Effect of fermentation by Bacillus subtilis on antioxidant and cytotoxic activities of black rice bran

Black rice bran was fermented with Bacillus subtilis KU3 isolated from Korean traditional food, Kimchi. Antioxidant and cytotoxic activities of the fermented black rice bran were investigated. Total phenolic and anthocyanin contents decreased from 171.54 mg GAE g^?1 and 2.31 mg g^?1 to 139.13 mg GAE g^?1 and 2.12 mg g^?1, respectively, after fermentation. Antioxidant activities determined by 1,1‐diphenyl‐2‐picrylhydrazyl (DPPH) radical scavenging, β‐carotene bleaching and ferric thiocyanate assay were correlated with total phenolic and anthocyanin contents. Non‐fermented black rice bran extract (NFBE) showed greater antioxidant activities than fermented black rice bran extract (FBE). Cytotoxic activities measured by MTT assay showed that both NFBE and FBE had over 50% activities. The cytotoxic activities of FBE against MCF‐7 and HeLa cells were 71.65% and 68.07%, respectively, at 8.0 mg mL^?1, but those of NFBE were lower than 50%. These results suggested that the cytotoxic activity of black rice bran improved through fermentation, while antioxidant activity reduced.     

Preparation of alcalase hydrolysed rice bran protein concentrate and its inhibitory effect on soybean lipoxygenase activity

Rice bran protein concentrate (RBPC) was prepared from defatted rice bran and hydrolysed by alcalase at different hydrolysis times. As the hydrolysis times increased, the degree of hydrolysis (DHs) increased. RBPC hydrolysate obtained at 50 min (RBPCH‐50) had the highest inhibitory efficiency on soybean lipoxygenase (LOX) activity (66%). The inhibition kinetics of the reaction analysed by Lineweaver–Burk plots indicates that RBPCH‐50 is a competitive inhibitor. RBPCH‐50 inhibited soybean LOX with an IC₅₀ of 11.73 μg μL^?1 RBPCH‐50, and the obtained K_I was 4.59 μg μL^?1 RBPCH‐50. LOX inhibitory activity of RBPCH‐50 was significantly higher than that of 50 and 100 ppm butylated hydroxyanisole (BHA) and 50, 100, and 200 ppm butylated hydroxytoluene (BHT) (P ≤ 0.05); however, LOX inhibitory activity of RBPCH‐50 was similar to that of 200 ppm BHA (P > 0.05). Therefore, RBPCH might potentially be used as a natural LOX inhibitor.     

Mineral composition,antioxidant and cytotoxic biopotentials of wild‐growing Ganoderma species (Serbia): G. lucidum (Curtis) P. Karst vs. G. applanatum (Pers.) Pat.

The aim of this work was to analyse mineral composition and chemical profile of two nonedible fungal species: Ganoderma lucidum and Ganoderma applanatum (Fru?ka Gora, Serbia) vs. their antioxidant (ABTS and A.E.A.C. assay) and cytotoxic biopotentials (MTT assay on MCF‐7). Both species were analysed for their content of macro‐ and microelements by atomic absorption spectrophotometry, while phenolic profile of EtOH and H₂O extracts was examined by LC‐MS/MS technique. Both species mostly contained the following ions: K⁺ > Ca²⁺ > Mg²⁺ > Mn²⁺ > Zn²⁺ > Cu²⁺ > Cr³⁺. Among nine phenolic compounds, the highest content of vanillic acid was detected in G. applanatum extracts while protocatechuic acid in EtOH extract and quinic acid in H₂O extract were mostly contained in G. lucidum. Ganoderma applanatum EtOH extract showed the best antioxidant activities related to its phenolic and flavonoid content. Further, the best cytotoxic effect after 72 h was observed in this extract as well.     

Production of bioactive proteins and peptides from the diatom Nitzschia laevis and comparison of their in vitro antioxidant activities with those from Spirulina platensis and Chlorella vulgaris

This research focuses on green production of bioactive proteins and hydrolysates from Nitzschia. A comparison of antioxidant activities was established between protein extracts and hydrolysates from Nitzschia and two other well‐known microalgae, chlorella and spirulina. Protein hydrolysates from these microalgae were produced using Alcalase^®, Flavourzyme^® and Trypsin. The hydrolysis process enhanced the antioxidant activities in general, especially those obtained using Alcalase^®. Nitzschia showed the highest (P < 0.05) total phenolic content/reducing capacity (2.4 ± 0.02 mg GAE/100 g) after 90 min of hydrolysis with Alcalase^®. The ABTS [2,2′‐Azino‐bis(3‐ethylbenzothiazoline‐6‐sulphonic acid)] radical scavenging activity (66.77 ± 0.00%) was highest (P < 0.05) after 120 min of hydrolysis, but DPPH (2,2‐Diphenyl‐1‐picrylhydrazyl radical) was low (29.59 ± 0.02%). A correlation between ABTS activity and total phenolic contents was the highest (P < 0.05) for protein hydrolysates from all three organisms using Alcalase^®, but superoxide anion radical scavenging activity was intermediate for Nitzschia. Therefore, Nitzschia protein hydrolysates have the potential to be used as antioxidants.     

Effects of overexpression of the alcohol acetyltransferase‐encoding gene ATF1 and disruption of the esterase‐encoding gene IAH1 on the flavour profiles of Chinese yellow rice wine

Alcohol acetyltransferase (AATase), which is mainly encoded by ATF1, is one of the most important enzymes for acetate ester synthesis. On the other hand, isoamyl acetate is degraded into a higher alcohol under the catalysis of IAH1‐encoded esterase. In this study, Chinese Saccharomyces cerevisiae was used as the parent strain to construct an ATF1 overexpression and IAH1 disruption mutant. The results show that after 5 days of pre‐fermentation, the concentrations of ethyl acetate, isoamyl acetate and isobutyl acetate in the yellow rice wines fermented with EY1 (pUC‐PIAK) increased to 468.94 mg L^?1 (which is approximately 22‐fold higher than that of the parent cell RY1), 99.86 and 7.69 mg L^?1 respectively. Meanwhile, isoamyl alcohol production was reduced to 56.37 mg L^?1 (which is approximately 50% of that produced by the parent strain RY1). Therefore, ATF1 overexpression and IAH1 disruption can significantly increase acetate esters contents and reduce isoamyl alcohol content in Chinese yellow rice wine, thereby paving the way for breeding an excellent yeast strain for high‐quality Chinese yellow rice wine production.     

Production of Agaricus brasiliensis mycelium from food industry residues as a source of antioxidants and essential fatty acids

Agaricus brasiliensis is a medicinal mushroom traditionally employed in prevention and treatment of cancer, used as immunostimulant and as a source of antioxidants. We investigated the chemical composition of the mycelium produced by submerged (SF) and solid‐state fermentation (SSF), using residues from food industry as substrates. After fermentation, antiradical activity and levels of antioxidants were enhanced, indicating that the micro‐organism produces these metabolites. For myceliated substrate (mushroom mycelia grown around and into the substrate particles) obtained by SSF, phenolics ranged from 18.57 to 70.46 mg g^?1 and flavonoids from 0.83 to 4.51 mg g^?1. For myceliated substrate obtained by SF, the variation was 27.19 to 66.99 mg g^?1 and 0.75 to 5.34 mg g^?1 for phenolics and flavonoids, respectively. The fatty acid profile determined by FT‐ICR MS and UPLC‐MS showed predominance of palmitic, linoleic, linolenic and oleic acids. Our findings indicated that mycelium has nutraceutical potential and can be incorporated in food supplements.     

The effect of enzymatic hydrolysis on the biological properties of Oenothera biennis,Borago officinalis and Nigella sativa seedcake by‐products from oil pressing

The biological properties of ethanolic (50%, v/v) extracts from Oenothera biennis, Borago officinalis, Nigella sativa seedcake before and after enzymatic hydrolysis by alpha‐amylase (EC 3.2.1.1) from Aspergillus oryzae, beta‐glucosidase (EC 3.2.1.21) and beta‐glucanase (EC 3.2.1.6) from Aspergillus niger combinations in a ratio of 1:1:1 were investigated. Total phenolic, flavonoid and reducing sugar content for O. biennis extract after enzymatic hydrolysis was, respectively, 0.5, 1.5 and 2 times higher in comparison with nonhydrolysed extract. Iron‐chelating and radical‐scavenging activity of O. biennis seedcake extract after hydrolysis (IC₅₀ = 0.076 mg mL^?1 and IC₅₀ = 0.050 mg mL^?1) was at a similar level as that nonhydrolyeed (IC₅₀ = 0.070 mg mL^?1 and IC₅₀ = 0.065 mg mL^?1). The antioxidant activity was two times higher after hydrolysis than before enzymatic hydrolysis of O. biennis seedcake extract. Also strong elastase inhibition activity has been shown to O. biennis seedcake extract before (IC₅₀ = 0.095 mg mL^?1) and after enzymatic hydrolysis (IC₅₀ = 0.07 mg mL^?1), respectively. Oenothera biennis and B. officinalis seedcake extracts before and after hydrolysis have stronger antibacterial activity against Pseudomonas aeruginosa strain in comparison with N. sativa seedcake.     

Raffinose family oligosaccharides in seed of Glycine max cv. Chiang Mai60 and potential source of prebiotic substances

Raffinose family oligosaccharides (RFOs) content in Glycine max seed of cultivar Chiang Mai60, a local soybean of Thailand, was investigated. RFOs and other low molecular weight sugars were extracted by 50% (v/v) ethanol and quantified by high‐performance liquid chromatography (HPLC). The prebiotic property of this extract was subsequently studied by in vitro method. The results showed that the concentrations of raffinose, stachyose and verbascose were 6.74 ± 1.62, 145.32 ± 18.74 and 1.60 ± 0.52 mg g^?1 dry seed, respectively, while glucose and sucrose were detected at 10.73 ± 1.35 and 13.28 ± 2.16 mg g^?1 dry seed, respectively. The growth of four Lactobacilli probiotics were increased significantly in a basal liquid medium supplemented with this ethanolic extract as carbon source compared to glucose supplementation. Subsequently, defined mixed culture was studied and it was found that growth stimulation of total Lactobacilli by extracted sugars resulted in the suppression of Escherichia coli and Salmonella enterica serovar Typhimurium growth. It could be concluded that this cultivar showed the RFOs‐rich content and a potential to be a source of an effective prebiotic substance for food application.     

Optimization of Aspergillus niger Fermentation for the Production of Glucose Oxidase

A number of nutritional factors influencing glucose oxidase (EC 1.1.3.4) production by Aspergillus niger NCIM 545 were studied. The synthesis of glucose oxidase by A. niger was investigated in two steps using submerged fermentation at 30 ± 2 °C and 180 rpm for 96 h. Primarily, nutritional components were selected by one-factor-at-a-time method, and the significance of each component with respect to glucose oxidase production was identified by Plackett–Burman design (seven variables including six nutritional viz. sucrose, sodium nitrate, peptone, calcium carbonate, magnesium sulfate, and potassium dihydrogen phosphate, and one dummy or unassigned variable were studied with eight experiments). In the second step, concentration of most significant factors and their interaction were studied with response surface methodology (central composite design). Each variable in the design was studied at five different levels, with all variables taken at a central coded value of zero. Considerable amount of glucose oxidase was produced from A. niger species with sucrose as the carbon source, sodium nitrate as the inorganic nitrogen source, and peptone as the organic nitrogen source. Glucose oxidase activity increased remarkably by 28.93 fold (from 0.00993 to 0.29 U ml⁻¹) with CaCO₃-supplemented media. The outcome of Plackett–Burman design showed CaCO₃, peptone, and MgSO₄ as significant parameters. Further optimization using a three-factor central composite design with 20 experiments increased yield of glucose oxidase from 0.29 to 2.05 U ml⁻¹ (sevenfold) with a decrease in cultivation time from 96 to 72 h.     

Study on inactivation mechanisms of Listeria grayi affected by pulse magnetic field via morphological structure,Ca2+ transmembrane transport and proteomic analysis

The inactivation mechanism of PMF on Listeria grayi (L. grayi) was investigated through the analysis of biological effects as well as the monitoring of morphology, membrane permeability and intracellular proteins of cells. Under the optimal inactivation conditions of PMF (2.5 T and 25 pulses), the analysis of transmission electron microscopy (TEM) and laser scanning confocal microscopy (LSCM) showed that PMF‐treated cells membranes were damaged, resulting in the increase in intracellular Ca²⁺ fluorescence intensities. A significant (P < 0.01) negative correlation between intracellular Ca²⁺ fluorescence intensities and the amount of colonies was found. Proteomic analysis showed that the mode‐of‐action of cells outer membrane, the stability of intracellular proteins and the metabolism‐related proteins might be affected by PMF. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that PMF treatment could affect the nitrogen, organic and energy metabolisms of L. grayi, inducing the death of cells finally.     

Effect of the main ingredients of Rhizoma gastrodiae on mycelial biomass and exopolysaccharide productions by submerged culture of Grifola frondosa

Various concentrations of the main ingredients of Rhizoma gastrodiae including gastrodin (GA) and p‐hydroxybenzyl alcohol (HA) as well as p‐hydroxylbenzaldehyde (HBA) were separately added into the cultural medium of Grifola frondosa in submerged culture. The results showed that the additive concentration of gastrodin from 0.10 to 0.35 g L^?1, p‐hydroxybenzyl alcohol from 0.05 to 0.30 g L^?1 and p‐hydroxylbenzaldehyde from 0.05 to 0.30 g L^?1 can significantly promote the EPS production by submerged culture of G. frondosa (P < 0.05). Among them, the addition of p‐hydroxylbenzaldehyde at 0.15 g L^?1 exhibited the best results, and the EPS productions reached 2.2511 ± 0.0378 g L^?1, with an increase of 45.03% compared with the control (without additives). Furthermore, all of the additives had an effective stimulatory effect on mycelial biomass. In addition, there is an important and novel breakthrough that p‐hydroxylbenzaldehyde was more effective than R. gastrodiae extract on increasing the EPS productions.     

Anti‐inflammatory properties of high pressure‐assisted extracts of Grifola frondosa in lipopolysaccharide‐activated RAW 264.7 macrophages

The aim of this study was to determine the in vitro anti‐inflammatory properties of the shake extract (SE) and the high pressure‐assisted extract (PE) of the mycelia of Grifola frondosa in a lipopolysaccharide‐stimulated RAW 264.7 macrophage model. The content of total polysaccharides and β‐glucans of PE at 600 MPa (PE‐600) was 41.2 and 6.2 mg g^?1 dry weight, respectively, which were significantly higher than SE extracts. The results showed that treatment with 500 μg mL^?1 of PE by 600 MPa (PE‐600) did not reduce RAW 264.7 cell viability but did significantly inhibit the production of LPS‐induced NO, PGE₂ and intracellular ROS. The PE‐600 inhibited the activation of NF‐kB and then reduced the production of LPS‐induced TNF‐α, IL‐6 and IL‐1β in a dose‐dependent manner. Thus, the PE could be used as an alternative extraction method for improving the extraction efficacy of G. frondosa and serve as an alternative source of anti‐inflammatory agents.     

Milk‐clotting enzymes produced by Aspergillus flavo furcatis strains on Amazonic fruit waste

Six strains of Aspergillus flavo furcatis were screened to investigate milk‐clotting enzyme production by fermentation in natural liquid medium. The growth media comprised extracts of cupuaçu exocarp+rice bran [10% or 20% (v/v) CE+RB] and açai waste+rice bran [10% or 20% (v/v) AW+RB] with or without supplementation of 0.1% (w/v) yeast extract and 0.5% (w/v) gelatin. Significant values of milk‐clotting activity were determined by A. flavo furcatis DPUA 1461 and DPUA 1608, in the standard and natural media, respectively. According to criteria of clot and whey formations, 8.3% of the samples tested were classified as strong coagulation, 41.70% showed weak coagulation and in 50% was not observed milk coagulation. The enzyme optimal action of DPUA 1608, the selected strain, was at 40 °C and pH 7.0. Milk‐clotting proteases were inhibited by pepstatin (94.72%) and moderately inhibited by the others metal ions tested.     

Mutation of Acetobacter pasteurianus by UV irradiation under acidic stress for high‐acidity vinegar fermentation

To obtain a mutant with higher vinegar production, a wild‐type industrial strain Acetobacter pasteurianus CICIM B7003 was pretreated in 50 g L^?1 acetic acid for 1 h and cultured for 12 h without adding any acetic acid; then, the enriched cells were mutated by UV under acidic stress (60 g L^?1 acetic acid). Using this method, A. pasteurianus CICIM B7003‐02, a mutant exhibiting best performance, was obtained. Notably, after repeated experiments, it was confirmed that B7003‐02 accumulated 103.81 ± 1.17 g L^?1 acetic acid within 160‐h batch cultivation in Erlenmeyer flasks, which was 49.2% higher than that of the wild type. Repeated batch fermentations with A. pasteurianus B7003‐02 were carried out for several runs in a Frings 8‐L acetator, and high‐acidity vinegars (90 ± 0.39 g L^?1) were produced. This work reveals the potential value for improvement in industrial vinegar production.     

Shelf life and quality of probiotic yogurt produced with Lactobacillus acidophilus and Gobdin

This study evaluated the effect of dry white mulberry and walnut paste (Gobdin, a traditional Turkish food) in probiotic yogurt on the survival of Lactobacillus acidophilus and yogurt properties. Six different yogurts were produced with 0%, 5% and 10% Gobdin using Lactobacillus bulgaricus + Streptococcus thermophilus and with 0%, 5% and 10% Gobdin using L. bulgaricus + S. thermophilus + L. acidophilus. The physical, chemical, microbiological and sensorial properties of the yogurts were evaluated based on storage at 4 ± 1 °C. Probiotic shelf life and the most suitable combinations were determined. The highest L. acidophilus count (8.65 log cfu g^?1) was found in the 5% Gobdin‐supplemented yogurt on the 7th day of storage, while the lowest count (8.11 log cfu g^?1) was found in the probiotic control yogurt on the 21st day. Although the L. acidophilus counts in the probiotic yogurts declined during storage, all values found throughout the 21‐day storage period were >8 log cfu g^?1. This is above the level necessary to provide the desired therapeutic effect in probiotic products (10⁶–10⁷ cfu g^?1). The highest overall acceptability score was obtained on the first day from the yogurt with 5% Gobdin. However, all yogurt samples had general acceptability scores between 7 and 8 points from a 9‐point maximum. Thus, this study determined that a new functional yogurt can be produced using L. acidophilus with 5% Gobdin.     

Characterization of Physical Properties of Flour and Starch Obtained from Gamma‐Irradiated White Rice

Physical and structural characteristics of rice flour and starch obtained from gamma‐irradiated white rice were determined. Pasting viscosities of the rice flour and starch, analyzed by using a Rapid Visco Analyser, decreased continuously with the increase in irradiation dosage. Differential scanning calorimetry showed that gelatinization onset, peak and conclusion temperatures of rice flour and starch changed slightly but the enthalpy change decreased significantly with increase of irradiation dosage. All irradiated starch displayed an A‐type X‐ray diffraction pattern like the native starch. Gel permeation chromatography showed that the blue value ratio of the first peak (amylopectin) to the second one (amylose) decreased with the increase of the irradiation dosage. The weight‐average molecular weight (M_w) and gyration radius (R_z) of amylopectin analyzed by using HPSEC‐MALLS‐RI (high‐performance size‐exclusion chromatography equipped with multiangle laser‐light scattering and refractive index detector) decreased gradually from 1.48×10⁹ (M_w) and 384.1 nm (R_z) of native rice starch to 2.36×10⁸ (M_w) and 236.8 nm of 9 kGy‐irradiated starch. The branch chain‐length distribution of amylopectins determined by HPAEC‐ENZ‐PAD (high‐performance anion‐exchange chromatography with amyloglucosidase post‐column on‐line reactor and pulsed amperometric detector) showed that gamma irradiation had no significant effect on the amylopectin branch chains with 13≤DP≤24 and 37≤DP, but produced more branch chains with 6≤DP≤12 when the irradiation dosage was less than 9 kGy. It might be deduced that gamma irradiation caused the breakage of the amylopectin chains at the amorphous regions, but had little effects on the crystalline regions of starch granules, especially at low dosage irradiation.     

Production of active lipase by Rhizopus oryzae from sugarcane bagasse: solid state fermentation in a tray bioreactor

This study deals with production of lipase in solid state fermentation by Rhizopus oryzae from sugarcane bagasse. A tray bioreactor was designed for the extracellular enzyme production. Daily, lipase production was evaluated at several incubation temperatures. Furthermore, the influence of temperature and humidity of the cabinet, depth of solid bed, particle size, initial moisture content and supplementary substrate (olive oil) as carbon source was investigated. The obtained results showed that bioreactor temperature of 45 °C, humidity of 80%, solid bed depth of 0.5 cm, particle size in the range of 0.335–1 mm, substrate initial moisture content of 80% for the top tray and 70% for the middle tray and supplementary substrate of 8% (v/w) olive oil led to maximum lipase production. Under optimum fermentation conditions after 72‐h incubation, maximum lipase activities for the top, middle and bottom trays were 215.16, 199.36 and 52.64 U gds^?1, respectively.     

Isolation,characterisation and sulphation of soluble polysaccharides isolated from Cucurbita maxima

Using hot water extraction, a large number of polysaccharides were obtained from Cucurbita maxima. A DEAE‐Sepharose CL‐6B chromatography column was used to isolate the major polysaccharides from C. maxima. Two fractions were obtained (LP2‐1 and LP2‐2). LP2‐1 and LP2‐2 consisted of neutral polysaccharides (MW: 1.02 × 10⁴ and 4.32 × 10⁸ g mol^?1, respectively) comprised mainly of galactose units. Analyses by FT‐IR spectrometry, partial acid hydrolysis, periodate oxidation, Smith degradation and GC‐MS indicated that LP2‐1 consisted of 85.3% (1→4) glycosidic linkages and 1.7% (1→3) or (1→6) glycosidic linkages. The LP2‐1 backbone consisted of (1→4)‐linked galactose units, which occasionally branched at O6 or O3. The branches were composed of (1→4)‐linked galactose and terminated with galactose (13%). Two sulphated derivatives (SLP2‐1 and SLP2‐2) with variable degrees of sulphation (DS) were obtained by the sulphur trioxide–pyridine method, without degradation of the polysaccharide. DS of PL2‐1 and PL2‐2 was 0.19 and 0.20, respectively.     

Comparative production of xylanase and the liberation of xylooligosaccharides from lignocellulosic biomass by Aspergillus brasiliensis BLf1 and recombinant Aspergillus nidulans XynC A773

We investigated the simultaneous production of xylanase and the liberation of xylooligosaccharides in rice husk solid‐state cultivations of Aspergillus brasiliensis BLf1 and by the recombinant Aspergillus nidulans XynC A773 strain. The bioprocess was optimised by experimental fractional design and response surface analysis. Results show that both fungi strains produced xylanases and their activities were dependent on the addition of basal medium, moisture content and the interactions between particle size and inoculum size, producing maximum xylanase activities of 230.7 U g^?1 substrate for A. brasiliensis, and 187.9 U g^?1 substrate for A. nidulans XynC. Xylooligosaccharides were liberated in the same cultures, with concentrations up to 17.6 mg g^?1 and 23.9 mg g^?1 of substrate for A. brasiliensis and A. nidulans XynC, respectively, both strains presenting similar profiles, with xylose residues varying from X3 to X6. These results suggest the possibility of lowering production costs of enzymes for food applications and oligosaccharides.     

Monitoring of the bacterial communities of bamboo shoots (Dendrocalamus latiflorus) during pickling process

The diversity and dynamics of the dominant bacterial communities arising during the pickling process of bamboo shoots (Dendrocalamus latiflorus) were investigated by nested polymerase chain reaction denaturing gradient gel electrophoresis combined with quantitative real‐time polymerase chain reaction. Results showed only several kinds of halophilic bacteria during early sampling time (0?3 days). After pickling for 7 days, Lactococcus lactis significantly increased (P < 0.05, quantities were 6.39 × 10⁵ copies μL^?1) and became the first dominant bacterium. After pickling for 14 days, Weissella sp. bands appeared and quickly became dominant on the 21st day (4.07 × 10⁶ copies μL^?1). As maturation progressed, Lc. lactis decreased in intensity whereas Weissella sp. increased in intensity. Finally, the quantities of Weissella sp. (2.50 × 10⁷ copies μL^?1) became higher than those of Lc. lactis (1.43 × 10⁷ copies μL^?1) after pickling for 56 days. The pickling process was initiated by Lc. lactis, followed by Lc. lactis and Weissella sp., and was finally succeeded by Weissella sp.