Dry extrusion pretreatment of cassava starch aided by sugarcane bagasse for improved starch saccharification.

The enzymatic hydrolysis of native starch lacks efficiency because starch is mostly confined in semi-crystalline granules. To address the challenges associated with gelatinization and render native cassava starch (CS) amenable to enzymatic hydrolysis (enzyme cocktail from Aspergillus awamori and Trichoderma reesei), dry-extrusion pretreatment of CS mixed with sugarcane bagasse (SB) was studied. Results showed that among the CS:SB mass ratios studied (1:1; 1:0.5 and 1:0.25), extruded CS:SB (1:0.25) gave the highest 3-hour glucose yield (71.5%) after enzymatic hydrolysis. Extrusion reduced CS:SB (1:0.25) crystallinity by 78% and increased the intensity of all major FTIR absorption bands by 67-202%. The optimum 3-hour glucose yield from extruded CS:SB (1:0.25) hydrolysis was 74.1%, which was 330% higher than from untreated CS. The water absorption and solubility indices of the treated biomass increased by 145% and 12,640%, respectively under the optimum conditions, aiding the hydrolysis process. The dry extrudates were easy to manipulate and store.

Comparative performance and reusability studies of lipases on syntheses of octyl esters with an economic approach.

A suitable immobilized lipase for esters syntheses should be selected considering not only its cost. We evaluated five biocatalysts in syntheses of octyl caprylate, octyl caprate, and octyl laurate, in which conversions higher than 90% were achieved. Novozym®ï»¿ 435 and non-commercial preparations (including a dry fermented solid) were selected for short-term octyl laurate syntheses using different biocatalysts loadings. By increasing the biocatalyst's loading the lipase's reusability also raised, but without strict proportionality, which resulted in a convergence between the lowest biocatalyst loading and the lowest cost per batch. The use of a dry fermented solid was cost-effective, even using loadings as high as 20.0% wt/wt due to its low obtaining cost, although exhibiting low productiveness. The combination of biocatalyst's cost, esterification activity, stability, and reusability represents proper criteria for the choice. This kind of assessment may help to establish quantitative goals to improve or to develop new biocatalysts.

Use of lignocellulose biomass for endoxylanase production by Streptomyces termitum.

Actinobacteria isolates from Brazilian Cerrado soil were evaluated for their ability to produce enzymes of the cellulolytic and xylanolytic complex using lignocellulose residual biomass. Preliminary semiquantitative tests, made in Petri plates containing carboxymethylcellulose and beechwood xylan, indicated 11 potential species producing enzymes, all belonging to the genus Streptomyces. The species were subsequently grown in pure substrates in submerged fermentation and analyzed for the production of enzymes endoglucanase, ß-glucosidase, endoxylanase, and ß-xylosidase. The best results were obtained for endoxylanase enzyme production with Streptomyces termitum(UFLA CES 93). The strain was grown on lignocellulose biomass (bagasse, straw sugarcane, and cocoa pod husk) that was used in natura or acid pretreated. The medium containing sugarcane bagasse in natura favored the production of the endoxylanase that was subsequently optimized through an experimental model. The highest enzyme production 0.387 U mL-1, (25.8 times higher), compared to the lowest value obtained in one of the trials, was observed when combining 2.75% sugar cane bagasse and 1.0 g L-1 of yeast extract to the alkaline medium (pH 9.7). This is the first study using S. termitum as a producer of endoxylanase.

Optimization of biohydrogen yield produced by bacterial consortia using residual glycerin from biodiesel production.

The aims of this study were to simplify the fermentation medium and to optimize the conditions of dark fermentation of residual glycerin to produce biohydrogen. It was possible to remove all micronutrients of fermentation medium and improve biohydrogen production by applying residual glycerin as feedstock. After statistical analysis of the following parameters pH, glycerin concentration and volatile suspended solids, the values of 5.5; 0.5g.L(-1) and 8.7g.L(-1), respectively, were defined as optimum condition for this process. It generated 2.44molH2/molglycerin, an expressive result when compared to previous results reported in literature and considering that theoretical yield of H2 from glycerol in dark fermentation process is 3molH2/molglycerol. This study allowed the improvement of yield and productivity by 68% and 67%, respectively.

Untreated Chlorella homosphaera biomass allows for high rates of cell wall glucan enzymatic hydrolysis when using exoglucanase-free cellulases.

BACKGROUND: Chlorophyte microalgae have a cell wall containing a large quantity of cellulose Iα with a triclinic unit cell hydrogen-bonding pattern that is more susceptible to hydrolysis than that of the cellulose Iß polymorphic form that is predominant in higher plants. This study addressed the enzymatic hydrolysis of untreated Chlorella homosphaera biomass using selected enzyme preparations, aiming to identify the relevant activity profile for the microalgae cellulose hydrolysis. Enzymes from Acremonium cellulolyticus, which secretes a complete pool of cellulases plus ß-glucosidase; Trichoderma reesei, which secretes a complete pool of cellulases with low ß-glucosidase; Aspergillus awamori, which secretes endoglucanases and ß-glucosidase; blends of T. reesei-A. awamori or A. awamori-A. cellulolyticus enzymes; and a purified A. awamori ß-glucosidase were evaluated. RESULTS: The highest initial glucan hydrolysis rate of 140.3 mg/g/h was observed for A. awamori enzymes with high ß-glucosidase, low endoglucanase, and negligible cellobiohydrolase activities. The initial hydrolysis rates when using A. cellulolyticus or T. reesei enzymes were significantly lower, whereas the results for the T. reesei-A. awamori and A. awamori-A. cellulolyticus blends were similar to that for the A. awamori enzymes. Thus, the hydrolysis of C. homosphaera cellulose was performed exclusively by the endoglucanase and ß-glucosidase activities. X-ray diffraction data showing negligible cellulose crystallinity for untreated C. homosphaera biomass corroborate these findings. The A. awamori-A. cellulolyticus blend showed the highest initial polysaccharide hydrolysis rate of 185.6 mg/g/h, as measured by glucose equivalent, in addition to the highest predicted maximum glucan hydrolysis yield of 47% of total glucose (w/w). T. reesei enzymes showed the lowest predicted maximum glucan hydrolysis yield of 25% (w/w), whereas the maximum yields of approximately 31% were observed for the other enzyme preparations. The hydrolysis yields were proportional to the enzyme ß-glucosidase load, indicating that the endoglucanase load was not rate-limiting. CONCLUSIONS: High rates of enzymatic hydrolysis were achieved for untreated C. homosphaera biomass with enzymes containing endoglucanase and ß-glucosidase activities and devoid of cellobiohydrolase activity. These findings simplify the complexity of the enzyme pools required for the enzymatic hydrolysis of microalgal biomass decreasing the enzyme cost for the production of microalgae-derived glucose syrups.

Amino acids interference on the quantification of reducing sugars by the 3,5-dinitrosalicylic acid assay mislead carbohydrase activity measurements.

This study evaluated the interference of the amino acids tryptophan, cysteine, histidine, tyrosine, hydroxyproline, leucine, proline, serine, glycine, valine, glutamic acid, phenylalanine, and methionine on the measurement of reducing sugars using a phenol-free 3,5-dinitrosalicylic acid (DNS) reagent. It was found that in reaction mixtures containing 20mM of either tryptophan, cysteine, histidine, tyrosine, or hydroxyproline the measurement of 3.7 mM glucose was overestimated by 76%, 50%, 35%, 18%, and 10%, respectively. The amino acids valine, glutamic acid, and phenylalanine did not affect the DNS reaction, while methionine decreased the color development by 5%. The measurement of glucose, xylose, arabinose, and cellobiose at the 3.7-12.4 mM range in the presence of 20 mM cysteine resulted in an overestimated concentration of 34.8-50%. Enzymatic assays for measuring xylanolytic and filter paper activity (FPAse) were conducted in the presence of 20-60 mM cysteine, and compared to cysteine-free assays. In the presence of cysteine, the measured xylanase activity increased threefold and the FPAse activity increased twofold due to the overestimation of the reducing sugar concentrations in the assays. The interference from cysteine was reduced to a maximum of 8.6% when a DNS reagent containing phenol was used.

The Use of HRP in Decolorization of Reactive Dyes and Toxicological Evaluation of Their Products.

This work studied the potential use of horseradish peroxidase (HRP) in the decolorization of the following textile dyes: Drimarene Blue X-3LR (DMBLR), Drimarene Blue X-BLN (DMBBLN), Drimarene Rubinol X-3LR (DMR), and Drimarene Blue CL-R (RBBR). Dyes were individually tested in the reaction media containing 120 mg·L(-1), considering the following parameters: temperature (20-45°C), H(2)O(2) concentration (0-4.44 mmol·L(-1)), and reaction time (5 minutes, 1 and 24 h). The following conditions: 35°C, 0.55 mmol·L(-1), and 1h, provided the best set of results of color removal for DMBLR (99%), DMBBLN (77%), DMR (94%), and RBBR (97%). It should be mentioned that only 5 minutes of reaction was enough to obtain 96% of decolorization for DMBLR and RBBR. After the decolorization reactions of DMBLR, DMR, and RBBR, it was possible to observe the reduction of Artemia salina mortality and the no significant increase in toxicity for the products generated from DMBBLN.

Dibenzothiophene oxidation by horseradish peroxidase in organic media: effect of the DBT:H2O2 molar ratio and H2O2 addition mode.

Its is well known that in the biodesulfurization (BDS) process the low water solubility of sulfur compounds hinders its transference from the oil phase to the cells being the rate-limiting step in the metabolism of dibenzothiophenes (DBT). Thus sulfur compounds derivatives with high water solubility could be more easily transported increasing the BDS efficiency. The present work performed a stepwise evaluation of the enzymatic oxidation of DBT by horseradish peroxidase (HRP). Reactions were carried out in monophasic organic media containing 25% (v/v) acetonitrile. The following parameters were evaluated: DBT:H2O2 molar ratio (1:1-1:20); H2O2 addition mode (single or stepwise); pH (6.0-8.0) and temperature (37-50 degrees C). Best results were observed in a reaction medium at pH 8.0 presenting HRP 0.06IUml(-1), DBT 0.267mM, DBT:H2O2 molar ratio of 1:20 (stepwise hydrogen peroxide addition) and incubated at 45 degrees C for 60min. Under these conditions 60% of DBT was converted into dibenzothiophene sulfoxide (12%) and dibenzothiophene sulfone (46%). The DBT oxidation rate observed in this work, of 5mmolmin(-1)g(-1) of HRP, was 250-fold higher than the BDS rate, 20mumolmin(-1)g(-1) of catalyst. As such a combined enzyme-microbial desulfurization process could be envisaged. Products were determined by HPLC RP C-18.