Purification of xylanases from Aureobasidium pullulans CCT 1261 and its application in the production of xylooligosaccharides.

Xylanolytic enzymes are involved in xylan hydrolysis, the main ones being endo-ß-1,4-xylanases (xylanases). This can be applied in the bioconversion of lignocellulosic materials into value-added products such as xylooligosaccharides (XOS). This study aimed to establish a protocol for the purification of xylanases, as well as to characterize and apply the purified enzyme extract in the production of XOS. The enzyme purification techniques studied were ammonium sulfate ((NH4)2SO4) and ethanol precipitation. Purification of xylanase by fractional precipitation (20-60%) with (NH4)2SO4 was more efficient than with ethanol because the salt precipitation reached a purification factor of 10.27-fold and an enzymatic recovery of 48.6% The purified xylanase exhibited optimum temperature and pH of 50 °C and 4.5, respectively. The Michaelis-Menten constant using beechwood xylan for the enzyme was 74.9 mg/mL. The addition of salts such as CaCl2, ZnCl2, and FeCl3 in the reaction medium increased the xylanase activity. Xylanase showed greater thermal stability (half-life = 169 h) at 45 °C and pH 4.5. Under these conditions and in the presence of Ca2+ (10 mmol/L) the enzyme was even more stable (half-life = 231 h). Total XOS contents (6.7 mg/mL) and the conversion of xylan to XOS (22.3%) between 2 and 24 h were statistically equal. The hydrolysates showed the majority composition of xylobiose, xylotriose, and xylose. The addition of Ca2+ ions did not contribute to an increase in the total XOS content or to a greater conversion of xylan into XOS, but the hydrolysate was richer in xylobiose and had a lower xylose content.

Agroindustrial byproduct-based media in the production of microbial oil rich in oleic acid and carotenoids.

This study focuses on the potential of Rhodotorula mucilaginosa CCT 7688 in simultaneous production of lipids and carotenoids in agroindustrial byproduct-based media and specially aims at establishing a process condition that guarantees high concentrations of both bioproducts, i.e., a carotenoid-rich microbial oil with potential economic value and health benefits attributed to carotenoids and fatty acids. Four different combinations of cultivation modes (batch and fed-batch) and alternative substrates (crude glycerol, sugarcane molasses and corn steep liquor) were tested. The M2-B assay, which comprises the use of an agroindustrial byproduct-based medium without any supplementation (70 g L-1 sugarcane molasses and 3.4 g L-1 corn steep liquor) and batch mode, was selected as the most promising one to produce both compounds. Total carotenoid production and total lipid content were 1794.2 µg L-1 and 43.2% (w/w), respectively, after 144 h of cultivation. The fatty acid profile showed predominance of oleic acid (69.9%) and palmitic acid (23.2%). Thus, R. mucilaginosa CCT 7688 may be used in simultaneous production of lipids and carotenoids successfully; its fatty acid profile is similar to that found in olive oil. Both compounds are economically interesting and have great possibility of future commercial applications.

Carotenoid Production by Rhodotorula mucilaginosa in Batch and Fed-Batch Fermentation Using Agroindustrial Byproducts.

Carotenoids are natural pigments that can be produced through biotechnological processes. However, the costs are relatively high and can be minimized by using lower-cost substrates as alternative nutrient sources. The fed-batch fermentation is one of the techniques used to obtain a high biomass concentration and/or maximum production. Thus, the aim of this work is to produce carotenoids in batch and fed-batch fermentation with the yeast Rhodotorula mucilaginosa CCT 7688 using agroindustrial byproducts in the culture medium. Carotenoid production was increased using experimental designs, which modified the concentration of the agroindustrial medium. In batch production the highest concentrations of total carotenoids (1248.5 µg/L) and biomass (7.9 g/L) were obtained in the medium containing 70 g/L sugar cane molasses and 3.4 g/L corn steep liquor at 25 °C and 180 rpm in 168 h, demonstrating an increase of 17% when compared to the standard yeast malt medium (1200 µg/L). In the fed-batch production, different feeding strategies were tested with 30 g/L sugar cane molasses and 6.5 g/L corn steep liquor, reaching a total carotenoid production of 3726 µg/L and biomass concentration of 16 g/L. Therefore, the strategy of the fed-batch process resulted in an increase in the carotenoid production of approx. 400% compared to that in the batch process (740.3 µg/L). Thus, the R. mucilaginosa strain has the potential to produce carotenoids in agroindustrial medium.

Different cell disruption methods for obtaining carotenoids by Sporodiobolus pararoseus and Rhodothorula mucilaginosa.

Since carotenoids are synthesized inside the cell, it is desirable to find an efficient method to extract carotegenic pigments. This study aimed at comparing the effectiveness of different chemical and mechanical techniques to disrupt the cell wall of Sporidiobolus pararoseus and Rhodotorula mucilaginosa yeasts isolated from environmental samples. Among the techniques under study, the ultrasonic bath and the abrasion with glass beads yielded the most promising results for S. pararoseus (84.8 ± 2.3 and 76.9 ± 2.1 µg/g, respectively). The ultrasonic bath yielded the highest specific concentration of carotenoids for R. mucilaginosa (193.5 ± 25.8 µg/g), while the biomass freezing process improved neither the extractability nor the specific concentration of carotenoids. Lyophilization increased the specific concentrations of carotenoids from S. pararoseus and R. mucilaginosa by 20 and 13.7%, respectively, while the freezing process did not significantly affect (p > 0.05) the recovery of carotenoids from both yeasts; thus, it may be eliminated from the process.

Maximization of beta-galactosidase production: a simultaneous investigation of agitation and aeration effects.

In this work, the agitation and aeration effects in the maximization of the beta-galactosidase production from Kluyveromyces marxianus CCT 7082 were investigated simultaneously, in relation to the volumetric enzyme activity and the productivity, as well as the analysis of the lactose consumption and production of glucose, and galactose of this process. Agitation and aeration effects were studied in a 2 L batch stirred reactor. A central composite design (2(2) trials plus three central points) was carried out. Agitation speed varied from 200 to 500 rpm and aeration rate from 0.5 to 1.5 vvm. It has been shown in this study that the volumetric enzyme production was strongly influenced by mixing conditions, while aeration was shown to be less significant. Linear models for activity and productivity due to agitation and aeration were obtained. The favorable condition was 500 rpm and 1.5 vvm, which lead to the best production of 17 U mL(-1) for enzymatic activity, 1.2 U mL(-1) h(-1) for productivity in 14 h of process, a cellular concentration of 11 mg mL(-1), and a 167.2 h(-1) volumetric oxygen transfer coefficient.