Optimization Of Fermentation Conditions Research Articles

Harnessing Oxidative Stress to Obtain Natural Riboflavin Secreting Lactic Acid Bacteria for Use in Biofortification.

Lactococcus lactis suffers from oxidative stress and riboflavin starvation at elevated temperatures due to dissolved oxygen, which can be relieved partially by exogenously supplied riboflavin. Here we explore whether this phenomenon can be harnessed to obtain riboflavin overproducing mutants. Using a riboflavin auxotrophic L. lactis strain as a riboflavin biosensor, we screened L. lactis cultures that had been exposed to temperature induced oxidative stress for up to one year. Riboflavin secreting mutants could readily be identified, some of which had arisen after just two weeks of exposure to oxidative stress. Whole genome sequencing revealed mutations in the riboswitch, which regulate riboflavin biosynthesis. Riboflavin secretion conferred a significant increase in tolerance to oxidative stress and enabled growth at high temperatures in the presence of dissolved oxygen. It was subsequently demonstrated that vigorous aeration at high temperature (37 °C) could prompt rapid emergence of riboflavin secreting mutants. The protective effect provided by riboflavin against oxidative stress may explain the natural occurrence of lactic acid bacteria (LAB) secreting riboflavin. By optimizing fermentation conditions and eliminating lactate formation, we achieved 64 mg/L riboflavin, the highest reported titer so far for LAB, which indicates great potential for use as a riboflavin fortification agent in food.

Sequential fermentation of Ginkgo biloba seeds by Bacillus subtilis natto and Lactobacillus plantarum enhanced nutrition, flavor and lipid-lowering activity.

Ginkgo biloba seeds (GBS) are rich in flavonoids, proteins and reducing sugar, and have been consumed as food and medicinal nuts for thousands of years. However, the presence of ginkgotoxins and their poor palatability limit people's consumption of them. This study used solid-state fermentation with Bacillus subtilis natto and Lactobacillus plantarum to enhance the safety and benefits of GBS. Optimized fermentation conditions increased the content of beneficial components like total flavonoids, soluble protein and reducing sugar while eliminating unpleasant odors (isoamyl aldehyde and hexanal) and reducing the toxin 4'-O-methylpyridoxine by 91.17%. Fermentation of GBS powder can significantly enhance its anti-inflammatory and antioxidant activities in vitro (P < 0.001). Furthermore, it exhibits a dose-dependent effect within a certain concentration range. Mixed fermentation (FBnLp) was evaluated for its effects on obesity and metabolic syndrome in mice fed a high-fat diet. FBnLp significantly reduced body and liver weight gain, prevented dyslipidemia and decreased inflammatory and oxidative stress compared to unfermented GBS. Histological analysis showed that FBnLp improved liver health by reducing fat accumulation and preventing non-alcoholic fatty liver disease. Meanwhile, it was found that feeding FBnLp increased the expression of CPT-1α, which regulates energy expenditure and fat breakdown, and downregulated the expression of SREBP-1c, FAS and ACC, which regulate fat synthesis. This research provides new insights and technological support for the application and development of FBnLp as a functional product, addressing key issues in its use and industry growth. © 2024 Society of Chemical Industry.

Machine learning-enhanced modeling and characterization for optimizing dietary Fiber production from Highland barley bran.

This study investigated the modification of highland barley bran through co-fermentation of Lactobacillus bulgaricus and Kluyveromyces marxianus, and developed a dynamic prediction model for DF content under these co-fermentation conditions using machine learning algorithms. The results showed that the XGBoost algorithm could predict changes in the DF component content (R2 = 0.9553(SDF/IDF), RMSE = 0.0464.) and identify optimal fermentation conditions. Under the optimal conditions, both strains exhibited synergistic effects, where the lactic acid produced by Lactobacillus bulgaricus and β-glucosidase produced by Kluyveromyces marxianus might facilitate IDF decomposition and conversion, resulting in a maximum SDF/IDF ratio of 0.6911. This led to a 27.65 % reduction in IDF content and a 19.11 % increase in SDF content. Moreover, the physicochemical and functional properties of DF were enhanced after co-fermentation. The structure of DF became more loose and porous, its thermal stability improved, and its water-holding, oil-holding, and swelling capacities increased by 53.54 %, 16.11 %, and 44.96 %, respectively, compared with the unfermented counterpart; In terms of adsorption characteristics, its glucose, cholesterol and nitrite adsorption capacities were also significantly improved. According to in vitro gastrointestinal simulated digestion, digestion would have a great impact on the fermented DF, which showed good antioxidant properties during the intestinal digestion stage.

The Bioavailability of Solid-State Fermented Feather Meal Using a Novel Feather-Degrading Bacterium Bacillus velezensis PN1 in Broilers

In this study, an effective feather-degrading bacterium was isolated and the solid-state fermentation condition for feather degradation was optimized. The resulting fermented feather meal (FFM) was evaluated for its bioavailability in broilers. Four Bacillus strains were examined for feather degradation rates, with Bacillus velezensis PN1 exhibiting the highest rate (83.24%, p &lt; 0.05). A 3 × 3 × 3 factorial design was used to test substrate moisture content (45%, 55%, 65%), temperature (27 °C, 37 °C, 47 °C), and incubation time (24, 48, 72 h) for optimized fermentation conditions. In vitro pepsin digestibility (IVPD) revealed a significant interaction between temperature and time (p &lt; 0.05), and the optimal performance was achieved at 37 °C for 72 h, followed by 37 °C for 48 h. Considering production time and cost, FFM2 (produced with 65% moisture at 37 °C for 48 h) was further compared with FFM1 (produced under the same conditions but at a lower temperature of 27 °C), and commercial hydrolyzed feather meal (HFM). IVPD did not differ significantly between FFM1, FFM2, and HFM, as they all showed significantly higher digestibility compared to raw feathers (RFs). FFM1, as well as FFM2, had significantly higher histidine and lysine concentrations than RF (p &lt; 0.05), while FFM2 had a significantly lower cysteine content (p &lt; 0.05). Based on fermentation conditions and amino acid composition, FFM2 was considered more suitable for large-scale production and was used in a broiler growth trial. The inclusion of 5% FFM2 in the broiler diet did not significantly affect body weight at 35 days compared to the 5% HFM group (p &gt; 0.05), although both groups showed significantly lower weight gain than the 5% fish meal (FM) group (p &lt; 0.05). The feed conversion ratio and performance efficiency factor did not differ significantly between the FFM2, HFM, and FM groups (p &gt; 0.05). In groups fed diets without supplemental crystalline amino acids, growth performance did not significantly differ between the HFM and FFM2 groups (p &gt; 0.05), although both performed significantly worse than groups with amino acid supplementation. In conclusion, FFM produced by B. velezensis PN1 through solid-state fermentation enhances feather bioavailability in poultry and can completely replace HFM when included at 5% in broiler diets, offering a potential sustainable alternative for poultry nutrition on a commercial scale.

Investigating the effect of fermentation byLactobacillus reuteri on biochemical properties of Ceylon Cinnamon

Abstract Introduction Fermentation is an important biotechnological process that improves the biochemical properties of natural raw materials by transforming high molecular compounds into more beneficial low molecular compounds through microbial biotransformation. Ceylon cinnamon (Cinnamomum zeylanicum Blume), globally recognized as 'true cinnamon', contains high molecular weight, complex compounds that are responsible for its renowned antioxidant, antibacterial, and therapeutic properties, making it a vital component in medicinal applications1. Aims The present study aimed to investigate the effect of fermentation by Lactobacillus reuteri on selected phytochemical contents, antioxidant, and antibacterial properties of bark extracts of Ceylon cinnamon (Cinnamomum zeylanicum Blume) and to determine the optimum conditions for fermentation of Cinnamon bark extract by L. reuteri. Method Authenticated Ceylon cinnamon bark (CCB) was extracted into water using the decoction method. The water extract was fermented by L. reuteri under controlled conditions (pH 6, incubation at 35 ºC) and optimized using two parameters; the inoculum density (1.5×108 cfu/ml and 7.5×108 cfu/ml) and incubation time (2 and 4 days). After fermentation, total polyphenolic content (TPC), total flavonoid content (TFC), DPPH free radical scavenging activity and antibacterial activity of non-fermented extracts (NFE) and fermented extracts (FE) of CCB were evaluated. Antibacterial activity was determined using the agar well diffusion method against Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. Ethical approval was not required since no human or animal study was done during the study. Results The extract fermented with an inoculum density of 7.5×108 cfu/ml and incubated for 4 days showed the highest TFC (19.3 ± 0.3 mg RE/g) and antioxidant activity (IC50: 97.2 ± 0.8 µg/ml) in comparison to that of NFEs (TFC:13.3 ± 0.3 mg RE/g and DPPH activity: IC50: 233.7 ± 12.8 µg/ml). Moreover, the fermented samples showed a reduction in TPC, indicating possible biotransformation of compounds. and no significant change in the antibacterial activity in comparison to the non-fermented samples. Discussion and Conclusion This study was mainly focused on investigating the effect of fermentation of Ceylon cinnamon bark aqueous extract by Lactobacillus reuteri. The increased antioxidant activity and total flavonoid content are notable and could be attributed to chemical transformations of some complex flavanols including catechin gallate and epicatechin gallate present in aqueous cinnamon to catechin and epicatechin monomers by the action of enzymes secreted by L. reuteri during the fermentation process2. Hence, this study suggests that fermentation using L. reuteri has a considerable positive impact on certain biochemical properties of cinnamon water extract. Cinnamon bark is a complex matrix containing various phytochemicals that could interact with bacterial strains in ways that are not fully understood. These interactions could lead to unexpected effects or limitations during fermentation.

Yeast Synthesis and Herbicidal Activity Evaluation of Aspterric Acid.

Aspterric acid (AA) is a novel natural product herbicide that targets dihydroxyacid dehydratase in plants. In this study, we introduced two distinct AA biosynthesis-related gene clusters into Saccharomyces cerevisiae and screened the core biosynthetic enzyme, sesquiterpene cyclase, from various fungi. The combination of sesquiterpene cyclase from Aspergillus taichungensis IBT 19404 and two cytochrome P450s from Penicillium brasilianum resulted in the optimal AA synthesis efficiency in yeast, with the highest titer of 33.21 mg/L achieved by optimizing fermentation conditions in shake flasks. Moreover, the herbicidal effects of AA on weed germination and growth were evaluated. Notably, AA strongly inhibited the germination of Amaranthus tricolor, Portulaca oleracea, Bidens pilosa, Lolium perenne, and Leptochloa chinensis. Furthermore, AA could also inhibit the shoot and root growth of weeds with a superior inhibitory effect on roots relative to shoots. Our work not only provides a sustainable method for the biosynthesis of AA in yeast but also paves the way for the application of AA as a preemergence herbicide.

Investigation of the Fermentation Process of Moringa oleifera Leaves and Its Effects on the Growth Performance, Antioxidant Capacity, and Intestinal Microbiome of Procambarus clarkii

Moringa oleifera is renowned for its high antioxidant activity. However, few studies have been conducted on its effects on aquatic animals. The aim of this experiment was to investigate the optimal fermentation process of M. oleifera leaves and to evaluate the effects of fermented M. oleifera leaves on crayfish (9.11 ± 0.3 g) in terms of growth performance, antioxidant capacity, and gut microbiological parameters. By optimizing the fermenting material/water ratio, fermentation time, temperature, and strain, the optimal fermentation conditions of a 10% water ratio + 48 h + 30 °C + inoculation with 2% B. amyloliquefaciens (107 CFU mL−1) were obtained. These conditions resulted in notable increases in the contents of the total protein, total phenols, flavonoids, and amino acids (p &lt; 0.05) while also leading to a notable decrease in the content of tannins in contrast to those of unfermented M. oleifera leaves (p &lt; 0.05). The fermented M. oleifera (FMO) leaves were incorporated at five concentrations, including 0% (control (CT)), 0.25% (0.25FMO), 0.5% (0.5FMO), 1% (1FMO), and 2% (2FMO). The results showed that the 1FMO group performed better in terms of the final body weight (FBW), weight gain rate (WGR), and specific weight gain rate (SGR) compared with the CT group (p &lt; 0.05). In addition, amylase and lipase activities were significantly higher in the 1FMO and 2FMO groups compared with the other groups (p &lt; 0.05). The fermented M. oleifera leaves significantly increased the catalase (CAT) activity in the crayfish (p &lt; 0.05). The superoxide dismutase (SOD) activity was significantly increased in the 0.25FMO, 1FMO, and 2FMO groups, and the malondialdehyde (MDA) content was significantly decreased while the glutathione peroxidase (GSH-PX) content was significantly increased in the 0.5FMO, 1FMO, and 2FMO groups (p &lt; 0.05). Furthermore, the 1FMO group was observed to significantly increase the abundance of Firmicutes while simultaneously reducing the abundance of Aeromonas (p &lt; 0.05) and adjusting the structure of the intestinal microbiome. In conclusion, this study established the optimal fermentation conditions for M. oleifera and obtained a product with high nutrient and low tannin contents. Furthermore, the incorporation of 1% FMO was demonstrated to facilitate growth, enhance the antioxidant capacity, and optimize the gut microbiology in crayfish.

Effect of Mixed Strains on Microbial Community and Flavor Metabolites in Fermentation Process of Chi-Flavor Baijiu.

The distinct flavor of chi-flavor baijiu (CFB) has garnered significant attention in China. After the optimization of fermentation conditions, Pichia anomala and Lactobacillus plantarum were introduced into the fermentation process to enhance the flavor. Samples inoculated with these mixed strains (SY) exhibited higher levels of alcohol (from 33.04 to 178.55 mg/L) and esters (from 49.51 to 130.20 mg/L) compared to the control group (KB). In SY, P. anomala and L. plantarum were the predominant microorganisms, while Pediococcus and Saccharomyces were more prevalent in KB. Moreover, 68 volatile flavor compounds were detected in SY, as opposed to 64 in KB. Notably, Pichia showed a positive correlation with key flavor compounds. The synergistic fermentation with exogenous strains led to a 52.38% increase in phenethyl alcohol and a 4.91% increase in ethyl lactate. Additionally, the levels of other flavor compounds, like ethyl acetate, γ-nonanolactone, and (E)-2-octenal, also increased. The results demonstrated that the addition of P. anomala and L. plantarum to the fermentation process of CFB significantly increased the contents of flavor compounds. This research reveals valuable insights into flavor enhancement and the microbial community dynamics in CFB production.

Degradation of Anti-Nutritional Factors in Maize Gluten Feed by Fermentation with Bacillus subtilis: A Focused Study on Optimizing Fermentation Conditions

Degradation of Anti-Nutritional Factors in Maize Gluten Feed by Fermentation with Bacillus subtilis: A Focused Study on Optimizing Fermentation Conditions

Preparation of safflower fermentation solution and study on its biological activity.

Safflower, a traditional Chinese medicine, is rich in chemical components including flavonoids, polysaccharides, and alkaloids. It exhibits pharmacological effects such as antioxidant, anti-inflammatory, anti-tumor, and anti-thrombosis properties, making it a valuable resource in the medical field. Furthermore, due to its antioxidant and anti-inflammatory effects, safflower is increasingly being utilized in the cosmetics industry. In this study, yeast was employed to ferment safflower, and the optimal fermentation conditions were established through single-factor experiments and response surface methodology. Subsequently, the antioxidant and anti-inflammatory efficacy of the safflower fermentation solution was assessed using both cellular and zebrafish models. Finally, the safety of the safflower fermentation solution was evaluated through a cosmetic eye irritation test. From a total of 20 yeast strains, YF-5 was identified as the dominant strain for safflower fermentation. By optimizing the fermentation conditions, it was established that the optimal parameters for YF-5 fermentation of safflower are as follows: a fermentation temperature of 36.55°C, a material-to-liquid ratio of 1:20.46, a fructose concentration of 6.20%, a fermentation duration of 72 h, and an inoculum volume of 4%. The biological activities of safflower, including its antioxidant and anti-inflammatory properties, were enhanced through yeast fermentation. In HaCaT cell and zebrafish oxidative damage assays, safflower fermentation solution inhibits the production of malondialdehyde (MDA) and increases superoxide dismutase (SOD) activity as well as total antioxidant capacity (T-AOC). In the RAW264.7 cell inflammatory damage assays, a 20% safflower fermentation solution was found to inhibit the release of TNF-α and NO in the inflammatory model, with inhibition rates of 30.94 and 28.86%, respectively. In the zebrafish inflammatory damage assays, the quantity of fluorescent neutral proteins in the 5% safflower fermentation solution was 0.7 times that observed in the dexamethasone (0.1 mg/mL) positive control group, indicating that its anti-inflammatory activity is comparable to that of dexamethasone (0.1 mg/mL). In the chicken embryo chorionic membrane experiment, it was observed that the safflower fermentation solution did not cause significant damage to the blood vessels of the chorionic allantoic membrane (CAM). This finding demonstrates that the safflower fermentation solution possesses a certain degree of safety. Safflower fermentation solution has antioxidant and anti-inflammatory bioactivities, and it has passed cosmetic safety evaluations. It can be used as a new natural cosmetic ingredient added to cosmetic products.

Optimization of fermentation conditions for physcion production of Aspergillus chevalieri BYST01 by response surface methodology.

This study aimed to optimize the culture conditions of the termite-derived fungus Aspergillus chevalieri BYST01 for the production of physcion, a characteristic component of the traditional herb rhubarb, which has been commercially approved as a botanical fungicide in China. First, potato dextrose broth was screened as the suitable basal medium for further optimization, with an initial yield of 28.0 mg/L. Then, the suitable carbon source, fermentation time, temperature, pH value, and the rotary shaker speed for physcion production were determined using the one-variable-at-a-time method. Based on the results of single factors experiments, the variables with statistically significant effects on physcion production were further confirmed using the Plackett-Burman design (PBD). Among the five variables, temperature, initial pH, and rotary shaker speed were identified as significant factors (P < 0.05) for physcion productivity in the PDB and were further analyzed by response surface methodology (RSM). Finally, we found that the maximum physcion production (82.0 mg/L) was achieved under the following optimized conditions:initial pH 6.6, rotary shaker speed of 177 rpm, temperature of 28 °C, and glucose concentration of 30 g/L in PDB medium after 11 d of fermentation. The yield of physcion under the optimized culture conditions was approximately threefold higher than that obtained using the basal culture medium. Furthermore, the optimum fermentation conditions in the 5-L bioreactor achieved a maximal physcion yield of 85.2 mg/L within 8 d of fermentation. Hence, response surface methodology proved to be a powerful tool for optimizing physcion production by A. chevalieri BYST01. This study may be helpful in promoting the application of physcion produced by A. chevalieri BYST01 to manage plant diseases.

Optimization of Fermentation and Biocontrol Efficacy of Bacillus atrophaeus XHG-1-3m2

Biological control plays an increasingly important role in various aspects of modern agriculture and forestry. Identifying biocontrol strains with commercial potential for effective disease management is currently a focal point in biological control research. In this study, Bacillus atrophaeus XHG-1-3m2, a strain with significant biocontrol potential against Wilsonomyces carpophilus causing shot hole disease in wild apricots, was developed. The study determined the antibacterial activity of the fermentation broth, the optimal fermentation medium composition and conditions, and explored its effectiveness in controlling Wilsonomyces carpophilus. The optimal fermentation medium for strain XHG-1-3m2 comprises 12.5 g/L yeast extract, 12.5 g/L soy peptone, 10.0 g/L sodium chloride, 1 g/L ammonium chloride, 1 g/L potassium dihydrogen phosphate, 1 g/L disodium hydrogen phosphate, and 0.5 g/L magnesium sulfate heptahydrate. With an initial pH of 7.0, a liquid volume of 40%, an inoculum volume of 3%, and shaking incubation at 28 °C for 24 h, the viable cell count reached 14 × 109 CFU/mL. In vitro and in vivo tests on leaves revealed that the fermentation broth and the biocontrol biofertilizer derived from this strain inhibited the leaf lesions caused by Wilsonomyces carpophilus on wild apricots, achieving inhibition rates of 94.62% and 82.46%, respectively.

A Novel Biosynthetic Strategy for Ginsenoside Ro: Construction of a Metabolically Engineered Saccharomyces cerevisiae Strain Using a Newly Identified UGAT Gene from Panax ginseng as the Key Enzyme Gene and Optimization of Fermentation Conditions.

Ginsenoside Ro, as one of the few oleanane-type ginsenosides, is well known for its unique molecular structure and biological activities. Currently, research on the biosynthesis of ginsenoside Ro is still in its early stages. Therefore, the establishment of a new ginsenoside Ro cell factory is of great significance for the in-depth development and utilization of genes related to ginsenoside Ro synthesis, as well as for the exploration of pathways to obtain ginsenoside Ro. In this study, we cloned endogenous constitutive promoters, terminators, and other genetic elements from S. cerevisiae BY4741. These elements were then sequentially assembled with the uridine diphosphate glucuronic acid transferase gene identified in our previously study (PgUGAT252645) and several other reported key enzyme genes, to construct DNA fragments used for integration into the genome of S. cerevisiae BY4741. By sequentially transferring these DNA fragments into chemically competent cells of engineering strains and conducting screening and target product detection, we successfully constructed an engineered S. cerevisiae strain (BY-Ro) for ginsenoside Ro biosynthesis using S. cerevisiae BY4741 as the host cell. Strain BY-Ro produced 253.32 μg/L of ginsenoside Ro under optimal fermentation conditions. According to subsequent measurements and calculations, this equates to 0.033 mg/g DCW, corresponding to approximately 31% of the ginsenoside Ro content found in plant samples. This study not only included a deeper investigation into the function of PgUGAT252645 but also provides a novel engineering platform for ginsenoside Ro biosynthesis.

Innovative Lactic Acid Production Techniques Driving Advances in Silage Fermentation

Lactic acid (LA) plays a crucial role in the silage process, which occurs through LA fermentation. Consequently, there is a strong correlation between lactic acid production and the efficiency of the silage. However, traditional methods face challenges like long fermentation times, low acid production, and unstable quality, limiting agricultural preservation. This paper aims to explore innovations in lactic acid production technologies and show how these technologies have driven the development of silage fermentation for agricultural conservation. First, the important role of LA in agricultural preservation and the limitations of traditional silage techniques are presented. Next, advancements in LA production methods are thoroughly examined, covering the selection of microbial strains and the substitution of fermentation substrates. Following this, new technologies for silage fermentation are explored, drawing from innovations in LA production. These include the selection of LA strains, optimization of fermentation conditions, and improvements in fermentation techniques. These innovations have proven effective in increasing LA production, improving feed quality, extending shelf life, and providing new solutions to enhance agricultural production and sustainability.

Lactic acid bacteria sequential fermentation improves viable counts and quality of fermented apple juice via generating two logarithmic phases

This study investigated the impact of lactic acid bacteria (LAB) sequential fermentation on viable counts and apple juice quality. The optimal fermentation conditions were obtained by a step-by-step optimization process, including pH 4.5, temperature 37 °C, the second inoculation time 16 h, total fermentation time 40 h and fermentation sequence (first 21,805 + 21,828, second 20,241). Under the optimal conditions, sequential fermentation allowed LAB to experience two logarithmic phases, increasing viable counts to 1.38 × 108 CFU/mL, exceeding simultaneous fermentation for 24 h and 40 h by 4.10 × 107 CFU/mL and 5.40 × 107 CFU/mL, respectively. This process enhanced sugar utilization, yielding more lactic acid and polyphenols. Furthermore, sequential fermentation improved DPPH (71.71 %) and ABTS (84.79 %) scavenging rates, and enriched volatile compounds, particularly beta-Damascenone, potentially contributing to floral and richer apple flavor. Sequential fermentation also achieved optimal sensory acceptability. This study proposes a novel strategy for high-density LAB fermentation to produce high-quality apple juice.

Characterization of newly isolated Bacillus cereus D3 Co-producing α-amylase and protease and its application in food raw materials

Bacillus cereus, which effectively secretes various extracellular enzymes, has been widely used in the food, feed, and fermentation industries. One strain co-producing α-amylase and protease was newly isolated from Daqu and identified as B. cereus D3. It was shown that the production of α-amylase and protease co-produced by B. cereus D3 reached 13.89 ± 0.07 and 2468.45 ± 38.96 U/mL through optimization of fermentation conditions (nutrients (g/L): 20 of glucose, 20 of urea, 0.7 of MnCl2; incubation conditions: 24 h of incubation, initial medium pH of 7.0, temperature of 37 °C, 8% inoculum size, and 30% working volume). Moreover, the enzymatic genetic co-expression exhibited a similar pattern to the changes in enzymatic activities (growth-phase-dependent pattern). The two enzymes performed excellent thermal stability and were stable over wide pH ranges of 6.0–11.0 and 2.0–8.0, respectively. Finally, the hydrolysis degrees of raw materials including rice, sorghum, corn, and soybeans by α-amylase from B. cereus D3 reached 66.59 ± 4.21, 57.99 ± 4.39, 57.44 ± 4.67, and 38.01% ± 1.73%, respectively, producing glucose and maltose as the main products. The hydrolysis degrees of proteinous materials including mackerel heads and soybeans by protease from B. cereus D3 were 39.93 ± 1.99 and 30.44% ± 1.39%. SDS-PAGE analysis displayed extensive breakdown of macromolecular proteins, forming peptides of varied sizes. This is a systematic study on the co-production of α-amylase and protease by B. cereus, which is crucial to widen the understanding of this species co-producing multi-enzymes and explore its potential application in the food fermentation industry.

Optimization of fermentation conditions for Bacillus velezensis TCS001 and evaluation of its growth promotion and disease prevention effects on strawberries

Bacillus velezensis TCS001 is a novel biocontrol bacterium with broad-spectrum antifungal activity and plant growth-promoting effects, holding great potential for development in agricultural production. This study optimized the fermentation conditions for B. velezensis TCS001 through single-factor experiments combined with response surface methodology, and developed a formulation for TCS001 suspension concentrate (TCS001-SC). The efficacy of TCS001-SC to promote the growth of strawberries and to prevent and control strawberry anthracnose was evaluated. The optimal liquid fermentation condition for TCS001 was determined to be 2.89 % soluble peanut cake powder, 3.0 % glucose, 3.0 % soluble starch, 0.002 % FePO4, 0.006 % KCl, 0.6 % NaCl, 0.05 % MgCl2·6H2O, 0.3 % K2HPO4, 0.15 % KH2PO4, 0.05 % CaCO3, 0.005 % MnSO4, a working volume of 31 % (77.5 mL/250 mL), a rotation speed of 173 r/min, a cultivation temperature of 28°C, an inoculum volume of 1.0 %, and a pH of 7.0. The 15 L fermenter upscale culture achieved a spore count of 9.46 × 109 CFU/mL, which was 2.01 times the spore count of 4.7 × 109 CFU/mL before optimization, and a preliminary TCS001-SC was developed with the fermented broth as the main component. Agar plate confrontation tests showed that TCS001 had antifungal activity against five types of anthracnose fungi, with inhibition rates ranging from 70.3 % to 87.2 %. TCS001-SC could promote the growth of strawberries and induce a rapid defense enzyme response in their leaves, enhancing the plant's resistance to pathogens. After treatment, individual strawberry plants showed significant increases in the number of leaves, fresh weight of stems and leaves, root fresh weight, leaf area, plant height, and the content of POD, SOD, CAT enzymes, GA, IAA, and ABA compared to the control.Additionally, it shows good prevention and control effects against strawberry anthracnose, with a control efficacy of 60.45% after five spray treatments at a concentration of 2 × 107 CFU/mL, which is not significantly different from the efficacy of commercial microbial agents such as Bacillus subtilis wettable powder, subsequent field trials will be conducted to determine its potential as a microbial pesticide. This study provides important support for the future industrial production and application of the strain TCS001.

Preparation and flavor characteristics of plant-based meat flavoring by mixed fermentation of multiple strains

In this study, plant-based meat flavoring was prepared by mixed fermentation and thermal reaction. Based on the changes of amino acid nitrogen, reducing sugar, and total acid during the fermentation process and combined with sensory evaluation, the optimal fermentation conditions were determined as 4 % total inoculation amount of Monascus purpureus, Actinomucor elegans, and Bacillus subtilis with an inoculation ratio of 1:1:1, and 36 h of fermentation time. The odor characterization of meat flavoring was identified by solid-phase microextraction–gas chromatography–mass spectrometry (SPME–GC–MS), aroma extract dilution analysis (AEDA), quantitative measurements, odor activity value (OAV) and aroma recombination and omission experiments. 64 volatile compounds and 28 odorants were identified. The compounds with FD ≥ 27 were precisely quantified by five-point standard calibration curve and OAVs were calculated, and those with OAV ≥ 1 were subjected to aroma recombination and omission experiments. Finally, furfuryl mercaptan, difurfuryl disulfide, 4-methyl-5-ethylthiazole, 1-octanal, and 2-acetylthiophene were determined to be the key aroma-active compounds. These compounds have been recognized as key aroma-active compounds or detected in other meats or meat flavorings. In this study, we prepared plant-based meat flavoring through mixed fermentation, and provide new ideas for the development of plant-based thermal reaction meat flavoring process.

Optimization of Liquid Fermentation of Acanthopanax senticosus Leaves and Its Non-Targeted Metabolomics Analysis

To enhance the nutritional value of Acanthopanax senticosus leaves (AL), a fermentation process was conducted using a probiotic Bacillus mixture, and the changes in chemical constituents and biological activities before and after fermentation were compared. A response surface methodology was employed to optimize the liquid fermentation conditions of AL based on their influence on polyphenol content. Non-targeted metabolomics analysis was performed using LC-MS/MS to reveal the differing profiles of compounds before and after fermentation. The results indicated that Bacillus subtilis LK and Bacillus amyloliquefaciens M2 significantly influenced polyphenol content during fermentation. The optimal fermentation conditions were determined to be a fermentation time of 54 h, a temperature of 39.6 °C, and an inoculum size of 2.5% (v/v). In comparison to unfermented AL, the total polyphenol and flavonoid contents, as well as the free radical scavenging capacities measured by DPPH and ABTS assays, and the activities of β-glucosidase and endo-glucanase, were significantly increased. The non-targeted metabolomics analysis identified 1348 metabolites, of which 829 were classified as differential metabolites. A correlation analysis between the differential metabolites of polyphenols, flavonoids, and antioxidant activity revealed that 13 differential metabolites were positively correlated with antioxidant activity. Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis of the differential metabolites identified 82 pathways, with two of the top 25 metabolic pathways related to flavonoids. This study explores the potential for enhancing the active ingredients and biological effects of AL through probiotic fermentation using Bacillus strains.

Optimization, purification and characterization of laccase from a new endophytic Trichoderma harzianum AUMC14897 isolated from Opuntia ficus-indica and its applications in dye decolorization and wastewater treatment.

Hazardous synthetic dye wastes have become a growing threat to the environment and public health. Fungal enzymes are eco-friendly, compatible and cost-effective approachfor diversity of applications. Therefore, this study aimed to screen, optimize fermentation conditions, and characterize laccase from fungal endophyte with elucidating its ability to decolorize several wastewater dyes. A new fungal endophyte capable of laccase-producing was firstly isolated from cladodes of Opuntia ficus-indica and identified as T. harzianum AUMC14897 using ITS-rRNA sequencing analysis. Furthermore, the response surface methodology (RSM) was utilized to optimize several fermentation parameters that increase laccase production. The isolated laccase was purified to 13.79-fold. GFC, SDS-PAGE revealed laccase molecular weight at 72kDa and zymogram analysis elucidated a single band without any isozymes. The peak activity of the pure laccase was detected at 50°C, pH 4.5, with thermal stability up to 50°C and half life span for 4h even after 24h retained 30% of its activity. The Km and Vmax values were 0.1mM, 22.22µmol/min and activation energy (Ea) equal to 5.71kcal/mol. Furthermore, the purified laccase effectively decolorized various synthetic and real wastewater dyes. Subsequently, the new endophytic strain produces high laccase activity that possesses a unique characteristic, it could be an appealing candidate for both environmental and industrial applications.