotenoids by molecular engineering.


Biomolecular Engineering | 2003

Carotenoids as protective response against oxidative damage in Dunaliella bardawil

Alonso Salguero; Benito de la Morena; Javier Vigara; José M. Vega; Carlos Vílchez; Rosa León

In the present work the relation between carotenoids production and cell response mechanisms to oxidative damage was studied. High light intensity and nitrogen starvation, both conditions, which may increase the oxidative damage in microalgae, significantly increased total carotenoids content in Dunaliella bardawil, the effect of N-starvation being more noticeable when acting synergetically with light on carotenoid production. S-starvation stimulated carotenoids production as much as N-starvation. The use of norflurazon, inhibitor of phytoene desaturase that blocks formation of epsilon-carotene from phytoene, caused a decrease of carotenoid content down to 5% that of the control cells incubated without the inhibitor. The decrease in the oxygen consumption rate of D. bardawil cells exposed to norflurazon suggests a connection between carotenoids desaturation and chloroplastic oxygen species dissipation processes reported in the literature for other algae. It is an indication of the carotenoids involvement in chloroplastic response mechanisms to oxidative damage.


Biomolecular Engineering | 2003

Microalgae mediated photoproduction of β-carotene in aqueous–organic two phase systems

Rosa León; Marta Martı́n; Javier Vigara; Carlos Vílchez; José M. Vega

Improving productivity is a usual requirement for most biotechnological processes, and the utilisation of two-phase aqueous organic systems has proved to be an effective way to improve the productivity of poorly water-soluble or toxic compounds. The high hydrophobicity of beta-carotene, which is highly demanded by the pharma and agrofood industry, makes it a good candidate for aqueous/organic biphasic photoproduction. In the present work we have investigated the viability of a two-phase system for the production of beta-carotene by the marine microalgae Dunaliella salina using decane as organic phase. Decane, with a logP(octanol) value of 5.6, showed no toxicity to Dunaliella cells for more than 72 h, and its ability for beta-carotene extraction is acceptable. Transferring Dunaliella cells from standard to carotenogenic conditions caused inhibition of chlorophyll production and induced a strong synthesis of beta-carotene. The two-phase aqueous/decane system was stable and beta-carotene content of the cells was increasing during 4-days. About 8% of the total carotenoids produced were excreted and extracted into the decane phase.


Advances in Experimental Medicine and Biology | 2007

Nuclear Transformation of Eukaryotic Microalgae Historical Overview, Achievements and Problems

Rosa León; Emilio Fernández

Transformation of microalgae is a first step in their use for biotechnological applications involving foreign protein production or molecular modifications of specific cell metabolic pathways. Since the first reliable achievements of nuclear transformation in Chlamydomonas, other eukaryotic microalgae have become transformed with molecular markers that allow a direct selection. Different methods--glass beads, electroporation, particle bombardment, or Agrobacterium--and constructions have been set up in several organisms and successfully used. However, some problems associated with efficiency, integration, or stability of the transgenes still persist and are analysed herein. Though the number of microalgae species successfully transformed is not very high, prospects for transformation of many more are good enough on the basis of what has been achieved so far.


Bioresource Technology | 2001

STUDIES ON THE SUITABILITY OF ALGINATE-ENTRAPPED CHLAMYDOMONAS REINHARDTII CELLS FOR SUSTAINING NITRATE CONSUMPTION PROCESSES

Carlos Vílchez; Inés Garbayo; Elena Markvicheva; Francisco Galván; Rosa León

Some aspects of the suitability of alginate beads entrapping Chlamydomonas reinhardtii cells for nitrate consumption from nitrate-containing waters were studied and discussed. Among 14 different metal cations tested as gel bead stabilizing agents, only calcium and barium formed beads showing nitrate-consuming activity. Pure calcium alginate cell entrapment resulted in the most suitable method for active cell immobilization compared to alginate-composite-gel beads based on poly-vinylcaprolactam (PVCL) and poly-vinylpyrrolidone (PVP). To perform a continuous nitrate consumption process, calcium alginate-entrapped cells were first grown in a 2.5 l airlift-loop reactor. A cell loading of about 150 microg Chl. g(-1) gel was achieved. Afterwards, five days nitrate consumption processes were performed and three different dilution rates were applied: (i) D < mu; (ii) D = mu; (iii) D > mu, where mu is the specific growth rate (h(-1)). The maximum consumption rates calculated for each dilution rate were: (i) 3.8, (ii) 6.4 and (iii) 7.2 mg nitrate mg(-1) Chl. h(-1). For low dilution rates (D < mu) some nitrite (< 300 microM) was excreted into the culture medium. However, this concentration of nitrite was not high enough to inhibit nitrate consumption.


Enzyme and Microbial Technology | 1999

Biotransformations in two-liquid-phase systems: Production of phenylacetaldehyde by oxidation of 2-phenylethanol with acetic acid bacteria

Francesco Molinari; Raffaella Gandolfi; Fabrizio Aragozzini; Rosa León; D.M.F. Prazeres

Abstract Phenylacetaldehyde can be obtained by oxidation of 2-phenylethanol with acetic acid bacteria in two-liquid-phase systems where the aldehyde is removed into the organic phase before its further conversion to acid. Two Acetobacter strains (ALEF and ALEG) were able to accumulate aldehyde when aliphatic hydrocarbons were used. A two-liquid-phase system, composed of water and isooctane (v/v, 1/1), was particularly suited for a significant accumulation of the aldehyde: Acetobacter sp. ALEG furnished 9 g/l of phenylacetaldehyde within 4 h starting from 10 g/l of alcohol and still 8 g/l were recovered after 24 h in the organic phase, whereas strain ALEF gave 3.5 g/l of aldehyde from 5.0 g/l of substrate. Acetobacter sp. ALEG also showed satisfactory long-term stability, being able to perform the transformation with 80% of the original activity after 3 days of contact with the solvent.


Photosynthesis Research | 2015

Production of carotenoids by microalgae: achievements and challenges

J. Varela; Hugo Pereira; Marta Vila; Rosa León

Carotenoids are a wide group of lipophylic isoprenoids synthesized by all photosynthetic organisms and also by some non-photosynthetic bacteria and fungi. Animals, which cannot synthesize carotenoids de novo, must include them in their diet to fulfil essential provitamin, antioxidant, or colouring requirements. Carotenoids are indispensable in light harvesting and energy transfer during photosynthesis and in the protection of the photosynthetic apparatus against photooxidative damage. In this review, we outline the factors inducing carotenoid accumulation in microalgae, the knowledge acquired on the metabolic pathways responsible for their biosynthesis, and the recent achievements in the genetic engineering of this pathway. Despite the considerable progress achieved in understanding and engineering algal carotenogenesis, many aspects remain to be elucidated. The increasing number of sequenced microalgal genomes and the data generated by high-throughput technologies will enable a better understanding of carotenoid biosynthesis in microalgae. Moreover, the growing number of industrial microalgal species genetically modified will allow the production of novel strains with enhanced carotenoid contents.


Biotechnology Progress | 2011

Overexpression of an exogenous phytoene synthase gene in the unicellular alga Chlamydomonas reinhardtii leads to an increase in the content of carotenoids.

Inmaculada Couso; Marta Vila; Herminia Rodríguez; M.A. Vargas; Rosa León

Phytoene synthase (PSY) catalyses the first step in the production of carotenoids, which has been described as a key regulatory step in the carotenoids biosynthetic pathway. PSY gene from Dunaliella salina was constitutively expressed in Chlamydomonas reinhardtii under the control of the RBCS2 and HSP70A promoters and targeted to the chloroplast by the RBCS2 transit peptide. DsPSY overexpression resulted in a stable increase in the corresponding PSY transcript level and in the content of carotenoids such as violaxanthin, lutein, and β‐carotene, reaching between 125 and 260% the levels in control untransformed cells.

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Inmaculada Couso

Spanish National Research Council

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Herminia Rodríguez

Spanish National Research Council

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Baldo F. Cordero

Spanish National Research Council

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