Exploitation of Aureobasidium pullulans NRRL Y-2311-1 xylanase in mulberry and rice flours-based gluten-free cookie formulation: Effects on dough properties and cookie characteristics.

Xylanases are mainly utilized in bakery industry for the hydrolysis of dietary fiber-based fractions. Their applications in gluten-free products have not been considered before. In the present study, the xylanase produced by Aureobasidium pullulans NRRL Y-2311-1 was utilized in a mulberry and rice flours-based gluten-free cookie formulation for the first time. Effects of various xylanase concentrations on gluten-free dough rheology and cookie characteristics were elucidated. Only rice flour-based cookie and only wheat flour-based cookie formulations were also prepared as comparison. Incorporation of xylanase into all cookie recipes resulted in softer cookie doughs with lower absolute stickiness. The hardness and absolute stickiness of the cookie doughs prepared by the mixture of mulberry and rice flours decreased by the addition of the enzyme into the formulation in a concentration-dependent manner. Enzyme concentrations above 100 U/100 g flour did not provide statistically significant further changes on gluten-free cookie doughs. Incorporation of xylanase into the cookie recipes resulted in increased baking loss and spread ratio in an enzyme concentration-dependent manner for all cookie types. Hardness values of both types of gluten-free cookies decreased by xylanase incorporation. Different effects on fracturability were observed depending on the cookie type and enzyme concentration. Enzyme concentration of 100 U/100 g flour provided mulberry and rice flours-based cookies with a more flexible and softer structure. No significant effects on color parameters of cookies were observed by xylanase incorporation.

Application of oligosaccharides in meat processing and preservation.

In recent decades, consumer preference and attention to foodstuff presented as healthy and with desirable nutritional information, has increased significantly. In this field, the meat industry has a challenging task since meat and meat products have been related to various chronic diseases. Functional ingredients have emerged in response to the increasing demand for healthier and more nutritious foods. On this matter, oligosaccharides such as fructooligosaccharides (FOS), xylooligosaccharides (XOS), galactooligosaccharides (GOS), and chitooligosaccharides (COS) have been presented as suitable ingredients for the meat industry with the aim of obtaining healthier meat derivatives (e.g. with low fat or sugar content, reduced amount of additives, and desirable functional properties, etc.). However, studies considering application of such oligomers in the meat sector are scarce. In addition, a large number of issues remain to be solved related both to obtaining and characterizing the oligosaccharides available in the industry and to the effect that these ingredients have on the features of meat products (mainly physicochemical and sensory). The study of new oligosaccharides, the methodologies for obtaining them, and their application to new meat products should be promoted, as well as improving knowledge about their effects on the properties of functional meat foods.

Valorization of egg shell as a detoxifying and buffering agent for efficient polymalic acid production by Aureobasidium pullulans NRRL Y-2311-1 from barley straw hydrolysate.

Stepwise formulation of a versatile and cost-effective medium based on barley straw hydrolysate and egg shell for efficient polymalic acid production by A. pullulans NRRL Y-2311-1 was carried out for the first time. The strain did not grow and produce polymalic acid when dilute acid pretreated barley straw hydrolysate (total fermentable sugars: 94.60 g/L; furfural: 1.01 g/L; hydroxymethylfurfural: 0.55 g/L; acetic acid: 5.06 g/L) was directly used in medium formulation without detoxification (e.g. charcoal pretreatment). When CaCO3 in the medium formulation was substituted with egg shell powder, efficient production of polymalic acid was achieved without a detoxification step. Utilization of 40 g/L of egg shell powder led to 43.54 g polymalic acid production per L with the productivity of 0.30 g/L/h and yield of 0.48 g/g. The bioprocess strategy used in this study can also be utilized for mass production of several other industrially important microbial organic acids and biomaterials.

Xylanase production by Aureobasidium pullulans on globe artichoke stem: Bioprocess optimization, enzyme characterization and application in saccharification of lignocellulosic biomass.

Statistical optimization of the factors affecting xylanase production by Aureobasidium pullulans NRRL Y-2311-1 on globe artichoke stem was performed for the first time. The optimization strategies used resulted in almost six-fold enhancement of xylanase production (66.48 U/ml). Biochemical and thermal characterization of the crude xylanase preparation was performed to elucidate its feasibility for different industrial applications. The optimum conditions for xylanase activity were pH 4.0 and 30-50°C. The enzyme was very stable over a wide pH range of 3.0-8.0. The thermal stability studies revealed an inactivation energy of 183 kJ/mol. Thermodynamic parameters (enthalpy, entropy, and Gibbs free energy) for thermal inactivation were also determined. Primary application of the crude xylanase preparation in saccharification of corn cob subjected to different pretreatment techniques has been evaluated. The crude xylanase preparation was very promising for saccharification of corn cob pretreated with aqueous ammonia. The maximum yield of reducing sugar was 357 mg/g dry substrate, which revealed that the crude xylanase from A. pullulans could be a very good alternative in saccharification of lignocellulosic biomass for biological fuel generation. This study also provides a basis for further exploitation of globe artichoke by-products in microbial production of several other industrially significant metabolites.

Single-step purification and characterization of an extreme halophilic, ethanol tolerant and acidophilic xylanase from Aureobasidium pullulans NRRL Y-2311-1 with application potential in the food industry.

An extracellular xylanase from Aureobasidium pullulans NRRL Y-2311-1 produced on wheat bran was purified by a single-step chromatographic procedure. The enzyme had a molecular weight of 21.6kDa. The optimum pH and temperature for xylanase activity were 4.0 and 30-50°C, respectively. The enzyme was stable in the pH range of 3.0-8.0. The inactivation energy of the enzyme was calculated as 218kJmol-1. The xylanase was ethanol tolerant and kept complete activity in the presence of 10% ethanol. Likewise, it retained almost complete activity at a concentration range of 0-20% NaCl. In general, the enzyme was resistant to several metal ions and reagents. Mg2+, Zn2+, Cu2+, K1+, EDTA and ß-mercaptoethanol resulted in enhanced xylanase activity. The Km and Vmax values on beechwood xylan were determined to be 19.43mgml-1 and 848.4Uml-1, respectively. The enzyme exhibits excellent characteristics and could, therefore, be a promising candidate for application in food and bio-industries.

Levan production by Zymomonas mobilis in batch and continuous fermentation systems.

Levan production in batch and continuous fermentation systems by Zymomonas mobilis B-14023 was investigated. The culture medium used in both of the fermentation systems contained sucrose and various organic nitrogen sources. Maximum concentration of levan was produced with yeast extract among the nitrogen sources tested. Response surface methodology was used to investigate the effects of three factors on the concentration of levan in batch cultures of Z. mobilis. Maximum levan concentration was 40.2 g/L and this concentration was reached at the optimum levels of process variables, which were 299.1 g/L initial substrate concentration, 42.3 h incubation time, and initial pH 6.0. Continuous fermentation experiments were done in packed bed bioreactor using Ca-alginate immobilized Z. mobilis cells. The highest levan concentration (31.8 ± 0.21 g/L) was obtained at a dilution rate of 0.14 h(-1) while maximum volumetric productivity (6.556 g/(Lh)) was obtained at a dilution rate of 0.22 h(-1). Increasing the dilution rate resulted in decreased levan and increased residual sugar concentrations.

A thermolabile aspartic proteinase from Mucor mucedo DSM 809: gene identification, cloning, and functional expression in Pichia pastoris.

In this study, the cDNA encoding the aspartic proteinase of Mucor mucedo DSM 809 has been identified by RNA ligased-mediated and oligo-capping rapid amplification of cDNA ends (RACE) technique. The gene contained an open reading frame of 1,200 bp and encoded for a signal peptide of 21 amino acid residues. Two N-glycosylation sites were observed within the identified sequence. The proteinase gene was cloned into the vector pGAPZαA and expressed in Pichia pastoris X-33 for the first time. The protein has been secreted in functionally active form into the culture medium. The expression system does not require any acid activation process. The factors affecting the expression level were optimized in shaking flask cultures. Maximum enzyme production was observed with an initial medium pH of 3.5 at 20 °C and 220 rpm shaking speed utilizing 4 % glucose as a carbon and energy source. The enzyme was purified with cation exchange chromatography and further studies revealed that the enzyme was secreted in glycosylated form. The purified enzyme exhibited remarkable sensitivity to thermal treatment and became completely inactivated after incubation at 55 °C for 10 min. These results indicated that the recombinant proteinase could be considered as a potential rennet candidate for the cheese-making industry.

Aspartic proteinases from Mucor spp. in cheese manufacturing.

Filamentous fungi belonging to the order of Mucorales are well known as producers of aspartic proteinases depicting milk-clotting activity. The biosynthesis level, the biochemical characteristics, and the technological properties of the resulting proteinases are affected by the producer strain and the mode of cultivation. While the milk-clotting enzymes produced by the Rhizomucor spp. have been extensively studied in the past, much less is known on the properties and potential applications of the aspartic proteinases obtained for Mucor spp. Indeed, several Mucor spp. strains have been reported as a potential source of milk-clotting enzymes having unique technological properties. Both submerged fermentation and solid substrate cultivation are proven alternatives for the production of Mucor spp. aspartic proteinases. This review provides an overview on the bioprocessing routes to obtain large amounts of these enzymes, on their structural characteristics as related to their functional properties, and on their industrial applications with focus on cheese manufacturing.