Production and Characterization of New Fibrinolytic Protease from Mucor subtillissimus UCP 1262 in Solid-State Fermentation

Fibrinolytic enzymes have received attention regarding their medicinal potential for thrombolytic diseases, a leading cause of morbidity and mortality worldwide. Various natural enzymes purified from animal, plant and microbial sources have been extensively studied. The aim of this work was to produce fibrinolytic protease by solid state fermentation using agro industrial substrates. Rhizopus arrhizus var. arrhizus UCP 1295 and Mucor subtillissimus UCP 1262 filamentous fungi species isolated from soil of Caatinga-PE, Brasil, were used as producer microorganisms. Wheat bran was shown to be the best substrate for the production of the enzyme and by using a 23 full factorial design the main effects and interactions of the quantity of the substrate wheat bran, moisture and temperature on the fibrinolytic enzyme production and protease were evaluated. The best results for fibrinolytic and protease activities, 144.58 U/mL and 48.33 U/mL, respectively, were obtained with Mucor subtillissimus UCP 1262 using as culture medium 3 g wheat bran, 50% moisture at a temperature of 25°C for 72 hours. The optimum temperature for the produced enzyme was 45°C and most of its original activity was retained after being subjected to 80°C for 120 min. The protease activity was enhanced by K+, Ca+ and Mn+;but with Cu+ there was an inhibition. The specificity to chromogenic substrate and the inhibition by PMSF indicates that it is a chymotrypsin-like serine protease. Presented results suggest that this enzyme produced by solid-state fermentation is an interesting alternative as a candidate for thrombolytic therapy.

Direct Solid-State Fermentation of Soybean Processing Residues for the Production of Fungal Chitosan by Mucor rouxii

The feasibility of utilizing soybean-processing residues such as soybean meal and hulls as substrates for chitosan production by the fungus Mucor rouxii ATCC 24905 via solid-state fermentation (SSF) was investigated. The effects of the type of soybean-based substrate, length of cultivation period, substrate moisture content, substrate pH, incubation temperature and extraction conditions on chitosan yield were determined. The results showed that a maximum fungal chitosan yield of up to 3.44% by dry substrate weight (34.4 g/kg) could be achieved using a pure soybean meal substrate with an initial moisture content of 50% (w/w) and pH of 5 - 6 incubated for six days at 25&degC. A more severe heat treatment (autoclaving vs. refluxing) resulted in higher chitosan extraction yields regardless of the strength of extraction reagents. Fourier transform infrared (FTIR) analysis of the fungal chitosan revealed its degree of deacetylation (DDA) to be between 55% and 60%.

Structure and immunomodulatory activity of Lentinus edodes polysaccharides modified by probiotic fermentation

Plant-based fermentations provide an untapped source for novel biotechnological applications.In this study,a probiotic named Lactobacillus fermentum 21828 was introduced to ferment Lentinus edodes.Polysaccharides were extracted from fermented and non-fermented L.edodes and purified via DEAE-52 and Sephadex G-100.The components designated F-LEP-2a and NF-LEP-2a were analyzed by FT-IR,HPGPC,HPAEC,SEM,GC-MS and NMR.The results revealed that probiotic fermentation increased the molecular weight from 1.16×10^(4) Da to 1.87×10^(4) Da and altered the proportions of glucose,galactose and mannose,in which glucose increased from 45.94%to 48.16%.Methylation analysis and NMR spectra indicated that F-LEP-2a and NF-LEP-2a had similar linkage patterns.Furthermore,their immunomodulatory activities were evaluated with immunosuppressive mice.NF-LEP and F-LEP improved immune organ indices,immunoglobulin(Ig G and Ig M)and cytokines concentrations;restored the antioxidation capacity of liver;and maintained the balance of gut microbiota.F-LEP displayed better moderating effects on the spleen index,immunoglobulin,cytokines and the diversity of gut microbiota than NF-LEP(200,400 mg/kg).Our study provides an efficient and environment-friendly way for the structural modification of polysaccharides,which helps to enhance their biological activity and promote their wide application in food,medicine and other fields.

The Advantages of Methane Production by Combined Fermentation of Lignite and Wheat Straw

Biogasification of coal is important for clean utilization of coal. Experiments on the fermentation of single lignite, single straw and their mixture were performed to explore the variation characteristics of gas production potential, microbial community and methanogenic metabolic pathways of mixture. Research has shown that mixed fermentation of lignite and straw significantly promoted biomethane production. The abundance of hydrolytic acidifying functional bacteria genera (Sphaerochaeta, Lentimicrobium) in mixed fermentation was higher than that in the fermentation of single lignite and single straw. The abundance of some key CAZy metabolic enzyme gene sequences in mixed fermentation group was increased, which was favorable to improve methane production. Aceticlastic methanogenesis was the most critical methanogenic pathway and acetic acid pathway was more competitive in methanogenic mode during peak fermentation. Macrogenomics provided theoretical support for the claim that mixed fermentation of coal and straw promoted biomethane metabolism, which was potentially valuable in expanding methanogenesis from mixed fermentation of lignite with different biomasses.

Postbiotics from Saccharomyces cerevisiae fermentation stabilize microbiota in rumen liquid digesta during grain-based subacute ruminal acidosis(SARA) in lactating dairy cows

Background Subacute ruminal acidosis(SARA)is a common metabolic disorder of high yielding dairy cows,and it is associated with dysbiosis of the rumen and gut microbiome and host inflammation.This study evaluated the impact of two postbiotics from Saccharomyces cerevisiae fermentation products(SCFP)on rumen liquid associated microbiota of lactating dairy cows subjected to repeated grain-based SARA challenges.A total of 32 rumen cannulated cows were randomly assigned to 4 treatments from 4 weeks before until 12 weeks after parturition.Treatment groups included a Control diet or diets supplemented with postbiotics(SCFPa,14 g/d Original XPC;SCFPb-1X,19 g/d Nutri Tek;SCFPb-2X,38 g/d Nutri Tek,Diamond V,Cedar Rapids,IA,USA).Grain-based SARA challenges were conducted during week 5(SARA1)and week 8(SARA2)after parturition by replacing 20%DM of the base total mixed ration(TMR)with pellets containing 50%ground barley and 50%ground wheat.Total DNA from rumen liquid samples was subjected to V3–V416S r RNA gene amplicon sequencing.Characteristics of rumen microbiota were compared among treatments and SARA stages.Results Both SARA challenges reduced the diversity and richness of rumen liquid microbiota,altered the overall composition(β-diversity),and its predicted functionality including carbohydrates and amino acids metabolic pathways.The SARA challenges also reduced the number of significant associations among different taxa,number of hub taxa and their composition in the microbial co-occurrence networks.Supplementation with SCFP postbiotics,in particular SCFPb-2X,enhanced the robustness of the rumen microbiota.The SCFP supplemented cows had less fluctuation in relative abundances of community members when exposed to SARA challenges.The SCFP supplementation promoted the populations of lactate utilizing and fibrolytic bacteria,including members of Ruminococcaceae and Lachnospiraceae,and also increased the numbers of hub taxa during non-SARA and SARA stages.Supplementation with SCFPb-2X prevented the fluctuations in the abundances of hub taxa that were positively correlated with the acetate concentration,andα-andβ-diversity metrics in rumen liquid digesta.Conclusions Induction of SARA challenges reduced microbiota richness and diversity and caused fluctuations in major bacterial phyla in rumen liquid microbiota in lactating dairy cows.Supplementation of SCFP postbiotics could attenuate adverse effects of SARA on rumen liquid microbiota.

Impact of sourdough fermentation on nutrient transformations in cereal-based foods:Mechanisms,practical applications,and health implications

Sourdough is often considered a healthy choice and quality improver for use in cereal production due to its unique microbial composition and fermentation properties.During sourdough fermentation of cereals,biotransformation of nutrients occurs,resulting in notable changes to proteins,carbohydrates,fats,vitamins,and minerals.Each nutrient undergoes specific transformations,providing various advantages for human health.Proteins undergo hydrolysis to produce small molecular weight peptides and amino acids that are more easily digested and absorbed by the human body.Carbohydrates break down to improve the digestibility and absorption of cereals and lower the glycemic index.Fatty acids experience oxidation to produce new substances with health benefits.Additionally,the application of sourdough fermentation can enhance the texture,flavor,and nutritional value of cereal foods while also extending their shelf life and improving food safety.In conclusion,sourdough fermentation has a broad range of applications in cereal food processing.Further research is encouraged to investigate the mechanisms and processes of sourdough fermentation to develop even more nutritious,healthy,and flavorful cereal-based foods.

Allergen degradation of bee pollen by lactic acid bacteria fermentation and its alleviatory effects on allergic reactions in BALB/c mice

Food allergy as a global health problem threatens food industry.Bee pollen(BP)is a typical food with allergenic potentials,although it performs various nutritional/pharmacological functions to humans.In this study,lactic acid bacteria(LAB)were used to ferment Brassica napus BP for alleviating its allergenicity.Four novel allergens(glutaredoxin,oleosin-B2,catalase and lipase)were identified with significant decreases in LAB-fermented BP(FBP)than natural BP by proteomics.Meanwhile,metabolomics analysis showed significant increases of 28 characteristic oligopeptides and amino acids in FBP versus BP,indicating the degradation of LAB on allergens.Moreover,FBP showed alleviatory effects in BALB/c mice,which relieved pathological symptoms and lowered production of allergic mediators.Microbial high-throughput sequencing analysis showed that FBP could regulate gut microbiota and metabolism to strengthen immunity,which were closely correlated with the alleviation of allergic reactivity.These findings could contribute to the development and utilization of hypoallergenic BP products.

Microbiomics and metabolomics insights into the microbial regulation on the formation of flavor components in the traditional fermentation process of Chinese Hongqu aged vinegar

This study aimed to investigate microbial succession and metabolic dynamics during the traditional fermentation of Hongqu aged vinegar,and explore the core functional microbes closely related to the formation of flavor components.Microbiome analysis demonstrated that Lactobacillus,Acetobacter,Bacillus,Enterobacter,Lactococcus,Leuconostoc and Weissella were the predominant bacterial genera,while Aspergillus piperis,Aspergillus oryzae,Monascus purpureus,Candida athensensis,C.xylopsoci,Penicillium ochrosalmoneum and Simplicillium aogashimaense were the predominant fungal species.Correlation analysis revealed that Acetobacter was positively correlated with the production of tetramethylpyrazine,acetoin and acetic acid,Lactococcus showed positive correlation with the production of 2-nonanone,2-heptanone,ethyl caprylate,ethyl caprate,1-hexanol,1-octanol and 1-octen-3-ol,C.xylopsoci and C.rugosa were positively associated with the production of diethyl malonate,2,3-butanediyl diacetate,acetoin,benzaldehyde and tetramethylpyrazine.Correspondingly,non-volatile metabolites were also detected through ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry.A variety of amino acids and functional dipeptides were identified during the traditional brewing of Hongqu aged vinegar.Correlation analysis revealed that Lactobacillus was significantly associated with DL-lactate,indolelactic acid,D-(+)-3-phenyllactic acid,pimelic acid,pregabalin and 3-aminobutanoic acid.This study is useful for understanding flavor formation mechanism and developing effective strategies for the suitable strains selection to improve the flavor quality of Hongqu aged vinegar.

Study on Solid Fermentation and Antioxidant Function of Natto

[Objectives]To study the optimum conditions of solid fermentation of natto with antioxidant function as an index.[Methods]Single factor experiment and orthogonal experiment were designed to study the effects of temperature,time,initial pH and inoculum amount on the antioxidant activity of natto solid fermentation.The optimum conditions of natto solid fermentation were determined and the antioxidant ac-tivity of natto extract was compared.[Results]The optimal fermentation conditions were as follows:temperature 32℃,initial pH 7.0,inocu-lation amount 8%,fermentation time 32 h.The hydroxyl radical scavenging rate of natto solid fermentation crude extract was the highest,which was 82.7%.The optimized nato fermentation extract showed stronger scavenging ability for-OH and O,:,and showed obvious dose-effect relationship.ICso was 3.63 and 4.24 mg/mL,respectively,and the scavenging efficiency was 1.3 and 1.9 times higher than that of the unoptimized fermentation extract,respectively.[Conclusions]Natto is rich in nattokinase and other functional factors,and its antioxidant ac-tivity can be improved by optimizing fermentation technology,so that natto products can be widely used,including cosmetic raw materials,nat-to skin care soap,health food and medicine,etc.,and have a broader development prospect.

Process Analysis of Mobile Fermentation of Spent White Hypsizygus marmoreus Substrate with Self-made Mobile Fermentation Bag

The self-made mobile fermentation bag for spent white Hypsizygus marmoreus substrate was developed, and the relevant fermentation process was studied. The results showed that under the condition of single addition of Lactobacillus,nitrogen-free extract was degraded into lactic acid, leading to decreased pH value.On day 10, the fermentation effect reached the best with Lactobacillus abundance of 5.12×10~7/ml, lactic acid content of 0.48% and strong acid flavor. At this time, the fermentation material was moist without mildew and agglomeration, and was suitable for livestock and poultry. However, after 10 days, undesirable acids and mildew generated, a large amount of lactic acid bacteria died, and the fermentation material turned black and agglomerated, and became unsuitable for feeding livestock and poultry. Throughout the fermentation process, the pH value first decreased continuously until to 4.0, and then remained stable. During the fermentation of spent white H. marmoreus substrate, the nitrogen-free extract and crude fiber contents decreased, the crude protein content increased, while other indicators remained unchanged.

Effects of Rosa roxburghii&edible fungus fermentation broth on immune response and gut microbiota in immunosuppressed mice

With the rise of probiotics fermentation in food industry,fermented foods have attracted worldwide attention.In this study,protective effects of Rosa roxburghii&edible fungus fermentation broth(REFB)on immune function and gut health in Cyclophosphamide induced immunosuppressed mice were investigated.Results showed that REFB could improve the immune organ index,and promote the proliferation and differentiation of splenic T lymphocytes.In addition,it attenuated intestinal mucosal damage and improved intestinal cellular immunity.REFB administration also up-regulated the expression of IL-4,INF-γ,TNF-α,T-bet and GATA-3 mRNA in small intestine.Furthermore,administration of REFB modulated gut microbiota composition and increased the relative abundance of beneficial genus,such as Bacteroides.It also increased the production of fecal short-chain fatty acids.These indicate that REFB has the potential to improve immunity,alleviate intestinal injury and regulate gut microbiota in immunosuppressed mice.

一株毛霉属真菌Mucor hiemalis发酵液的化学成分研究

研究一株毛霉属真菌Mucor hiemalis发酵液的化学成分。运用ITS序列分析法进行分子生物学鉴定,并使用MEGA7.0软件构建NJ(Neighbor-Joining)系统发育进化树,最终鉴定该菌株为Mucor hiemalis。利用正相硅胶、Sephadex LH-20凝胶柱层析进行分离纯化,通过核磁共振波谱和高分辨质谱等技术进行结构鉴定。从Mucor hiemalis发酵液中分离鉴定了6个化合物,分别为油酸(1),2-甲氧基乙基乙酸酯(2),邻苯二甲酸二丁酯(3),(22E,24R)-5α,8α-过氧麦角甾-6,22-二烯-3β-醇(4),亚油酸甘油三酯(5)和2,3-丁二醇(6)。以上所有化合物均首次从该菌中分离得到。其中,(22E,24R)-5α,8α-过氧麦角甾-6,22-二烯-3β-醇(4)是一种甾体类化合物,对细胞炎症表现出显著的抗炎活性,这将对该菌株后期的综合利用与开发提供了一定理论基础。

Mucor sp.JX23发酵液生物催化肉桂醛选择加氢制肉桂醇

研究了Mucor sp.JX23发酵液直接生物催化肉桂醛选择加氢制肉桂醇的反应,并考察了肉桂醛质量浓度、反应pH值、温度、时间等因素对反应转化率和选择性的影响。结果表明,Mucor sp.JX23发酵液的最适催化条件为:肉桂醛质量浓度3g/L,反应体系的初始pH=6.0,28℃反应70h。在此条件下,肉桂醛的转化率为82.9%,肉桂醇的选择性为90.4%。

γ-亚麻酸产生菌Mucor sp. EIM-10的筛选及分子鉴定

为了获得高产γ-亚麻酸(γ-linolenic acid,GLA)的菌株,利用苏丹黑染色法筛选获得1株GLA产生菌EIM-10,通过摇瓶培养,其生物量可达11.882 g/L,菌丝体油脂含量可达18.86%。气相色谱-质谱(GC-MS)分析表明其γ-亚麻酸质量分数(占总脂肪酸)高达27.68%。为进一步鉴定该菌株,克隆测定了该菌18S rRNA基因序列,并对其进行系统进化树分析,结果表明该菌属于毛霉属,与Mucor racemosus、Mucor plumbeus、Mucor ramosissimus与Mucorcircinelloides同属一个分支。

Mucor sp. JX23发酵液生物催化肉桂酸降解生成苯乙酮

对以Mucor sp. JX23发酵液生物催化肉桂酸降解生成苯乙酮的反应进行了研究。考察了肉桂酸浓度、反应体系pH值、反应温度和反应时间对反应的影响。结果表明:以Mucor sp.JX23为催化剂的反应具有高转化率和高选择性。在肉桂酸浓度为5.0g/L,反应体系pH=6.0,反应温度28-30℃,反应时间2d的条件下,肉桂酸的转化率为89.2%,苯乙酮的产量达3.1g/L,没有发现其他副产物。

Emerging peril of post-dengue mucormycosis:A case report

Rationale:Dengue fever is a leading cause of death in tropical and subtropical countries.Although most patients have a self-limited febrile illness,the viral infection can induce virus-mediated host changes,making immunocompetent persons susceptible to deadly fungal infections.However,there are only a few reports of such an association.Here we present a case of this deadly co-infection.Patient’s Concern:A 17-year-old male patient was diagnosed with dengue fever.He presented to us with facial swelling,periorbital edema,and black discoloration over the palate during the second week of his illness.Diagnosis:Diagnostic tests confirmed the presence of fungal hyphae.A diagnosis of post-dengue mucormycosis was made.No other comorbidity or underlying immune deficit was detected.Interventions:The patient underwent surgical debridement and antifungal treatment.Outcomes:The patient recovered and showed signs of palatal healing with an advancing mucosal edge.Lessons:Dengue virus and mucor co-infection has brought to light a new pathogenic paradigm.Clinicians need to be aware of this emerging medical condition and maintain a high index of suspicion for mucor co-infections while treating dengue patients.

Medicinal Solid Fermentation Engineering of Chinese Traditional Medicinal Fungal

Medicinal solid fermentation technology of fungal was unique within the traditional Chinese medicine （TCM） and one of the oldest bio-pharmaceutical technology in the history, for example, the production of medicated leaven named ＂Shenqu＂, the history of which had continued for more than 1,500 years. These kinds of productions were widly used and had a close relationship with TCM. The development of this technology had through stages of ＂yeast production＂, ＂solid culture＂, ＂solid fermentation＂, and nowadays toward a series engineering of solid fermentation with promising prospects. The author described the development of its history briefly, and the representative products and the characteristics of production process of every stage, respectively. This research focused on taking ＂Trametes robinioplila fermented substance＂, ＂T. robinioplila ＆ A. membranaceus fermented substance＂ and ＂T. wilfordii Hook.f. ＆ G. lucidum fermented substance＂ as examples to introduce R ＆ D ideas and important findings of＂common fermentation＂ and ＂bi-directional fermentation＂ which were recommended as studying hotspots. There were many unique and important functions in the fields of TCM about all kinds of＂fermentation substances＂. It is infered that there may exist some other innovate technologies such as ＂Multi-fungal fermentation＂, ＂Special elements fermentation＂, etc. which are remained to be researched and developed continuely. All kinds of＂fermentation substances＂ are looked forward to show unique and important roles in the fields of TCM.

一株产纤维素酶真菌Mucor amphibiorum RSC1的分离鉴定及其产酶培养基的初步优化

[目的]对一株产纤维素酶真菌进行分离鉴定,以期为产纤维素酶菌株种类的丰富提供试验依据。[方法]通过刚果红平板染色法初筛和摇瓶发酵产酶复筛从农田稻草堆的土壤中筛选出可产纤维素酶的真菌菌株,并对其发酵产酶培养基进行了单因素试验优化,同时对该菌进行了形态学和18S rRNA分子生物学鉴定。[结果]分离得到的菌株为1株产纤维素酶丝状真菌RSC1,发酵产酶培养基优化结果显示,培养基中含1.3%CMC-Na,0.1%酵母膏和0.1%MgSO4时其产酶活性最高。对该菌进行的形态学和分子生物学鉴定结果表明,丝状真菌RSC1与毛霉属真菌Mucor amphibiorum相似度为97%,故将该丝状真菌命名为Mucor amphibiorum RSC1。[结论]该研究鉴定了所分离纯化的菌株为Mucor amphibiorum RSC1,该结果为产纤维素酶菌株种类的丰富奠定了基础。

Influence of nitrogen status on fermentation performances of non-Saccharomyces yeasts:a review

Nitrogen,one of the most crucial nutrients present in grapes and musts,plays a key role in yeast activities during alcoholic fermentation.Such influences are imposed on yeast growth and fermentation performances including the formation of secondary metabolites.Saccharomyces cerevisiae,the main yeast responsible for fermentation,has been studied extensively regarding nitrogen impacts.On the other hand,a similar study for non-Saccharomyces yeasts,whose contributions to winemaking have gradually been acknowledged,remains to be fully explored,with a few studies being reported.This review starts by discussing nitrogen impacts on non-Saccharomyces yeast growth and fermentation kinetics in different case scenarios,then proceeds to summarize the nitrogen preferences of individual yeast strains with regulation mechanisms elucidated by recent studies.Detailed discussions on the influences on the production of volatile compounds and proposed pathways therein are made,followed by future work suggested as the final section.In summarizing the nitrogen impacts on non-Saccharomyces yeasts throughout alcoholic fermentation,this review will be helpful in obtaining a more comprehensive view on these non-conventional wine yeasts in terms of nutrient requirements and corresponding volatile production.Research gaps will therefore be elucidated for future research.

The Influence of Gratering, Fermentation and Variety on the Physio-Chemical Quality of Cassava Starch

The study was conducted to determine the influence of gratering and fermentation parameters on the physicochemical quality of starch obtained from two cassava varieties in Sierra Leone (i.e., SLICASS 11 and SLICASS 6). Fresh cassava roots harvested from the Department of Agricultural Engineering were peeled, washed and grated before fermentation and starch extraction. Fermentation was carried out under separate aerobic and anaerobic conditions for a period of Seven days. Physicochemical analyses were conducted at the Postharvest Food and Bioprocess Engineering Laboratory of the Department of Agricultural and Bio-System Engineering, Njala University to determine the solubility, water absorption capacity and swelling power of starch extracts obtained from various experimental treatments. Fermentation method and duration had significant effects on the solubility, water absorption capacity (WAC) and swelling power (SP) of starch extracts. Maximum solubility and WAC were recorded on the fifth day, for both cassava varieties tested, with apparent significant difference resulting from the two gratering bands (i.e., with 1.5-inch nail hole and 4-inch nail hole sizes, respectively). While swelling power increased consistently with temperature for starch obtained from SLICASS-11 variety, an irregular pattern was observed for SLICASS-6 variety. A multiple correlation analysis proposes a significant and weak correlation between temperature, WAC (+0.150) and swelling power (+0.048). Also multiple correlation analyses suggest a significant correlation between fermentation period, the functional properties of starch extracts obtained from both fermentation methods and cassava varieties (i.e., solubility (−0.226), water absorption capacity (+0.301) and swelling power (+0.329)).