UV Protective, Antioxidant, Antibacterial and Compostable Polylactic Acid Composites Containing Pristine and Chemically Modified Lignin Nanoparticles.

Polylactic acid (PLA) films containing 1 wt % and 3 wt % of lignin nanoparticles (pristine (LNP), chemically modified with citric acid (caLNP) and acetylated (aLNP)) were prepared by extrusion and characterized in terms of their overall performance as food packaging materials. Morphological, mechanical, thermal, UV-Vis barrier, antioxidant and antibacterial properties were assayed; appropriate migration values in food simulants and disintegration in simulated composting conditions were also verified. The results obtained indicated that all lignin nanoparticles succeeded in conferring UV-blocking, antioxidant and antibacterial properties to the PLA films, especially at the higher filler loadings assayed. Chemical modification of the fillers partially reduced the UV protection and the antioxidant properties of the resulting composites, but it induced better nanoparticles dispersion, reduced aggregates size, enhanced ductility and improved aesthetic quality of the films through reduction of the characteristic dark color of lignin. Migration tests and disintegration assays of the nanocomposites in simulated composting conditions indicated that, irrespectively of their formulation, the multifunctional nanocomposite films prepared behaved similarly to neat PLA.

Yarrowia lipolytica: a model yeast for citric acid production.

Every year more than 2 million tons of citric acid (CA) are produced around the world for industrial uses. Although initially extracted from citrus, the low profitability of the process and the increasing demand soon stimulated the search for more efficient methods to produce CA. Currently, most world CA demand (99%) is satisfied by fermentations with microorganisms, especially filamentous fungi and yeasts. CA production with yeasts has certain advantages over molds (e.g. higher productivity and easier cultivation), which in the last two decades have triggered a clear increase in publications and patents devoted to the use of yeasts in this field. Yarrowia lipolytica has become a model yeast that proved to be successful in different production systems. Considering the current interest evidenced in the literature, the most significant information on CA production using Y. lipolytica is summarized. The relevance on CA yields of key factors such as strains, media formulation, environmental conditions and production regimes is thoroughly discussed, with particular focus on increasing CA productivity. Besides, the possibility of tuning the mentioned variables to reduce concomitant isocitric acid production-the biggest disadvantage of using yeasts-is analyzed. Available methods for CA purification/quantification are also discussed.

Acetylation of bacterial cellulose catalyzed by citric acid: Use of reaction conditions for tailoring the esterification extent.

Bacterial cellulose (BC) nanoribbons were partially acetylated by a simple direct solvent-free route catalyzed by citric acid. The assay of reaction conditions within chosen intervals (i.e. esterification time (0.5-7h), catalyst content (0.08-1.01mmol/mmol AGU), and temperature (90-140°C)), illustrated the flexibility of the methodology proposed, with reaction variables which can be conveniently manipulated to acetylate BC to the required degree of substitution (DS) within the 0.20-0.73 interval. Within this DS interval, characterization results indicated a surface-only process in which acetylated bacterial cellulose with tunable DS, preserved fibrous structure and increased hydrophobicity could be easily obtained. The feasibility of reusing the catalyst/excess acylant in view of potential scale-up was also illustrated.

Use of grape pomaces to produce biomass of aKomagataella pastoris strain expressing a bovine chymosin activity.

The use of agroindustrial wastes not only decreases bioprocesses and disposal costs but also contributes to the upgrading of the residues. An active recombinant methanol-inducible bovine chymosin has been expressed in our laboratory in the yeastKomagataella pastoris, and grape pomace extracts (GRE) were proposed as a convenient C-energy source for the biomass production of the genetically engineered strain. Carbon and nitrogen sources, growth factors, and initial pH conditions were selected by classical methodology; thereafter, growth conditions optimization was performed using statistical designed experiments (DoEs). In the presence of (in g·L(-1)) 67.0 monosaccharides (glucose and fructose) from GRE, 5.0 (NH4)2SO4, and 10.0 sugar cane molasses (CMz), a yield of 20.0 g·L(-1) cell dry weight (CDW) was obtained aerobically after 60 h incubation at 28°C and pH 4.0. Applying a fed-batch strategy with methanol:sorbitol as the enzyme inducers, a chymosin production of 8.53 International Milk Clotting Units (IMCU) per mg protein was obtained in the supernatant. The results presented show that through a statistical design, a simple, cheap, and easy to prepare culture medium could be developed using two agroindustrial derivatives (GRE and CMz) to obtain a higher value added product.

Surface esterification of cellulose nanofibers by a simple organocatalytic methodology.

Bacterial cellulose nanofibers were esterified with two short carboxylic acids by means of a simple and novel organic acid-catalyzed route. The methodology proposed relayed on the use of a non-toxic biobased α-hydroxycarboxylic acid as catalyst, and proceeded under moderate reaction conditions in solventless medium. By varying the esterification interval, acetylated and propionized bacterial cellulose nanofibers with degree of substitution (DS) in the 0.02-0.45 range could be obtained. Esterified bacterial cellulose samples were characterized by means of Solid-State CP/MAS (13)C Nuclear Magnetic Resonance spectroscopy (CP/MAS (13)C NMR), Fourier Transform Infrared spectroscopy (FTIR), X-ray Diffraction (XRD), Thermogravimetric Analysis (TGA) and chosen hydrophobicity test assays. TGA results showed that the esterified nanofibers had increased thermal stability, whereas XRD data evidenced that the organocatalytic esterification protocol did not alter their crystallinity. The analysis of the ensuing modified nanofibers by NMR, FTIR, XRD and TGA demonstrated that esterification occurred essentially at the surface of bacterial cellulose microfibrils, something highly desirable for changing their surface hydrophilicity while not affecting their ultrastructure.

Optimization of biomass production of a mutant of Yarrowia lipolytica with an increased lipase activity using raw glycerol

The yeast Yarrowia lipolytica accumulates oils and is able to produce extracellular lipases when growing in different carbon sources including glycerol, the principal by-product of the biodiesel industry. In this study, biomass production of a novel mutant strain of Y. lipolytica was statistically optimized by Response Surface Methodology in media containing biodiesel-derived glycerol as main carbon source. This strain exhibited distinctive morphological and fatty acid profile characteristics, and showed an increased extracellular lipase activity. An organic source of nitrogen and the addition of 1.0 g/l olive oil were necessary for significant lipase production. Plackett-Burman and Central Composite Statistical Designs were employed for screening and optimization of fermentation in shaken flasks cultures, and the maximum values obtained were 16.1 g/l for biomass and 12.2 Units/ml for lipase, respectively. Optimized batch bioprocess was thereafter scaled in aerated bioreactors and the values reached for lipase specific activity after 95 % of the glycerol had been consumed, were three-fold higher than those obtained in shaken flasks cultures. A sustainable bioprocess to obtain biomass and extracellular lipase activity was attained by maximizing the use of the by-products of biodiesel industry.

Optimización de la producción de biomasa usando glicerol crudo, de una cepa mutante de Yarrowia lipolytica con actividad incrementada de lipasa. La levadura Yarrowia lipolytica acumula aceites y produce una lipasa extracelular al crecer en diferentes fuentes de carbono, entre ellas el glicerol, principal subproducto de la creciente industria del biodiésel. En el presente trabajo, se optimizó mediante la metodología de superficies de respuesta la producción de biomasa de una nueva cepa mutante de Y. lipolytica, empleando medios con glicerol derivado de la industria del biodiésel como principal fuente de carbono. Esta cepa presentó características morfológicas y perfil de ácidos grasos distintivos, y una mayor actividad de lipasa extracelular. Para obtener una producción significativa de lipasa extracelular, fue necesario el agregado de una fuente orgánica de nitrógeno y de 1 g/l de aceite de oliva. Se utilizaron los diseños estadísticos de Plackett-Burman y central compuesto para la selección y la optimización de las fermentaciones en frascos agitados; los máximos valores de biomasa y de lipasa obtenidos fueron de 16,1 g/l y 12,2 unidades/ml, respectivamente. Luego, el bioproceso en lote optimizado se escaló a biorreactores aireados, y los valores de actividad específica de lipasa alcanzados después de haberse consumido el 95 % del glicerol fueron tres veces más altos que los obtenidos en los cultivos en frascos agitados. En suma, se desarrolló un bioproceso sostenible para la obtención de biomasa y de una actividad de lipasa extracelular, que a la vez maximiza el uso de subproductos de la industria del biodiésel.

Optimization of biomass production of a mutant of Yarrowia lipolytica with an increased lipase activity using raw glycerol.

The yeast Yarrowia lipolytica accumulates oils and is able to produce extracellular lipases when growing in different carbon sources including glycerol, the principal by-product of the biodiesel industry. In this study, biomass production of a novel mutant strain of Y. lipolytica was statistically optimized by Response Surface Methodology in media containing biodiesel-derived glycerol as main carbon source. This strain exhibited distinctive morphological and fatty acid profile characteristics, and showed an increased extracellular lipase activity. An organic source of nitrogen and the addition of 1.0 g/l olive oil were necessary for significant lipase production. Plackett-Burman and Central Composite Statistical Designs were employed for screening and optimization of fermentation in shaken flasks cultures, and the maximum values obtained were 16.1 g/l for biomass and 12.2 Units/ml for lipase, respectively. Optimized batch bioprocess was thereafter scaled in aerated bioreactors and the values reached for lipase specific activity after 95 % of the glycerol had been consumed, were three-fold higher than those obtained in shaken flasks cultures. A sustainable bioprocess to obtain biomass and extracellular lipase activity was attained by maximizing the use of the by-products of biodiesel industry.

[Studies of viability and vitality after freezing of the probiotic yeast Saccharomyces boulardii: physiological preconditioning effect]. / Estudios de la viabilidad y la vitalidad frente al congelado de la levadura probiótica Saccharomyces boulardii: efecto del preacondicionamiento fisiológico.

The aim of this study was to evaluate the vitality and viability of the probiotic yeast Saccharomyces boulardii after freezing/thawing and the physiological preconditioning effect on these properties. The results indicate that the specific growth rate (0.3/h(-1)) and biomass (2-3 x10(8)cells/ml) of S. boulardii obtained in flasks shaken at 28 degrees C and at 37 degrees C were similar. Batch cultures of the yeast in bioreactors using glucose or sugar-cane molasses as carbon sources, reached yields of 0.28 g biomass/g sugar consumed, after 10h incubation at 28 degrees C; the same results were obtained in fed batch fermentations. On the other hand, in batch cultures, the vitality of cells recovered during the exponential growth phase was greater than the vitality of cells from the stationary phase of growth. Vitality of cells from fed-batch fermentations was similar to that of stationary growing cells from batch fermentations. Survival to freezing at -20 degrees C and subsequent thawing of cells from batch cultures was 0.31% for cells in exponential phase of growth and 11.5% for cells in stationary phase. Pre-treatment of this yeast in media with water activity (a(w)) 0.98 increased the survival to freezing of S. boulardii cells stored at -20 degrees C for 2 months by 10 fold. Exposure of the yeast to media of reduced a(w) and/or freezing/thawing process negatively affected cell vitality. It was concluded that stress conditions studied herein decrease vitality of S. boulardii. Besides, the yeast strain studied presented good tolerance to bile salts even at low pH values.

Estudios de la viabilidad y la vitalidad frente al congelado de la levadura probiótica Saccharomyces boulardii: efecto del preacondicionamiento fisiológico / Studies of viability and vitality after freezing of the probiotic yeast Saccharomyces boulardii: physiological preconditioning effect

El objetivo de este estudio fue evaluar la vitalidad y la viabilidad de la levadura probiótica Saccharomycesboulardii después de su congelación y descongelación, y el efecto del preacondicionamiento fisiológico sobredichas propiedades. Los resultados indican que la velocidad específica de crecimiento (0,3 h–1) y la biomasa(2-3 × 108 células/ml) de S. boulardii obtenida en frascos agitados tanto a 28 como a 37 °C fueron semejantes.El cultivo de esta levadura en biorreactores en lote, utilizando glucosa o melaza de caña de azúcar como fuentede carbono, alcanzó rendimientos de 0,28 g de biomasa/g azúcar consumida tras 10 h de cultivo a 28 °C,obteniéndose resultados similares en fermentaciones en lote alimentado. Además, en los cultivos en lote, lavitalidad de las células en fase de crecimiento exponencial fue mayor que la de las células en fase estacionaria.En cambio, en el lote alimentado, la vitalidad de las células fue semejante a la del lote en fase estacionaria.La supervivencia a la congelación a –20 °C y posterior descongelación de las células de fermentaciones en loteen fase de crecimiento exponencial fue del 0,31% y la de fase estacionaria del 11,5%. El pretratamiento de estalevadura en medios de actividad de agua (aw) 0,98 aumentó 10 veces la supervivencia al congelado de las célulasalmacenadas a –20 °C durante 2 meses. La exposición de las levaduras a medios de aw reducida y/o elproceso de congelación/descongelación afectaron negativamente la vitalidad celular. Se concluye que condicionesde estrés como las estudiadas en este trabajo disminuyen la vitalidad de S. boulardii. Además, la cepaestudiada presentó buena tolerancia a las sales biliares aun a bajos valores de pH del cultivo(AU)

The aim of this study was to evaluate the vitality and viability of the probiotic yeast Saccharomyces boulardiiafter freezing/thawing and the physiological preconditioning effect on these properties. The results indicatethat the specific growth rate (0.3/h–1) and biomass (2-3 × 108 cells/ml) of S. boulardii obtained in flasks shakenat 28 °C and at 37 °C were similar. Batch cultures of the yeast in bioreactors using glucose or sugar-canemolasses as carbon sources, reached yields of 0.28 g biomass/g sugar consumed, after 10 h incubation at 28 °C;the same results were obtained in fed batch fermentations. On the other hand, in batch cultures, the vitality ofcells recovered during the exponential growth phase was greater than the vitality of cells from the stationaryphase of growth. Vitality of cells from fed-batch fermentations was similar to that of stationary growing cellsfrom batch fermentations. Survival to freezing at –20 °C and subsequent thawing of cells from batch cultureswas 0.31% for cells in exponential phase of growth and 11.5% for cells in stationary phase. Pre-treatment ofthis yeast in media with water activity (aw) 0.98 increased the survival to freezing of S. boulardii cells stored at–20 °C for 2 months by 10 fold. Exposure of the yeast to media of reduced aw and/or freezing/thawing processnegatively affected cell vitality. It was concluded that stress conditions studied herein decrease vitality ofS. boulardii. Besides, the yeast strain studied presented good tolerance to bile salts even at low pH values(AU)

Selective antimicrobial activity of chitosan on beer spoilage bacteria and brewing yeasts.

Chitosan (0.1 g l(-1)), assayed in a simple medium, reduced the viability of four lactic acid bacteria isolated during the beer production process by 5 logarithmic cycles, whereas activity against seven commercial brewing yeasts required up to 1 g chitosan l(-1). Antimicrobial activity was inversely affected by the pH of the assay medium. In brewery wort, chitosan (0.1 g l(-1)) selectively inhibited bacterial growth without altering yeast viability or fermenting performance.