Daniela Vieira Cortez

Individual and interaction effects of vanillin and syringaldehyde on the xylitol formation by Candida guilliermondii

The effect of lignin degradation products liberated during chemical hydrolysis of lignocellulosic materials on xylose-to-xylitol bioconversion by Candida guilliermondii FTI 20037 was studied. Two aromatic aldehydes (vanillin and syringaldehyde) were selected as model compounds. A two-level factorial design was employed to evaluate the effects of pH (5.5-7.0), cell concentration (1.0-3.0 g l(-1)), vanillin concentration (0-2.0 g l(-1)) and syringaldehyde concentration (0-2.0 g l(-1)) on this bioprocess. The results showed that in the presence of vanillin or syringaldehyde (up to 2.0 g l(-1)) the cell growth was inhibited to different degrees with a complete inhibition of the yeast growth when the mixture of both (at 2.0 g l(-1) each) was added to the fermentation medium. The xylitol yield was not significantly influenced by vanillin, but was strongly reduced by syringaldehyde, which showed a more pronounced inhibitor effect at pH 7.0. The yeast was also able to convert vanillin and syringaldehyde to the corresponding aromatic acids or alcohols and their formation was dependent of the experimental conditions employed.

Optimization of β-xylosidase recovery by reversed micelles using response surface methodology

β-Xylosidase recovery by reversed micelles using CTAB cationic surfactant was performed under different experimental conditions. A 2 2 -full orthogonal design with center points was employed to optimize the recovery of this enzyme and to evaluate the influence of the factors CTAB (A) and butanol concentration (B) on the enzyme extraction. A mathematical model to represent the enzyme recovery as a function of A and B was proposed. According to the model, the CTAB and butanol concentrations necessary to attain the maximum β-xylosidase recovery (43%) were 0.26 M and 29.4%, respectively. The results showed that the recovery value predicted by the model was similar to that obtained experimentally (44.3%).

CTAB, Triton X-100 and freezing–thawing treatments of Candida guilliermondii: Effects on permeability and accessibility of the glucose-6-phosphate dehydrogenase, xylose reductase and xylitol dehydrogenase enzymes

Cells of Candida guilliermondii (ATCC 201935) were permeabilised with surfactant treatment (CTAB or Triton X-100) or a freezing-thawing procedure. Treatments were monitored by in situ activities of the key enzymes involved in xylose metabolism, that is, glucose-6-phosphate dehydrogenase (G6PD), xylose reductase (XR) and xylitol dehydrogenase (XD). The permeabilising ability of the surfactants was dependent on its concentration and incubation time. The optimum operation conditions for the permeabilisation of C. guilliermondii with surfactants were 0.41 mM (CTAB) or 2.78 mM (Triton X-100), 30°C, and pH 7 at 200 rpm for 50 min. The maximum permeabilisation measured in terms of the in situ G6PD activity observed was, in order, as follows: CTAB (122.4±15.7U/g(cells)) > freezing-thawing (54.3 ± 1.9U/g(cells))>Triton X-100 (23.5 ± 0.0U/g(cells)). These results suggest that CTAB surfactant is more effective in the permeabilisation of C. guilliermondii cells in comparison to the freezing-thawing and Triton X-100 treatments. Nevertheless, freezing-thawing was the only treatment that allowed measurable in situ XR activity. Therefore, freezing-thawing permeabilised yeast cells could be used as a source of xylose reductase for analytical purposes or for use in biotransformation process such as xylitol preparation from xylose. The level of in situ xylose reductase was found to be 13.2 ± 0.1 U/g(cells).

Optimization of D-xylose to xylitol biotransformation by Candida guilliermondii cells permeabilized with Triton X-100

Abstract The effect of NADP+ and glucose-6-phosphate (G6P) on the biotransformation of D-xylose to xylitol by cells of Candida guilliermondii permeabilized with surfactant Triton X-100 was evaluated. The experimental runs were performed with 12 g L−1 of permeabilized cells and a reaction medium composed of Tris–HCl buffer (0.1 M pH 7), D-xylose (57 g L−1), and MgCl2.6H2O (5 mM). The levels of NADP+ (from 0.0 to 1.7 mM) and G6P (from 0.00 to 0.17 M) were varied according a 22-full factorial composed design. Under optimized conditions (NADP+ 0.5 mM and 0.05 M G6P), the xylitol volumetric productivity (QP) and yield factor (YP/S) predicted were 1.86 ± 0.03 g L−1 h− 1 and 0.64 ± 0.03 g g−1, respectively. These values were 94% and 19% higher than those obtained with unpermeabilized cells under fermentation conditions (0.97 g L−1 h−1 and 0.53 g g−1, respectively). On the basis of the results, it can be concluded that xylitol production by biotransformation with cells of C. guilliermondii permeabilized with Triton X-100 is a promising alternative to the fermentative process.

Micelas reversas de lecitina de soja: uma alternativa para purificação de proteínas

No presente trabalho, estudaram-se os efeitos de diversos parâmetros sobre a extracao das proteinas caseina e albumina de soro bovino empregando micelas reversas de lecitina de soja. Independentemente da condicao empregada, a extracao da albumina apresentou baixo rendimento (variando de 0% a 4%, aproximadamente), resultado de um significativo efeito de exclusao por tamanho. Com relacao a caseina, o rendimento da extracao aumentou 23 vezes com o aumento do tempo de agitacao, ou seja, com o maior tempo de contato entre a proteina e o sistema de micelas reversas. A adicao de 1-hexanol ao sistema, usado como co-solvente, foi efetiva, aumentando a solubilizacao da caseina em 36%, sendo os rendimentos da extracao desta proteina muito influenciados pelo pH. Os valores maximos de eficiencia obtidos foram de 20% em pH 7,9, 80% em pH 5,4 e 100% em pH 5,0 (pH proximo ao pI da proteina).

The Realm of Lipases in Biodiesel Production

Lipases are the enzymes known for the hydrolytic activity on carboxylic fatty ester bonds. The industrial interest in lipases is due to their application in a wide array of products: in detergents and cleaning products, in pharmaceutical applications, in the food industry, and on the production of biodiesel. Biodiesel, i.e. short-chain-acyl fatty ester, is mainly produced via the transesterification of fatty-acyl glycerides or esterification of fatty acids, both reactions with a short chain alcohol. Lipases can catalyze both said reactions with high specificity, producing biodiesel at high yields at low temperature. With the significant advances in biodiesel production over the last decades, coupled with a strong industrial partnership, the costs of utilizing lipases as catalysts have dropped significantly. The production of lipases became popularized in the industry due to advances not only in the reaction mechanisms, and in better understanding of lipase-producing microorganisms, but to cost-effective utilization practices. Immobilization is the practice responsible for the initial breakthrough innovation that allowed efficient reutilization of lipases, thus reducing the cost per batch. There was, and still there is, numerous advances in the development of immobilizing matrices and novel utilization pathways of immobilized enzymes available in the literature. More recently, other methods of using lipase in biodiesel production have been developed, e.g. via the utilization of whole-cell and fermented solid with lypolytic activity, and by the use of lipase in liquid formulations. Over the last years, there has been an increased interest in developing next-generation biodiesel, i.e., the one produced from alternative lipid feedstock, such as microbial and residual lipids, and by utilizing ethanol as acyl agent, instead of methanol. There has also been prominent advances in the reactor engineering aspect of lipase-derived biodiesel, by promoting more efficient batch processes, and the development of lower-cost continuous processing. The present chapter reviews the recent literature in the important field of using lipases in biodiesel production, and critically describes the opportunities and challenges present in such applications.

Seleção de espécies do gênero Penicillium produtoras de lipase ligada ao micélio para aplicação em hidrólise de óleos vegetais

SCREENING OF SPECIES FROM THE GENUS Penicillium PRODUCING CELL BOUND LIPASES TO BE APPLIED IN THE VEGETABLE OIL HYDROLYSIS. Ten species of Penicillium genus isolated from different habitats were evaluated as mycelium bound lipase producers to be used in the hydrolysis of vegetable oils. Using olive oil as an inducer three species (P. italicum AT4421, P. janthinellum CCT3162 and P. purpurogenum AT2008) were able to produce lipases having high mycelium bound activities (>150 U g) and were further characterized in relation to their biochemical and kinetic properties and specificity using vegetable oils having majority fatty acids composition in C12:0 (coconut); C16:0 (palm); C18:1 (canola) and C18:1 (soybean). All the three lipases could enrich the medium with fatty acids according to their respective selectivity and the reaction hydrolysis was found to enhance at least three folds under ultrasonic irradiations. For P. purpurogenum lipase the highest hydrolysis degree (66.8 ± 0.2%) was attained with coconut oil. Both P. italicum and P. janthinellum lipases showed high selectivity for canola oil, resulting in hydrolysis degrees of 79.9 ± 0.5% and 63.5 ± 0.6%, respectively. Analysis of the hydrolysates confirmed that the majority of the fatty acids released by P. italicum and P. janthinellum lipases was composed by oleic acid, and P. purpurogenum lipase the hydrolysate contained approximately 50% of lauric acid.

Otimização do processo de extração de β-xilosidase por sistemas micelares reversos

β-xylosidase, produced by Penicillium janthinellum, was extracted by using the reversed micellar system of the CTAB (cetyl trimethyl ammonium bromide) cationic surfactant containing isooctane, hexanol and butanol. The combined effects of CTAB and butanol concentrations on enzyme extraction were studied by using the surface response methodology. As a function of the results, it was proposed a mathematical model to describe the process of β-xylosidase extraction. This model predicted a maximal enzyme extraction yield of 35.05±6.4% under the following conditions: pH = 8.0, CTAB = 0.20 M, hexanol = 5% (v/v), and butanol = 20% (v/v). Experimentally, a recovery value of about 38% was attained, and showed that the model is appropriate.