Comparison of releasing bound phenolic acids from wheat bran by fermentation of three Aspergillus species

Wheat bran was fermented at 28 °C for 7 days under 70% humidity by Aspergillus niger, Aspergillus oryzae and Aspergillus awamori. Total phenolic content (TPC) of the unfermented sample was 1531.5 μg g^?1 wheat bran. After the fermentation of Aspergillus awamori, Aspergillus oryzae and Aspergillus niger, TPC reached 5362.1, 7462.6 and 10 707.5 μg g^?1, respectively. The antioxidant activity in the extractions of fermented wheat bran also increased significantly compared with the unfermented sample (P < 0.05). Aspergillus niger showed the greatest capacity to release bound ferulic acid (416.6 μg g^?1). Aspergillus oryzae and Aspergillus awamori had the advantages of releasing more chlorogenic acid (84.0 μg g^?1) and syringic acid (142.3 μg g^?1), respectively. The destructive effect of Aspergillus niger on wheat bran structure was the strongest, followed by that of Aspergillus oryzae. This effect of Aspergillus niger may be due to its higher cellulase, xylanase, arabinofuranosidase and β‐xylosidase activities. Besides, Aspergillus oryzae possessed higher β‐glucosidase activity, and Aspergillus awamori had higher α‐amylase and feruloyl esterase activities. Aspergillus niger may be the best to release bound phenolic acids in the three Aspergillus species. These will provide the helpful information for understanding mechanism of the fermentation by Aspergillus species releasing bound phenolic in wheat bran.     

Production of α‐amylase by Aspergillus oryzae As 3951 in solid state fermentation using spent brewing grains as substrate

BACKGROUND: The optimisation of nutrient levels for the production of α‐amylase by Aspergillus oryzae As 3951 in solid state fermentation (SSF) with spent brewing grains (SBG), an inexpensive substrate and solid support, was carried out using response surface methodology (RSM) based on Plackett–Burman design (PBD) and Box–Behnken design (BBD). RESULTS: In the first optimisation step a PBD was used to evaluate the influences of related factors. Corn steep liquor, CaCl₂ and MgSO₄ were found to be the most compatible supplements to the substrate of SBG and influenced α‐amylase activity positively. In the second step the concentrations of these three nutrients were optimised using a BBD. The final concentrations (g/g dry substrate basis) in the medium optimised with RSM were 1.8% corn steep liquor, 0.22% CaCl₂ and 0.2% MgSO₄ · 7H₂O using SBG as the solid substrate. The average α‐amylase activity reached 6186 U g^?1 dry substrate under the optimised conditions at 30 °C after 96 h. Under the optimised conditions of SSF an approximately 17.5% increase in enzyme yield was observed. CONCLUSION: SBG was found to be a good substrate for the production of α‐amylase by A. oryzae As 3951 under SSF. Copyright © 2007 Society of Chemical Industry     

Effect of substrate and fermentation conditions on pectinase and cellulase production by <Emphasis Type="Italic">Aspergillus niger</Emphasis> NCIM 548 in submerged (SmF) and solid state fermentation (SSF)

The present study deals with the optimization of substrate and fermentation conditions for the production of both pectinase and cellulase by Aspergillus niger NCIM 548 under same fermentation conditions in submerged fermentation (SmF) and solid state fementation (SSF) using a central composite face centered design of response surface methodology (RSM). As per statistical design, the optimum conditions for maximum production of pectinase (1.64 U/mL in SmF and 179.83 U/g in SSF) and cellulase (0.36 U/mL in SmF and 10.81 U/g in SSF) were, time 126 h, pH 4.6, and carbon source concentration 65 g/L in SmF and were time 156 h, pH 4.80, and moisture content 65% in SSF. The response surface modeling was applied effectively to optimize the production of both pectinase and cellulase by A. niger under same fermentation conditions to make the process cost-effective in both submerged and solid state fermentation using agro industrial wastes as substrate.     

Recovery of residual nutrients from agri-food byproducts using a combination of solid-state fermentation and insect rearing

The present research develops an innovative nutrient recovery strategy for capturing of un-utilised nutrients from agri-food byproducts using a combination of solid state fermentation (SSF) and insect rearing. SSF of borage and flaxseed meals were performed using GRAS organisms to release indigenous nutrients and to produce additional nutrients. Proximate analysis of the SSF-meals showed increases in both lipid and protein contents. Black soldier fly larvae (BSFL) were then grown on these SSF-meals for 12 day. The meals fermented singularly with Lactobacillus plantarum or Aspergillus niger displayed up to 30% enhancement in BSFL biomass, and dual fermentation with L. plantarum and Aspergillus oryzae resulted in an additional 10% enhancement. These examinations showed SFL use over-90% of proteins and lipids in SSF-meals. The results indicate that fermentation of these low-value meals can boost the efficacy of larval growth and the recovery of nutrients from agricultural byproducts as larval biomass.     

High production of Aspergillus niger β‐glucosidase at pilot‐scale and application for hydroxytyrosol release from olive by‐product

This study reports a very simple and fast method to collect extract with high amounts of hydroxytyrosol, biotransforming Olea europaea L. leaf extract and olive mill wastewater by a β‐glycosidase produced locally by Aspergillus niger. These extracts are considered as natural substances from a vegetal source; possessing a higher concentration of simple phenols, characterised by a stronger antioxidant capacity which could be commercially found as dietetic products and food integrators. The proposed process may prove useful for a further application for recycling O. europaea by‐product. In this study, a β‐glucosidase for A. niger was optimised in submerged culture conditions and produced at pilot‐scale. The optimal culture conditions for extracellular β‐glucosidase production by the submerged fermentation of A. niger were investigated using various parameters, including different substrate, nitrogen source, C/N ratio.     

Large-scale production and application of leucine aminopeptidase produced by <Emphasis Type="Italic">Aspergillus oryzae</Emphasis> LL1 for hydrolysis of chicken breast meat

Leucine aminopeptidase (LAP) production by Aspergillus oryzae LL1 (LAP LL1) was scaled up by varying fermenting conditions. The maximal LAP activity for 5, 20, 250, and 2,000 L fermenters were 0.22, 0.16, 0.19, and 0.12 U mL⁻¹, respectively. LAP LL1 production coincides with increasing pH for all scale cases. A pilot plant scale hydrolysis experiment using LAP LL1 was prepared by removing the cells from 1,000 L of A. oryzae LL1 culture and concentrating the supernatant to ∼60 L, with a LAP LL1 activity yield of ∼20% and a specific LAP activity of 0.054 U mg⁻¹, comparable to Flavourzyme. Using a casein substrate, LAP LL1 specific protease activity (50.9 U mg⁻¹) is 43% higher than Flavourzyme (35.6 U mg⁻¹). We believe LAP LL1 has potential use as an industrial enzyme for food processing. Comparing the hydrolysis effects on chopped chicken breast meat (100 g), the degree of hydrolysis (D_H) of LAP LL1 (50.6%) was about twice as effective as Flavourzyme (25.6%). On a pilot-plant scale hydrolysis experiment of chopped chicken breast meat (54 kg), the D_H of LAP LL1 was 45.6%. Adding 30 μmol L⁻¹ zinc increased D_H by 33.3% for LAPs LL1 but zinc did not enhance Flavourzyme hydrolysis. Together, this study provides possibility that LAP LL1 has significant potential for application in the food industry. Shie-Jea Lin and Yi-Hong Chen have contributed equally to this work.     

Comparison of the antioxidant activities of roasted and explosive puffed coffees

We investigated the explosive process effect on antioxidant activities of coffee bean. The total polyphenol contents of powdered extract of explosive puffing coffee bean at 0.75 MPa (PEP 7.5) and powdered extract of explosive puffing coffee bean at 0.9 MPa (PEP 9.0) were at a significantly higher than that of the powdered extract of roasting coffee bean (PER) (P < 0.05). PEP 7.5 showed the highest levels of 3‐CQA (86.23 μg mg^?1), 4‐CQA (43.71 μg mg^?1), and 5‐CQA (31.66 μg mg^?1), and PEP 9.0 had also similar levels of chlorogenic acids, with 3‐CQA (77.99 μg mg^?1), 4‐CQA (43.71 μg mg^?1), and 5‐CQA (30.32 μg mg^?1). PEP 7.5 and PEP 9.0 showed relatively higher antioxidant capacities in DPPH, ABTS, taurine, FRAP, and β‐carotene/linoleic acid assays. PEPs partly recovered the HepG2 cell damage induced by t‐BOOH. These results suggest that puffed coffee has beneficial health effects, and could be used for the development of novel processed coffee products.     

Effects of a mixed koji culture of Aspergillus oryzae HG‐26 and Aspergillus niger HG‐35 on the levels of enzymes,antioxidants and phenolic compounds in soy sauce during the fermentation process

Soy sauce contains a number of bioactive components, which have been shown to possess strong antioxidative properties. The objective of this study was to compare the enzyme activities, antioxidant activity and phenolic compounds content of soy sauces prepared from a mixed koji (SSAON, inoculated with Aspergillus oryzae HG‐26 and Aspergillus niger HG‐35), with those of soy sauces made from a koji culture containing only Aspergillus oryzae HG‐26 koji (SSAO). The total phenolic content (TPC), total flavonoid content (TFC), content of three isoflavone aglycones (daizein, glycitein and genistein) and the antioxidant activity of the SSAON were found to be higher than those of the SSAO during moromi fermentation. In addition, they showed strong positive correlations with the antioxidant potential of the soy sauce. This study has demonstrated the potential of a mixed‐culture koji, based upon A. orzyae and A. niger, for the production of soy sauce with an increased level of bioactive components.     

Aqueous Extraction of Coconut Oil by an Enzyme-Assisted Process

A pre-extraction enzyme treatment of copra meal was incorporated into a rural wet copra oil extraction process to investigate the possibility of improving the traditional process by the enzyme treatment. The enzymes used in the study were combinations of a protease from Aspergillus niger, a cellulase/hemicellulase preparation from Trichoderma reseei, and α-amylase from A oryzae and a pectinase from A niger (all were crude preparations). Finely milled copra meal samples were mixed with water and the enzymes and incubated at 37°C for 6 h or more. After the treatment, the meal was extracted using a water flotation technique. Yield increases of about 50%, relative to experimental controls, were observed, suggesting the possibility of using the enzyme-assisted oil extraction methods to improve upon traditional copra processing. © 1997 SCI.     

Reduction of raffinose oligosaccharides in chickpea (Cicer arietinum) flour by crude extracellular fungal α-galactosidase

The efficiency of crude extracelluar α-galactosidases from Cladosporium cladosporides, Aspergillus oryzae and A niger in reducing the raffinose and stachyose content in chickpea flours was studied and compared with other traditional treatments. The optimum pH for α-galactosidase activity was found to be 4·5 for A oryzae and 5·0 for Cl cladosporides and A niger, while the optimum temperature of enzyme activity was 40°C for Cl cladosporides and 50°C for A oryzae and A niger. The specific activities of α-galactosidase from Cl cladosporides, A oryzae and A niger were 3·35, 3·94 and 5·94 units μg⁻¹ protein, respectively. The enzyme activity was stable between pH 4·0 and 7·0 for A oryzae and A niger and between pH 5·0 and 7·0 for Cl cladosporides. The enzymes were thermostable when incubated at temperature ranges of 40–60°C for Cl cladosporides and 40–50°C for A oryzae and A niger. The optimum conditions for removing the raffinose and stachyose were obtained by incubating chickpea flours with 30 ml of crude fungal α-galactosidase extract (290, 210 and 130 units ml⁻¹ for Cl cladosporides, A oryzae and A niger, respectively) for 3 h at the optimum conditions of each strain. Crude fungal α-galactosidases reduced the raffinose oligosaccharides content in chickpea flours by 100%, while germination reduced the raffinose content by 69% and stachyose content by 75%. Other traditional techniques reduced the raffinose content by 13–49% and stachyose content by 10–32%. © 1998 Society of Chemical Industry     

Characterization of an extracellular β‐d‐fructofuranosidase produced by Aspergillus niveus during solid‐state fermentation (SSF) of cassava husk

β‐d ‐Fructofuranosidases are biotechnologically important enzymes produced by various organisms. Here, Aspergillus niveus produced an extracellular β‐d ‐fructofuranosidase during SSF of cassava husk. This enzyme was purified 8.5‐fold (recovery of 5.2%). A 37‐kDa protein band was observed after 8% SDS‐PAGE. Native molecular mass is 91.2 kDa. Optimal temperature and pH of activity were 55°C and 4.5, respectively. The enzyme was stable at 50°C for 1 hr, and 80% of its activity was retained after 1 hr at pH 8.0. The enzymatic activity was improved by Mn²⁺, was resistant to most solvents, and was inhibited by Triton X‐100 and Tween 20. K_m and V_max with sucrose were 22.98 mM and 120.48 U/mg of protein, respectively. With Mn²⁺, these values were 16.31 mM and 0.30 U/mg of protein. The enzyme did not hydrolyze inulin and for this reason can be considered a true invertase. Thus, A. niveus β‐d ‐fructofuranosidase holds promise for invert sugar production.

Practical applications

β‐d ‐Fructofuranosidase is an enzyme that can be applied to different industrial sectors, especially food and beverage industries. It is responsible for the hydrolysis of sucrose and yields an equimolar mixture of D‐glucose and D‐fructose, named as inverted sugar syrup, with broad applications in the confectionery industry. The Aspergillus niveus enzyme hydrolyzed only sucrose here and can be considered a true invertase, showing its potential for application to invert sugar production. Besides, the use of cassava husk for enzyme production means an interesting utilization route of this agroindustrial residue. Thus, characterization of this enzyme is an important step for identification of its potential for practical applications.     

Optimization of process conditions for the production of a prolylendopeptidase by <Emphasis Type="Italic">Aspergillus niger</Emphasis> ATCC 11414 in solid state fermentation

The effect of 8 factors [(with/without) daily mixing and moisture control, incubation time (t), temperature, ratio between dry substrate mass and bed’s cross section area (MA), inoculum size (spores/g), wheat germ content (WG), initial pH, and moisture content (M)] in the production of a prolyl endopeptidase (PEP) by Aspergillus niger ATCC 11414 in solid state fermentation (SSF) was tested. Contribution of all the factors was significant (p<0.05); main effects were those of MA, t, and M. The 4 interactions that presented high interaction severity indexes involved the WG. Under optimized conditions PEP and protease activity were 9.76±0.06 and 3.6×10⁶±1.5×10⁵ U/kg, respectively. The enzyme was partially purified (ammonium sulfate precipitation, dialysis, DEAE-Sepharose ionexchange); it has a molecular weight of 66 kDa (SDS-PAGE), and maximum activity was exhibited at pH 4 and 50°C. The enzyme is stable in a wide pH range (2.2–10) and at temperatures lower than 70°C.     

Optimisation of culture conditions and development of a novel fed‐batch strategy for high production of β‐galactosidase by Kluyveromyces lactis

The aim of this study was to enhance β‐galactosidase production by Kluyveromyces lactis CICC1773. Firstly, the optimum culture conditions were obtained by response surface methodology, and the maximum β‐galactosidase activity reached 20.6 U mL^?1, about two‐fold increase than that of the initial conditions (initial fermentation medium and conditions). To further improve β‐galactosidase production, a new fed‐batch strategy based on pH feedback control was developed successfully in a 7‐L fermenter, using 400 g L^?1 lactose as feeding medium. As a result, the β‐galactosidase activity and productivity reached up to 111.61 U mL^?1 and 5.31 U/(mL·h), enhanced by 15.3‐fold and 17.6‐fold superior than the results of initial conditions, respectively. To our knowledge, β‐galactosidase activity obtained was the highest value among the results reported by nonrecombinant strains. These results demonstrated that the new fed‐batch strategy based on optimum culture conditions could be automatic control easily and be conductive to further scale up for industrial fermentation.     

Production of feed enzymes from citrus processing waste by solid‐state fermentation with Eupenicillium javanicum

Bingtang sweet orange processing waste was utilised to produce four feed enzymes (Endoglucanase, β‐glucosidase, pectinase and xylanase) by the solid‐state fermentation (SSF) with Eupenicillium javanicum. The factors related with SSF including moisture content, temperature, initial pH, time, carbon source (0.5 g), nitrogen sources (0.05 g), inorganic mineral salts (0.1 g) were investigated separately. The corresponding optimal condition was: moisture content 80% (w/w), temperature 30 °C, natural pH, time 96 h, wheat bran 0.5 g, (NH₄)₂SO₄ 0.05 g or NaNO₃ 0.05 g, CaCl₂ 0.1 g. The L₉(3⁴) orthogonal experiment results showed that the optimal condition for producing above multiple enzymes was: moisture content 80% (w/w), temperature 30 °C, wheat bran 1 g, (NH₄)₂SO₄ 0.05 g, NaNO₃ 0.05 g, CaCl₂ 0.1 g, fermentation time 96 h and natural pH. Under this condition, the average activity of Endoglucanase (CMCase), β‐glucosidase, pectinase and xylanase by E. javanicum could reach 46.80, 49.64, 51.87 and 106.42 U g^?1, respectively, which was significantly higher than those in single factor experiments. Our present results demonstrated that E. javanicum could also be an effective and useful fungus for multienzyme preparation especially for β‐glucosidase and xylanase from citrus processing wastes.     

Multiple forms of β‐galactosidase from mango (Mangifera indica L Alphonso) fruit pulp

Mango (Mangifera indica L cv Alphonso) was found to contain three isoforms (I, II and III) of β‐galactosidase which, upon purification on Sephadex G‐200, had relative abundances of 44, 38 and 18%, respectively. The total specific activity increased from 20 to 727 µmol l^?1 upon purification, representing a ～36‐fold increase with a recovery of 0.28 U U^?1. The optimal pH for activity and stability were in the ranges 3.6–4.3 and 4–6.2, respectively. The optimal temperature for β‐galactosidase activity was between 42 and 47 °C with T_m in the range 45–51 °C. The K_m for pNP‐β‐galactopyranoside was 0.98, 1.11 and 0.95 mM , and V_max was 0.56, 0.53 and 0.35 µmol pNP min^?1, respectively for isoforms I, II and III. Hg²⁺ caused strong inhibition, whereas galacturonic acid, galactose, xylose, fucose and mannose slightly inhibited the activity of β‐galactosidase isoforms. The apparent molecular weights by GPC were 78, 58 and 91 kDa for isoforms I, II and III, respectively. The ability of these isoforms to degrade the endogenous substrate (arabinogalactan) possibly suggests a role in pectin dissolution during tissue softening/fruit ripening. Copyright © 2004 Society of Chemical Industry     

Production and Application of Xylanase from Penicillium sp. Utilizing Coffee By-products

The lignocellulosic coffee by-products such as coffee pulp, coffee cherry husk, silver skin, and spent coffee were evaluated for their efficacy as a sole carbon sources for the production of xylanase in solid-state fermentation using Penicillium sp. CFR 303. Among the residues, coffee cherry husk was observed to produce maximum xylanase activity of 9,475 U/g. The process parameters such as moisture (50%), pH (5.0), temperature (30 °C), particle size (1.5 mm), inoculum size (20%), fermentation time (5 days), carbon source (xylose), and nitrogen source (peptone) were optimized and the enzyme activity was in the range of 19,560–20,388 U/g. The enzyme production was further improved to 23,494 U/g with steam as a pre-treatment. The extracellular xylanase from the fungal source was purified to homogeneity from culture supernatant by ammonium sulfate fractionation, DE32-cellulose with a recovery yield of 25.5%. It appeared as a single band on SDS-PAGE gel with a molecular mass of approximately 27 kDa. It had optimum parameters of 50 °C temperature, pH 5.0, K _m 5.6 mg/mL, and V _max 925 μmol mg⁻¹ min⁻¹ with brichwood xylan as a substrate. The crude enzyme hydrolysed lignocellulosic substrate as well as industrial pulp. Production of xylanase utilizing coffee by-products constitutes a renewable resource and is reported for the first time.     

Optimized Production of Tannase from Cashew Testa using Aspergillus niger MTCC 5898

The production of enzymes by bioprocessing is a good alternative to add value to agro-industry residues. Production cost of enzymes, the major constraint in bioprocessing, may be brought down by multifaceted approaches which include the use of cheap agro-residues for the microbial-based production of the enzyme, and the use of cost-efficient aerobic cell growth strategies and anaerobic-combined strategies like solid state fermentation (SSF). The current study investigated the production of tannase by Aspergillus niger CEPC 11 (MTCC 5898) on an abundantly available waste byproduct cashew testa under solid state A. niger growth. The optimum values of parameters obtained through response surface methodology (RSM) were cashew testa (23%), K₂HPO₄ (3.40 mM), sodium chloride (0.47 mM) and temperature (32–35°C). Optimization of the amount of cashew testa, content of the medium and incubation temperature resulted in a 3.02-fold increase from 97.32–301.70 U/g DS of tannase.     

Reduction efficiency of the neurotoxin β‐ODAP in low‐toxin varieties of Lathyrus sativus seeds by solid state fermentation with Aspergillus oryzae and Rhizopus microsporus var chinensis

Solid state fermentation of several low‐toxin varieties of grass pea (Lathyrus sativus L) seeds with Aspergillus oryzae and Rhizopus microsporus var chinensis removed the neurotoxin β‐ODAP (3‐N‐oxalyl‐L ‐2,3‐diaminopropanoic acid) to a considerable degree from the seed meal. The detoxification efficiency was statistically significant and ranged from 52.4% (p < 0.01) to 82.2% (p < 0.001), which was lower than for a high‐toxin variety processed by the same fermentation procedure (94.8%, p < 0.001). While the content of β‐ODAP decreased, those of other free protein amino acids, especially glutamic acid, histidine, threonine, tyrosine, valine and lysine, increased dramatically in the fermented seeds. Efforts to remove the neurotoxin from Lathyrus sativus either by breeding or by food processing to obtain toxin‐free grass pea seeds have been made worldwide for several decades. The efficiencies of various reported processing methods are summarised and compared. © 2000 Society of Chemical Industry     

Purification and characterisation of a novel α‐L‐rhamnosidase exhibiting transglycosylating activity from Aspergillus oryzae

A novel α‐L‐rhamnosidase was isolated and purified from Aspergillus oryzae NL‐1. The enzyme was purified 13.2‐fold by ultrafiltration, ion exchange and gel filtration chromatography with an overall recovery of 6.4% and specific activity of 224.4 U/mg, and the molecular mass of its subunit was approximately 75 kDa. Its optimal temperature and pH were 65 °C and 4.5, respectively. The enzyme was stable in the pH range 3.5–7.0, and it showed good thermostability at higher temperatures. The K_M, kcat and kcat/K_M values were 5.2 mm , 1624 s^?1 and 312 s^?1 mm ^?1 using pNPR as substrates, respectively. Moreover, the enzyme exhibited transglycosylating activity, which could synthesise rhamnosyl mannitol through the reactions of transglycosylation with inexpensive rhamnose as the glycosyl donor. Our findings indicate that the enzyme has potential value for glycoside synthesis in the food industry.     

Statistical optimization,partial purification,and characterization of coffee pulp β-glucosidase and its application in ethanol production

Extracellular β-glucosidase was produced using coffee pulp as a sole carbon source by Penicillium verrucosum by solid state fermentation and 897.36±59 U/g enzyme activity was obtained. Increase in 2.21-fold of enzyme activity on optimizing the bioprocess parameters by response surface methodology based on central composite rotatable design is illustrated. Maximum production level of 1,991.17 U/g was obtained with optimum values of pH 4.2, moisture 66.8%, and fermentation duration of 56 h. The enzyme was partially purified and the enzyme activity was optimum at 50°C temperature and at pH 6. The metal ions such as Mg²⁺, Zn²⁺, Ca²⁺, K⁺, detergents, and chelator such as EDTA were effective and further increased the β-glucosidase activity. On application of β-glucosidase for simultaneous saccharifiation and fermentation, 3.3% ethanol was obtained. Thus, this study provides insight on exploitation of P. verrucosum for synthesis of of β-glucosidase using coffee pulp which is available abundantly in coffee processing industries.