Destarched Wheat Bran Research Articles

Extraction of arabinoxylans from wheat bran using hydrothermal processes assisted by heterogeneous catalysts

The extraction/hydrolysis process of arabinoxylans from destarched wheat bran was studied in this work using different mesoporous silica supports and the corresponding RuCl3-based catalysts. The effects of temperature, time, catalyst supports and ruthenium catalysts were investigated and discussed in terms of the arabinoxylans extraction yield and their polymerization degree. Relatively high temperatures (180°C), short extraction times (10min) and RuCl3 supported on Al-MCM-48 led to a high amount of extracted arabinoxylans (78%) with a low molecular weight (9kDa). Finally, a relation between the operating conditions, the arabinoxylans extraction yield and the molecular weight was stablished based on the obtained results.

Replacement of carbohydrate binding modules improves acetyl xylan esterase activity and its synergistic hydrolysis of different substrates with xylanase.

BackgroundAcetylation of the xylan backbone was a major obstacle to enzymatic decomposition. Removal of acetyl groups by acetyl xylan esterases (AXEs) is essential for completely enzymatic hydrolysis of xylan. Appended carbohydrate binding modules (CBMs) can promote the enzymatic deconstruction of plant cell walls by targeting and proximity effects. Fungal acetyl xylan esterases are strictly appended to cellulose-specific CBM1. It is still unclear whether xylan-specific CBMs have a greater advantage than CBM1 in potentiating the activity of fungal deacetylating enzymes and its synergistic hydrolysis of different substrates with xylanase.ResultsThree recombinant AXE1s fused with different xylan-specific CBMs, together with wild-type AXE1 with CBM1 and CBM1-deleted mutant AXE1dC, were constructed in this study. The optimal temperature and pH of recombinant AXE1s was 50 °C and 8.0 (except AXE1dC-CBM6), respectively. Cellulose-specific CBM1 in AXE1 obviously contributed to its catalytic action against substrates compared with AXE1dC. However, replacement of CBM1 with xylan-specific CBM4-2 significantly enhanced AXE1 thermostability and catalytic activity against soluble substrate 4-methylumbelliferyl acetate. Whereas replacements with xylan-specific CBM6 and CBM22-2 were more effective in enzymatic release of acetic acid from destarched wheat bran, NaClO2-treated wheat straw, and water-insoluble wheat arabinoxylan compared to AXE1. Moreover, replacement with CBM6 and CBM22-2 also resulted in higher degree releases of reducing sugar and acetic acid from different substrates when simultaneous hydrolysis with xylanase. A good linear relationship exists between the acetic acid and reducing sugar release.ConclusionsOur findings suggested that the replacement with CBM6 and CBM22-2 not only significantly improved the catalysis efficiency of AXE1, but also increased its synergistic hydrolysis of different substrates with xylanase, indicating the significance of targeting effect in AXE1 catalysis mediated by xylan-specific CBMs.Electronic supplementary materialThe online version of this article (doi:10.1186/s12896-016-0305-6) contains supplementary material, which is available to authorized users.

Feruloyl esterases from Schizophyllum commune to treat food industry side-streams

Agro-industrial side-streams are abundant and renewable resources of hydroxycinnamic acids with potential applications as antioxidants and preservatives in the food, health, cosmetic, and pharmaceutical industries. Feruloyl esterases (FAEs) from Schizophyllum commune were functionally expressed in Pichia pastoris with extracellular activities of 6000UL−1. The recombinant enzymes, ScFaeD1 and ScFaeD2, released ferulic acid from destarched wheat bran and sugar beet pectin. Overnight incubation of coffee pulp released caffeic (>60%), ferulic (>80%) and p-coumaric acid (100%) indicating applicability for the valorization of food processing wastes and enhanced biomass degradation. Based on substrate specificity profiling and the release of diferulates from destarched wheat bran, the recombinant FAEs were characterized as type D FAEs. ScFaeD1 and ScFaeD2 preferably hydrolyzed feruloylated saccharides with ferulic acid esterified to the O-5 position of arabinose residues and showed an unprecedented ability to hydrolyze benzoic acid esters.

An acidic feruloyl esterase from the mushroom Lactarius hatsudake: A potential animal feed supplement

An acidic feruloyl esterase (lhFAE) from Lactarius hatsudake was purified to homogeneity, resulting in a high specific activity of 10.3U/mg toward methyl ferulate as substrate, which was 33.2-fold higher than that of the crude enzyme. lhFAE was a monomeric protein with a molecular mass of 55kDa. All three internal amino acid sequences exhibited 100% identity with the Laccaria bicolor carbohydrate esterase family 9 protein. The enzyme demonstrated high activity toward methyl ferulate (MFA) at an optimum pH of 4.0 and an optimum temperature at 30°C, and under these conditions, the Km and Vmax were 0.54mM and 13.84Umg−1, respectively. The purified FAE exhibited preference for methyl caffeate over methyl ferulate, and was least active on methyl p-coumarate. Interestingly, above 80% of the enzyme activity was preserved in the presence of metal ions at 5.0mM concentration, which indicated that lhFAE was highly metal-tolerant. lhFAE was able to release ferulic acid from de-starched wheat bran (DSWB) both in the absence and presence of xylanase, and 70.2% (of the total amount of) ferulic acid (FA) was released by the synergistic action of xylanase. The result showed that lhFAE has good acid and metal ion stability, and is suitable to be used as an animal feed supplement.

Thermotolerant hemicellulolytic and cellulolytic enzymes from Eupenicillium parvum 4-14 display high efficiency upon release of ferulic acid from wheat bran.

To characterize the hemicellulolytic and cellulolytic enzymes from novel fungi, and evaluate the potential of novel enzyme system in releasing ferulic acid (FA) from biomass resource. A hemicellulolytic and cellulolytic enzyme-producing fungus 4-14 was isolated from soil by Congo red staining method, and identified as Eupenicillium parvum based on the morphologic and molecular phylogenetic analysis. The optimum temperature of fungal growth was 37°C. Hemicellulolytic and cellulolytic enzymes were produced by this fungus in solid-state fermentation (SSF), and their maximum activities were 554, 385, 218, 2·62 and 5·25Ug(-1) for CMCase, xylanase, β-glucosidase, FPase and FAE respectively. These enzymes displayed the best catalytic ability at low pH values (pH 4·5-5·0). The optimum temperatures were 70°C, 70°C, 75°C and 55°C for CMCase, β-glucosidase, xylanase and FAE respectively. CMCase, xylanase and FAE were stable at different pHs or high temperature (60°C). Enzymatic hydrolysis experiment indicated that the maximum (76·8±4)% of total alkali-extractable FA was released from de-starched wheat bran by the fungal enzyme system. High activities of thermotolerant CMCase, β-glucosidase, xylanase and FAE were produced by the newly isolated fungus E.parvum 4-14 in SSF. The fungal enzyme system displayed high efficiency at releasing FA from wheat bran. This study provides a new fungal strain for researches of novel hemicellulolytic and cellulolytic enzymes and will improve the bioconversion and utilization of agricultural by-products.

Solid-state fermentation for the production of biomass valorizing feruloyl esterase

Abstract Aspergillus niger ATCC 13497 has been used to produce feruloyl esterase type A enzyme. The process parameters were optimized to attain a maximum of 11984.1 U/gds of enzyme activity within a period of 72 h using destarched wheat bran as a substrate. The purified enzyme studied in the present work displays a pH optimum of 5, and a temperature optimum of 37 °C. The enzyme has a size of 35 kDa and exhibited moderate thermostability, with 41% of activity retained at 90 °C. Ni +2 and Co +2 ions were found to have stimulatory activity on the enzyme with 66% and 78% increase in activity respectively. Cu+2 inhibited the activity by 88%. Statistical optimization enhanced the enzyme activity by 2.47 fold.

Heterologous production and characterization of a chlorogenic acid esterase from Ustilago maydis with a potential use in baking

Ustilago maydis, an edible mushroom growing on maize (Zea mays), is consumed as the food delicacy huitlacoche in Mexico. A chlorogenic acid esterase from this basidiomycete was expressed in good yields cultivating the heterologous host Pichia pastoris on the 5L bioreactor scale (reUmChlE; 45.9UL−1). In contrast to previously described chlorogenic acid esterases, the reUmChlE was also active towards feruloylated saccharides. The enzyme preferred substrates with the ferulic acid esterified to the O-5 position of arabinose residues, typical of graminaceous monocots, over the O-2 position of arabinose or the O-6 position of galactose residues. Determination of kcat/Km showed that the reUmChlE hydrolyzed chlorogenic acid 18-fold more efficiently than methyl ferulate, p-coumarate or caffeate. Phenolic acids were released by reUmChlE from natural substrates, such as destarched wheat bran, sugar beet pectin and coffee pulp. Treatment of wheat dough using reUmChlE resulted in a noticeable softening indicating a potential application of the enzyme in bakery and confectionery.

The saccharification of destarched wheat bran with microwave-assisted acid treatment

ABSTRACTDestarched wheat bran was hydrolyzed for saccharification using a microwave-assisted acid treatment. The result of the wheat bran with treatment obtained the maximum reducing sugar content of 13.2 g/L with the glucose content of 1.5 g/L, xylose content of 4.8 g/L, and arabinose content of 3.2 g/L under the optimized conditions with microwave power of 400 w, 30 min, and 2% concentration of H2SO4. The scanning electron microscope and infrared spectroscopy showed that the wheat bran could be greatly degraded using the treatment of microwave-assisted acid.

Biochemical characterization of a halotolerant feruloyl esterase from Actinomyces spp.: refolding and activity following thermal deactivation

Ferulic acid esterases (FAE, EC. 3.1.1.73) hydrolyse the linkage between hemicellulose and lignin and thus have potential for use in mild enzymatic pretreatment of biomass as an alternative to thermochemical approaches. Here, we report the characterization of a novel FAE (ActOFaeI) obtained from the bacterium, Actinomyces sp. oral which was recombinantly expressed in Escherichia coli BL21 in two forms: with and without its putative signal peptide. The truncated form was found to have <10 % relative activity compared to the full length and was more prone to aggregation after purification. The enzyme with retained peptide demonstrated 2 to 4-fold higher activity against methyl caffeate and methyl p-coumarate, with specific activities of 477.6 and 174.4 U mg(-1) respectively, than the equivalent activities of the benchmark FAE from Aspergillus niger A and B. ActOFaeI retained activity over a broad pH range with a maximum at 9 but >90 % relative activity at pH 6.5 and an optimum reaction temperature of 30 °C. ActOFaeI increased activity by 15% in high salt conditions (1000 mMNaCl) and its thermal unfolding temperature improved from 41.5 °C in standard buffer to 74 °C in the presence of 2500 mM sodium malonate. ActOFaeI also released ferulic acid from destarched wheat bran when combined with a xylanase preparation. After treatment above the thermal denaturation temperature followed by cooling to room temperature, ActOFaeI demonstrated spontaneous refolding into an active state. ActOFaeI displays many useful characteristics for enzymatic pretreatment of lignocellulose and contributes to our understanding of this important family.

Heterologous production of a feruloyl esterase from Pleurotus sapidus synthesizing feruloyl-saccharide esters.

The feruloyl esterase (FAE) gene EST1 from the basidiomycete Pleurotus sapidus was heterologously expressed in Escherichia coli and Pichia pastoris. Catalytically active recombinant Est1 was secreted using P. pastoris as a host. For expression in P. pastoris, the expression vector pPIC9K was applied. The EST1 gene was cloned with an N-terminal α-mating factor pre-pro sequence and expressed under the control of a methanol inducible alcohol oxidase 1 promotor. Est1 was purified to homogeneity using ion exchange and hydrophobic interaction chromatography. The recombinant Est1 showed optima at pH 5.0 and 50°C, and released ferulic acid from saccharide esters and from the natural substrate destarched wheat bran. Substrate specificity profile and descriptor-based analysis demonstrated unique properties, showing that Est1 did not fit into the current FAE classification model. Transferuloylation synthesis of feruloyl-saccharide esters was proven for mono- and disaccharides.

Enhancing the Thermostability of Feruloyl Esterase EstF27 by Directed Evolution and the Underlying Structural Basis.

To improve the thermostability of EstF27, two rounds of random mutagenesis were performed. A thermostable mutant, M6, with six amino acid substitutions was obtained. The half-life of M6 at 55 °C is 1680 h, while that of EstF27 is 0.5 h. The Kcat/Km value of M6 is 1.9-fold higher than that of EstF27. The concentrations of ferulic acid released from destarched wheat bran by EstF27 and M6 at their respective optimal temperatures were 223.2 ± 6.8 and 464.8 ± 11.9 μM, respectively. To further understand the structural basis of the enhanced thermostability, the crystal structure of M6 is determined at 2.0 Å. Structural analysis shows that a new disulfide bond and hydrophobic interactions formed by the mutations may play an important role in stabilizing the protein. This study not only provides us with a robust catalyst, but also enriches our knowledge about the structure-function relationship of feruloyl esterase.

Non-alkaline solubilization of arabinoxylans from destarched wheat bran using hydrothermal microwave processing and comparison with the hydrolysis by an endoxylanase

Destarched wheat bran (DWB) contained 47.7% hemicellulose, with an arabinose/xylose ratio (A/X) equal to 0.54. When hydrolyzed by an endoxylanase, around 25% of the DWB was solubilized within 4h mostly as monomer-free arabinoxylans (AX) and predominantly arabinoxylo-oligosaccharides (AXOS), with A/X of 0.22. After 20h, the solid residue still contained more than 35% non-cellulosic saccharides. Then, microwave heating in pressure vessels was used to extract the AX from DWB under acid to neutral pH according to a Box–Behnken experimental design. The effects of temperature, time and pH were assessed on: (1) matter solubilization, (2) its content in total and free monosaccharides, (3) HMF and 2-furaldehyde, (4) A/X of the released polymers, (5) AX remaining in the residues. Those responses were also plotted against the combined severity factors (CSF). Such processes led to fast and efficient solubilizations of the hemicellulose into AX/AXOS/monosaccharides with variable yields and proportions. Three independent runs were conducted with the parameters optimizing the total saccharides of the solubilized matter in order to verify the model. Both processes can lead to tailored AX/AXOS mixtures with various A/X, according to the intended application. The microwave-screened parameters may be upscaled in a hydrothermal reactor in a multi-product biorefinery perspective.

Immobilization of feruloyl esterases on magnetic nanoparticles and its potential in production of ferulic acid

Feruloyl esterase plays an indispensable role in hydrolyzing plant cell walls, and ferulic acid will be released as by-product, which has potential applications in food and medicine industry. This study presents immobilization of partially purified feruloyl esterases from the fermentation liquor of recombinant Pichia on magnetic Fe3O4 nanoparticles. Furthermore, the optimal conditions for the immobilization and some characteristics of immobilized enzyme were studied. The optimal immobilization conditions observed were enzyme 0.2mg (1mg/mL, 0.2mL), magnetic Fe3O4 nanoparticles 4mg, pH 6.0, immobilization time 3h. The results showed that the optimal reaction temperature of immobilized enzyme was increased from 45°C to 55°C. Thermal stability of the immobilized enzyme also had an improvement, the residual activity retained 80% and 69% after 120h at 40°C and 50°C, respectively, while free enzymes only showed 45% and 40% remnant activity at the same condition. The immobilized enzyme also exhibited good operational stability and 52.4% of its initial activity was observed during the fifth cycle. In terms of the thermal and operational stability, the immobilized enzyme could be better used in many more applications than the free enzymes. At last the destarched wheat bran was taken as substrate for immobilized and free feruloyl esterases to produce ferulic acid, the maximum ferulic acid yield was 11.2% and 12.3%, respectively, indicating a great potential in industrial application.

Characterization of cell wall components of wheat bran following hydrothermal pretreatment and fractionation

BackgroundPretreatments are a prerequisite for enzymatic hydrolysis of biomass and production of ethanol. They are considered to open up the plant cell wall structure by altering, moving or solubilizing lignin and hydrolyzing a proportion of hemicellulosic moieties. However, there is little information concerning pretreatment-induced changes on wheat bran cell wall polymers and indeed on changes in cell wall phenolic esters in bran or other lignocellulosic biomass. Here, we evaluate polymeric changes (chemical and physical) as a result of selected hydrothermal pretreatment conditions on destarched wheat bran using controlled polymer extraction methods. Quantification of cell wall components together with soluble oligosaccharides, the insoluble residues and ease of extractability and fractionation of biomass residues were conducted.ResultsPretreatment solubilized selected arabinoxylans and associated cross-linking ferulic and diferulic acids with a concomitant increase in lignin and cellulosic glucose. The remaining insoluble arabinoxylans were more readily extractable in alkali and showed considerable depolymerization. The degree of arabinose substitution was less in xylans released by higher concentrations of alkali. The recalcitrant biomass which remained after pretreatment and alkali extraction contained mostly cellulosic glucose and Klason lignin. Pretreatment generated small but insignificant amounts of yeast-inhibiting compounds such as furfural and hydroxymethyl furfural.As such, simultaneous saccharification and fermentation of the hydrothermally pretreated bran resulted in increased ethanol yields compared to that of the control (97.5% compared to 63% theoretical).ConclusionHydrothermal pretreatment of destarched wheat bran resulted in degradation and depolymerization of the hemicellulosic arabinoxylans together with some breakdown of cellulosic glucose. This was accompanied by a significant reduction in the cross-linking phenolic acids such as ferulic and diferulic acids. The results suggest that hydrothermal pretreatment enhances enzymatic digestibility of the cellulose not only by depolymerization and solubilization of the hemicelluloses but by breakdown of interpolymeric phenolic cross-links between the remaining insoluble polymers. This allows easier access of hydrolytic enzymes by opening or loosening of the cell wall thus resulting in enhanced saccharification of cellulose and subsequent fermentation to ethanol. The reduction in cinnamic acids by selected breeding or biotechnological approaches could provide a useful basis for improved saccharification and fractionation of wheat bran polysaccharides.

A chlorogenic acid esterase with a unique substrate specificity from Ustilago maydis.

An extracellular chlorogenic acid esterase from Ustilago maydis (UmChlE) was purified to homogeneity by using three separation steps, including anion-exchange chromatography on a Q Sepharose FF column, preparative isoelectric focusing (IEF), and, finally, a combination of affinity chromatography and hydrophobic interaction chromatography on polyamide. SDS-PAGE analysis suggested a monomeric protein of ∼71 kDa. The purified enzyme showed maximal activity at pH 7.5 and at 37°C and was active over a wide pH range (3.5 to 9.5). Previously described chlorogenic acid esterases exhibited a comparable affinity for chlorogenic acid, but the enzyme from Ustilago was also active on typical feruloyl esterase substrates. Kinetic constants for chlorogenic acid, methyl p-coumarate, methyl caffeate, and methyl ferulate were as follows: Km values of 19.6 μM, 64.1 μM, 72.5 μM, and 101.8 μM, respectively, and kcat/Km values of 25.83 mM(-1) s(-1), 7.63 mM(-1) s(-1), 3.83 mM(-1) s(-1) and 3.75 mM(-1) s(-1), respectively. UmChlE released ferulic, p-coumaric, and caffeic acids from natural substrates such as destarched wheat bran (DSWB) and coffee pulp (CP), confirming activity on complex plant biomass. The full-length gene encoding UmChlE consisted of 1,758 bp, corresponding to a protein of 585 amino acids, and was functionally produced in Pichia pastoris GS115. Sequence alignments with annotated chlorogenic acid and feruloyl esterases underlined the uniqueness of this enzyme.

Effect of polygalacturonase and feruloyl esterase from Aspergillus tubingensis on demucilage and quality of coffee beans

To explore the potential for the application of Aspergillus tubingensis enzyme extract in coffee processing, the effects of crude enzymes on coffee beans were studied. Yields of polygalacturonase and feruloyl esterases obtained from solid-state fermentation using A. tubingensis, with pectin and de-starched wheat bran as carbon sources, were 15.9U/mL and 2.44U/mL, respectively. Crude enzyme extracts removed the mucilage of coffee cherries within 3h, which is substantially more efficient than traditional fermentation. Additionally, the viscosity of coffee mucilage was reduced to 80% by a 3-h treatment with the crude enzyme extract at 50°C. The titratable acidity and organic acids in coffee beverages were also decreased to half the amounts of those in the traditionally fermented group. The total chlorogenic acid in the green beans decreased to 67.3%; however, a decline of only 14.3% was observed in the traditionally fermented group. On the other hand, chlorogenic acid lactones in the roasted beans were reduced to 63.9%, and a 37.2% decline in the chlorogenic acid content was detected.

Extraction by three processes of arabinoxylans from wheat bran and characterization of the fractions obtained

Arabinoxylans (AXs) were extracted from destarched wheat bran (DWB) according to different processes, with a view to their production at industrial scale. Two fractions (F3a and F3b, respectively, purified on 10kDa and 100kDa ultrafiltration membranes) were obtained with low yields by treating DWB with an endoxylanase and this process left a solid residue exhausted in enzyme-extractable AXs (EDWB). F1 and F2 AX fractions were obtained by treatment with sodium hydroxide of, respectively, DWB and EDWB. The fraction F4 resulted from a hydrothermal treatment of EDWB in a pressure reactor, followed by ethanol precipitation. The different AX fractions were characterized and compared for the composition in monosaccharides, for the contents in fats and in ferulic, phytic and uronic acids, for the molecular mass distribution and the degrees of methylation and acetylation. The alkaline extractions gave one deesterified AX population with molecular mass (MM) higher than 670kDa and arabinose/xylose ratios (Ara/Xyl) around 1. The enzyme and thermal treatments yielded AXs with two main populations in size-exclusion chromatography (the largest one at 5–12.5kDa and a second one at 140–160kDa), having overall Ara/Xyl of, respectively, 0.7 and 0.5 for both processes. These data bring information about the influence of the process on the characteristics of AX fractions obtained from pretreated wheat bran. Here are also reported processes that enabled to recover enzyme-unextractable AXs from DWB, including an original and up-scalable hydrothermal extraction. Phytate contents of isolated AXs are described for the first time.

A halotolerant type A feruloyl esterase from Pleurotus eryngii

An extracellular feruloyl esterase (PeFaeA) from the culture supernatant of Pleurotus eryngii was purified to homogeneity using cation exchange, hydrophobic interaction, and size exclusion chromatography. The length of the complete coding sequence of PeFaeA was determined to 1668 bp corresponding to a protein of 555 amino acids. The catalytic triad of Ser-Glu-His demonstrated the uniqueness of the enzyme compared to previously published FAEs. The purified PeFaeA was a monomer with an estimated molecular mass of 67 kDa. Maximum feruloyl esterase (FAE) activity was observed at pH 5.0 and 50 °C, respectively. Metal ions (5 mM), except Hg2+, had no significant influence on the enzyme activity. Substrate specificity profiling characterized the enzyme as a type A FAE preferring bulky natural substrates, such as feruloylated saccharides, rather than small synthetic ones. Km and kcat of the purified enzyme for methyl ferulate were 0.15 mM and 0.85 s−1. In the presence of 3 M NaCl activity of the enzyme increased by 28 %. PeFaeA alone released only little ferulic acid from destarched wheat bran (DSWB), whereas after addition of Trichoderma viride xylanase the concentration increased more than 20 fold.

Comprehensive Study of Valuable Lipophilic Phytochemicals in Wheat Bran

Wheat bran, the major side-stream generated in the milling of wheat grains in the production of white flour, contains significant quantities of carbohydrate and proteins. While not interfering with flour utilization, the bran could be considered as an important feedstock within a biorefinery concept. Wheat bran also contains some amounts of lipids that can be used as a source of valuable phytochemicals. Gas chromatography and mass spectrometry analysis of the lipid composition of destarched wheat bran demonstrated that the predominant lipids found in wheat bran were free fatty acids (ca. 40% of total lipids), followed by acylglycerols (40%). Additionally, important amounts of alkylresorcinols (13% of total lipids) and steroid compounds (hydrocarbons, ketones, free sterols, sterol glycosides, sterol esters, and sterol ferulates) (7% of total lipids) were also present among the lipids of wheat bran. The use of wheat bran as a valuable source of phytochemicals of interest in the context of a wheat bran biorefinery is discussed.

Cloning, expression of a feruloyl esterase from Aspergillus usamii E001 and its applicability in generating ferulic acid from wheat bran

A cDNA gene (AufaeA), which encodes a mature polypeptide of the type-A feruloyl esterase from Aspergillus usamii E001 (abbreviated to AuFaeA), was cloned and heterologously expressed in Pichia pastoris GS115. One transformant, labeled as P. pastoris GSFaeA4-8, expressing the highest recombinant AuFaeA (reAuFaeA) activity of 10.76U/ml was selected by the flask expression test. The expressed reAuFaeA was purified to homogeneity with an apparent molecular weight of 36.0kDa by SDS-PAGE analysis, and characterized using the model substrate of methyl ferulate (MFA). The purified reAuFaeA was optimally active at pH 5.0 and 45°C, and highly stable at pH 4.0-6.5 and 45°C or below. Its activity was not significantly affected by metal ions tested and EDTA. The K m and V max of reAuFaeA towards MFA were 4.64mM and 115.5U/mg, respectively. High-performance liquid chromatography analysis showed that only 9.7% of total alkali-extractable ferulic acid (FA) was released from destarched wheat bran by reAuFaeA alone. The released FA increased to 36.5% when reAuFaeA was used together with a recombinant Aspergillus usamii GH family 11 xylanase A, indicating a synergistic interaction between them.