ABSTRACT
Chitinases are promising enzymes for a multitude of applications, including chitooligosaccharide (COS) synthesis for food and pharmaceutical uses and marine waste management. Owing to fungal diversity, fungal chitinases may offer alternatives for chitin degradation and industrial applications. The rapid reproduction cycle, inexpensive growth media, and ease of handling of fungi may also contribute to reducing enzyme production costs. Thus, this study aimed to identify fungal species with chitinolytic potential and optimize chitinase production by submerged culture and enzyme characterization using shrimp chitin. Three fungal species, Coriolopsis byrsina, Trichoderma reesei, and Trichoderma harzianum, were selected for chitinase production. The highest endochitinase production was achieved in C. byrsina after 168 h cultivation (0.3 U mL- 1). The optimal temperature for enzyme activity was similar for the three fungal species (up to 45 and 55 ºC for endochitinases and exochitinases, respectively). The effect of pH on activity indicated maximum hydrolysis in acidic pH (4-7). In addition, the crude T. reesei extract showed promising properties for removing Candida albicans biofilms. This study showed the possibility of using shrimp chitin to induce chitinase production and enzymes that can be applied in different industrial sectors.
ABSTRACT
The increase in the generation of chicken feathers, due to the large production of the poultry industry, has created the need to search for ecologically safer ways to manage these residues. As a sustainable alternative for recycling keratin waste, we investigated the ability of the bacterium Ochrobactrum intermedium to hydrolyze chicken feathers and the valorization of the resulting enzymes and protein hydrolysate. In submerged fermentation with three different inoculum sizes (2.5, 5.0, and 10.0 mg of bacterial cells per 50 mL of medium), the fastest degradation of feathers was achieved with 5.0 mg cells, in which a complete decomposition of the substrate (96 h) and earlier peaks of keratinolytic and caseinolytic activities were detected. In the resulting protein hydrolysate, we noticed antioxidant and Fe2+ and Cu2+ chelating activities. ABTS scavenging, Fe3+-reducing ability and metal chelating activities of the fermentative samples followed the same trend of feather degradation; as feather mass decreased in the media, these activities increased. Furthermore, we noticed about 47% and 60% dispersion of established 7-day biofilms formed by S. aureus after enzymatic treatment for 5 h and 24 h, respectively. These findings highlight the potential use of this bacterium as an environmentally friendly alternative to treat this poultry waste and offer valuable products.
ABSTRACT
For 2G ethanol production, pentose fermentation and yeast tolerance to lignocellulosic hydrolyzate components are essential to improve biorefinery yields. Generally, physicochemical pre-treatment methodologies are used to facilitate access to cellulose and hemicellulose in plant material, which consequently can generate microbial growth inhibitory compounds, such as furans, weak acids, and phenolic compounds. Because of the unsatisfactory yield of wild-type Saccharomyces cerevisiae during pentose fermentation, the search for xylose-fermenting yeasts tolerant to microbial growth inhibitors has gained attention. In this study, we investigated the ability of the yeasts Pichia guilliermondii G1.2 and Candida oleophila G10.1 to produce ethanol from xylose and tolerate the inhibitors furfural, 5-hydroxymethylfurfural (HMF), acetic acid, formic acid, ferulic acid, and vanillin. We demonstrated that both yeasts were able to grow and consume xylose in the presence of all single inhibitors, with greater growth limitation in media containing furfural, acetic acid, and vanillin. In saline medium containing a mixture of these inhibitors (2.5-3.5 mM furfural and HMF, 1 mM ferulic acid, 1-1.5 mM vanillin, 10-13 mM acetic acid, and 5-7 mM formic acid), both yeasts were able to produce ethanol from xylose, similar to that detected in the control medium (without inhibitors). In future studies, the proteins involved in the transport of pentose and tolerance to these inhibitors need to be investigated.
Subject(s)
Furans , Xylose , Xylose/metabolism , Furans/metabolism , Ethanol/metabolism , Pichia/metabolism , Furaldehyde/pharmacology , Biomass , Saccharomyces cerevisiae/metabolism , Pentoses/metabolism , Fermentation , Phenols/metabolism , Formates/metabolismABSTRACT
The intestinal microbiota plays an important role in the immune response against viral infections, modulating both innate and adaptive immune responses. The cytokine storm is associated with COVID-19 severity, and the patient's immune status is influenced by the intestinal microbiota in a gut-lung bidirectional interaction. In this study, we evaluate the intestinal microbiota of Brazilian patients in different post-COVID-19 periods, and correlate this with clinical data and the antibiotic therapy used during the acute phase. DNA extracted from stool samples was sequenced and total anti-SARS-CoV-2 antibodies and C-reactive protein were quantified. Compared with controls, there were significant differences in the microbiota diversity in post-COVID-19 patients, suggesting an intestinal dysbiosis even several months after acute disease resolution. Additionally, we detected some genera possibly associated with the post-COVID-19 dysbiosis, including Desulfovibrio, Haemophillus, Dialister, and Prevotella, in addition to decreased beneficial microbes, associated with antibiotic-induced dysbiosis, such as Bifidobacterium and Akkermansia. Therefore, our hypothesis is that dysbiosis and the indiscriminate use of antibiotics during the pandemic may be associated with post-COVID-19 clinical manifestations. In our study, 39% (n = 58) of patients reported symptoms, including fatigue, dyspnea, myalgia, alopecia, anxiety, memory loss, and depression. These data suggest that microbiota modulation may represent a target for recovery from acute COVID-19 and a therapeutic approach for post-COVID-19 sequelae.
Subject(s)
COVID-19 , Gastrointestinal Microbiome , Acute Disease , Dysbiosis/microbiology , Humans , PandemicsABSTRACT
Xylooligosaccharides (XOs) are a promising class of prebiotics capable of selectively stimulating the growth of the beneficial intestinal microbiota against intestinal pathogens. They can be obtained from xylan present in residual lignocellulosic material from agriculture. Thus, in this study we produced XOs by extracting xylan from sugarcane bagasse and hydrolyzing it using the GH10 xylanase from Thermoascus aurantiacus expressed by Pichia pastoris. An alkaline method to extract xylan is described, which resulted in 83.40% of xylan recovery and low amounts of cellulose and lignin. The enzymatic hydrolysate exhibited a mixture of XOs containing mainly xylobiose, xylotriose and xylotetraose. These oligosaccharides stimulated the growth of Lactobacillus casei, L. rhamnosus, L. fermentum and L. bulgaricus strains, which were able to produce organic acids, especially acetic acid. These findings demonstrate the possibility to redirect crop by-products to produce XOs and their use as a supplement to stimulate the growth of probiotic strains.
Subject(s)
Probiotics , Saccharum , Thermoascus , Cellulose , Endo-1,4-beta Xylanases/genetics , Glucuronates , Hydrolysis , Oligosaccharides , XylansABSTRACT
ß-Glucosidases have been extensively investigated to integrate the enzyme complex for cellulose fiber saccharification and for improving the aroma of wine. To produce these enzymes, greater attention has been given to filamentous fungi and bacteria, and few investigations have targeted the potential applications of enzymes secreted by yeasts. Addressing this issue, in this study, ß-glucosidases were produced by the Pichia ofunaensis and Trichosporon multisporum yeasts, via solid state fermentation with wheat bran as a substrate. When using p-Nitrophenyl ß-d-glucopyranoside (pNPG) as an enzyme substrate, maximum ß-glucosidase activities were detected at pH 5.5-6.0 and 50-60 °C for P. ofunaensis, and pH 5-6 and 55 °C for T. multisporum. Both enzymes were able to hydrolyze cellobiose and exhibited stability over a wide range of pH (3.5-9.0) for 24 h at 4 °C, thermostability up to 50 °C for 1 h and tolerance to 10 mM phenolic compounds. Negative modulation on enzyme activity was observed in the presence of Cu2+, Fe3+, Zn2+, Al3+ and Hg2+, while both ß-glucosidases were tolerant to 30% methanol, isopropanol and acetone. In the presence of ethanol and glucose, enzymes from P. ofunaensis were the more active and stable of the two. These enzymes, especially the P. ofunaensis ß-glucosidases, could be tested in enology for improving the aroma of wine and for integrating a cellulolytic complex to produce 2G ethanol.
ABSTRACT
Yeast isolates from flowers and fruits from a Brazilian forest were studied. The yeasts were identified at species and strain level by PCR-RFLP and PCR-RAPD, respectively. The 46 isolated yeasts were classified into 11 different species belonging to the genera Candida, Diutina, Hanseniaspora, Meyerozyma, Pichia, Rhodotorula, and Torulaspora. A total of 20 different strains were found. In order to ascertain the probiotic potential, the resistance to gastrointestinal conditions, autoaggregation, and hydrophobicity assays were studied, along with the capacity to form biofilm. The results indicate that, although most of the strains presented better results than Saccharomyces boulardii (the only strain recognized as a probiotic yeast), four strains were the most promising, namely, Rhodotorula mucilaginosa 32, Meyerozyma caribbica 35, and Diutina rugosa 12 and 45, according to the Duncan test. Several biotechnological properties were evaluated. D. rugosa inhibited Dekkera bruxellensis. The assimilation or fermentation of seven sugars was tested, and only five of the yeasts did not show a capacity to assimilate any of the sugars under aerobic conditions. However, all strains were able to ferment at least one of the sugars under anaerobic conditions. As far as enzyme production is concerned, positive results were only found for the enzymes' amylase, pectinase, and protease. D. rugosa 42 and Hanseniaspora opuntiae 18, followed of Pichia kluyveri 26, showed high values for the production of melatonin. In conclusion, the results of this study show that several non-Saccharomyces present probiotic characteristics, and these have good potential for industrial applications in the food or biotechnology industries.
Subject(s)
Biotechnology , Ecosystem , Fruit , Probiotics , Biotechnology/trends , Fermentation , Fruit/microbiology , Random Amplified Polymorphic DNA Technique , Sugars/metabolism , Yeasts/geneticsABSTRACT
Dysbiosis, associated with barrier disruption and altered gut-brain communications, has been associated with multiple sclerosis (MS). In this study, we evaluated the gut microbiota in relapsing-remitting patients (RRMS) receiving disease-modifying therapies (DMTs) and correlated these data with diet, cytokines levels, and zonulin concentrations. Stool samples were used for 16S sequencing and real-time PCR. Serum was used for cytokine determination by flow cytometry, and zonulin quantification by ELISA. Pearson's chi-square, Mann-Whitney, and Spearman's correlation were used for statistical analyses. We detected differences in dietary habits, as well as in the gut microbiota in RRMS patients, with predominance of Akkermansia muciniphila and Bacteroides vulgatus and decreased Bifidobacterium. Interleukin-6 concentrations were decreased in treated patients, and we detected an increased intestinal permeability in RRMS patients when compared with controls. We conclude that diet plays an important role in the composition of the gut microbiota, and intestinal dysbiosis, detected in RRMS patients could be involved in increased intestinal permeability and affect the clinical response to DTMs. The future goal is to predict therapeutic responses based on individual microbiome analyses (personalized medicine) and propose dietary interventions and the use of probiotics or other microbiota modulators as adjuvant therapy to enhance the therapeutic efficacy of DMTs.
Subject(s)
Multiple Sclerosis, Relapsing-Remitting , Multiple Sclerosis , Bacteroides , Brazil , Dysbiosis , Humans , PermeabilityABSTRACT
Keratinases are proteolytic enzymes with a particular ability to cleave peptide bonds in keratin, and in other proteins. Due to their broad-spectrum of activity, keratinases are considered viable substitutes for chemical and thermal treatments of protein-rich industrial by-products. Among these protein residues, special attention has been given to keratinous materials (feathers, hair, horns, etc.), which disposal through harsh conditions methods, such as acid/alkaline hydrolysis or incineration, is not considered ecologically safe. Microbial keratinolytic enzymes allow for keratin degradation under mild conditions, resulting in keratin hydrolysates containing undamaged amino acids and peptides. In this review article, we offer perspectives on the relevance of these unique biocatalysts and their revolutionary ascent in industries that generate keratin-rich wastes. Additionally, we share insights for applications of keratinases and protein hydrolysates in agriculture, animal feed, cosmetics, phamaceuticals, detergent additives, leather processing, and others. Due to the scientific importance of keratinases and their potential use in green technologies, searching for bacterial and fungal species that efficiently produce these enzymes may contribute to the sustainability of industries.
Subject(s)
Keratins/chemistry , Peptide Hydrolases/metabolism , Biocatalysis , Industrial Waste/analysis , Peptide Hydrolases/genetics , Protein Engineering , ProteolysisABSTRACT
Derived compounds from lignin have been used as substrates for chemical and biological processes for obtainment bioproducts. The ferulic acid is a lignocellulosic biomass whose biotransformation in flavors compounds was described. The objective of this study was the bioconversion of ferulic acid to 4-vinylguaiacol by Klebsiella pneumoniae TD 4.7. The biotransformation of commercial ferulic acid into 4-vinylguaiacol in a semi synthetic liquid medium containing the ferulic acid at an initial concentration of 300 mg L-1 reached 32.4%. The ferulic acid obtained from alkaline hydrolysis of the sugar cane bagasse at 300 mg L-1 allowed the yield of 1.3 mmol L-1 of 4-vinylguaiacol, corresponding to 81.7% of the ferulic acid content. The data indicated that the bacterial strain decarboxylated the ferulic acid to 4-vinylguaiacol and the presence of an active cell associated ferulic acid decarboxylase. The enzyme showed maximum activity at pH 5.5 and 40 °C and was stable at pH range 4.5 to 9.0 and temperature up 20 to 45 °C. According to these biochemical properties and performance to bioconversion of ferulic acid to 4-vinylguaiacol, this enzyme could be viable for application in food industry.
Subject(s)
Coumaric Acids , Klebsiella pneumoniae , Biotransformation , Coumaric Acids/metabolism , Klebsiella pneumoniae/metabolism , LigninABSTRACT
Hashimoto thyroiditis (HT) is the most common autoimmune disease worldwide, characterized by chronic inflammation and circulating autoantibodies against thyroid peroxidase and thyroglobulin. Patients require hormone replacement with oral levothyroxine, and if untreated, they can develop serious adverse health effects and ultimately death. There is a lot of evidence that the intestinal dysbiosis, bacterial overgrowth, and increased intestinal permeability favor the HT development, and a thyroid-gut axis has been proposed, which seems to impact our entire metabolism. Here, we evaluated alterations in the gut microbiota in Brazilian patients with HT and correlated this data with dietary habits, clinical data, and systemic cytokines and zonulin concentrations. Stool samples from 40 patients with HT and 53 controls were analyzed using real-time PCR, the serum cytokine levels were evaluated by flow cytometry, zonulin concentrations by ELISA, and the dietary habits were recorded by a food frequency questionnaire. We observed a significant increase (p < 0.05) in the Bacteroides species and a decrease in Bifidobacterium in samples of patients with HT. In addition, Lactobacillus species were higher in patients without thyroid hormone replacement, compared with those who use oral levothyroxine. Regarding dietary habits, we demonstrated that there are significant differences in the consumption of vegetables, fruits, animal-derived proteins, dairy products, saturated fats, and carbohydrates between patients and control group, and an inverse correlation between animal-derived protein and Bacteroides genus was detected. The microbiota modulation by diet directly influences the inflammatory profile due to the generated microbiota metabolites and their direct or indirect action on immune cells in the gut mucosa. Although there are no differences in systemic cytokines in our patients with HT, we detected increased zonulin concentrations, suggesting a leaky gut in patients with HT. These findings could help understand the development and progression of HT, while further investigations to clarify the underlying mechanisms of the diet-microbiota-immune system axis are still needed.
Subject(s)
Dysbiosis/immunology , Gastrointestinal Microbiome/immunology , Hashimoto Disease/immunology , Intestines/immunology , Adult , Bacteria/classification , Bacteria/genetics , Cytokines/blood , Cytokines/immunology , Cytokines/metabolism , Dysbiosis/microbiology , Feces/microbiology , Feeding Behavior , Female , Haptoglobins/immunology , Haptoglobins/metabolism , Hashimoto Disease/blood , Hashimoto Disease/microbiology , Humans , Intestines/microbiology , Intestines/physiology , Male , Middle Aged , Permeability , Protein Precursors/blood , Protein Precursors/immunology , Protein Precursors/metabolism , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNAABSTRACT
Fungal enzymes are widely used in technological processes and have some interesting features to be applied in a variety of biosynthetic courses. Here, free and substrate-immobilised lipases from Fusarium verticillioides P24 were obtained by solid-state fermentation using wheat bran as substrate and fungal carrier. Based on their hydrolytic and transesterification activities, the lipases were characterised as pH-dependent in both reactions, with higher substrate conversion in an alkaline environment. Thermally, the lipases performed well from 30 to 45 °C, being more stable in mild conditions. Organic solvents significantly influenced the lipase selectivity using different vegetable oils as fatty acid source. Omega(ω)-3 production in n-hexane achieved 45% using canola oil, against ≈ 18% in cyclohexane. However, ω-6 production was preferably produced for both solvents using linseed oil with significant alterations in the yield (≈ 79% and 49% for n-hexane and cyclohexane, respectively). Moreover, the greatest enzyme selectivity for ω-6 led us to suppose a lipase preference for the Sn1 position of the triacylglycerol. Lastly, a transesterification reaction was performed, achieving 90% of ester conversion in 72 h. This study reports the characterisation and use of free and substrate-immobilised lipases from Fusarium verticillioides P24 as an economic and efficient method for the first time.
Subject(s)
Biocatalysis , Enzymes, Immobilized/chemistry , Fungal Proteins/chemistry , Fusarium/enzymology , Lipase/chemistry , EsterificationABSTRACT
A protease from the fungus Mucor subtilissimus URM 4133, capable of producing bioactive peptides from goat casein, was purified. SDS-PAGE and zymography showed a molecular mass of 30 kDa. The enzyme was active and stable in a wide pH range (6.0-10.5) and (5.0-10.5), respectively. Optimum temperature was at 45-50 °C and stability was above 80 % (40 °C/2 h). Activity was not influenced by ions or organic substances (Triton, Tween, SDS and DMSO), but was completely inhibited by PMSF, suggesting that it belongs to the serine protease family. The Km and Vmax were 2.35 mg azocasein.mL-1 and 333.33 U.mg protein-1, respectively. Thermodynamic parameters of irreversible denaturation (40-60 °C) were enthalpy 123.63 - 123.46 kJ.mol-1, entropy 120.24-122.28 kJ.mol-1 and Gibbs free energy 85.97 - 82.45 kJ.mol-1. Any peptide sequences compatible with this protease were found after analysis by MALDI-TOF, which suggests that it is a new serine protease.
ABSTRACT
The main organochlorinated compounds used on agricultural crops are often recalcitrant, affecting nontarget organisms and contaminating rivers or groundwater. Diuron (N-(3,4-dichlorophenyl)-N',N'-dimethylurea) is a chlorinated herbicide widely used in sugarcane plantations. Here, we evaluated the ability of 13 basidiomycete strains of growing in a contaminated culture medium and degrading the xenobiotic. Dissipation rates in culture medium with initial 25 mg/L of diuron ranged from 7.3 to 96.8%, being Pluteus cubensis SXS 320 the most efficient strain, leaving no detectable residues after diuron metabolism. Pycnoporus sanguineus MCA 16 removed 56% of diuron after 40 days of cultivation, producing three metabolites more polar than parental herbicide, two of them identified as being DCPU and DCPMU. Despite of the strong inductive effect of diuron upon laccase synthesis and secretion, the application of crude enzymatic extracts of P. sanguineus did not catalyzed the breakdown of the herbicide in vitro, indicating that diuron biodegradation was not related to this oxidative enzyme.
Subject(s)
Basidiomycota/metabolism , Biodegradation, Environmental , Diuron/pharmacokinetics , Herbicides/pharmacokinetics , Water Pollutants, Chemical/pharmacokinetics , Agaricales/metabolism , Basidiomycota/drug effects , Basidiomycota/growth & development , Cytochrome P-450 Enzyme System/metabolism , Diuron/toxicity , Herbicides/toxicity , Hydrocarbons, Chlorinated/pharmacokinetics , Hydrocarbons, Chlorinated/toxicity , Laccase/metabolism , Models, Biological , Nitrogen/metabolism , Polyporaceae/metabolism , Rainforest , Water Pollutants, Chemical/toxicityABSTRACT
BACKGROUND: Atrazine is one of the most widespread chlorinated herbicides, leaving large bulks in soils and groundwater. The biodegradation of atrazine by bacteria is well described, but many aspects of the fungal metabolism of this compound remain unclear. Thus, we investigated the toxicity and degradation of atrazine by 13 rainforest basidiomycete strains. RESULTS: In liquid medium, Pluteus cubensis SXS320, Gloelophyllum striatum MCA7, and Agaricales MCA17 removed 30, 37, and 38%, respectively, of initial 25 mg L- 1 of the herbicide within 20 days. Deficiency of nitrogen drove atrazine degradation by Pluteus cubensis SXS320; this strain removed 30% of atrazine within 20 days in a culture medium with 2.5 mM of N, raising three metabolites; in a medium with 25 mM of N, only 21% of initial atrazine were removed after 40 days, and two metabolites appeared in culture extracts. This is the first report of such different outcomes linked to nitrogen availability during the biodegradation of atrazine by basidiomycetes. The herbicide also induced synthesis and secretion of extracellular laccases by Datronia caperata MCA5, Pycnoporus sanguineus MCA16, and Polyporus tenuiculus MCA11. Laccase levels produced by of P. tenuiculus MCA11 were 13.3-fold superior in the contaminated medium than in control; the possible role of this enzyme on atrazine biodegradation was evaluated, considering the strong induction and the removal of 13.9% of the herbicide in vivo. Although 88% of initial laccase activity remained after 6 h, no evidence of in vitro degradation was observed, even though ABTS was present as mediator. CONCLUSIONS: This study revealed a high potential for atrazine biodegradation among tropical basidiomycete strains. Further investigations, focusing on less explored ligninolytic enzymes and cell-bound mechanisms, could enlighten key aspects of the atrazine fungal metabolism and the role of the nitrogen in the process.
Subject(s)
Agaricales/drug effects , Agaricales/metabolism , Atrazine/metabolism , Laccase/metabolism , Agaricales/growth & development , Agaricales/isolation & purification , Atrazine/pharmacology , Biodegradation, Environmental , Culture Media , Environmental Pollutants/metabolism , Extracellular Matrix/enzymology , Fungal Proteins/metabolism , Nitrogen/metabolism , Polyporaceae/drug effects , Polyporaceae/metabolism , Rainforest , Species SpecificityABSTRACT
OBJECTIVES: Keratinases are proteolytic enzymes that emerge as an alternative for dealing with the disposal of chicken feathers. In this study, we aimed to investigate the keratin-degrading enzymes secreted by the fungus Coriolopsis byrsina and their partial biochemical characterization to adapt their use for keratin decomposition, detergent additive applications, and collagen degradation. RESULTS: We observed the secretion of different proteolytic enzymes that possessed caseinolytic activity that peaked at pH 7.0-9.0 and 60-70 °C and at pH 10.5 and 55-60 °C, and keratinolytic activity that reached a maximum at pH 7.0-7.5 and 40-55 ºC and at pH 9.0 and 55 °C. Keratinolytic activity was maintained at approximately 63% of residual activity for 1 h at 50 °C. The caseinolytic activity at pH 10.5 remains stable until 1 h at 50 °C, and this is in contrast to the activity at pH 8.5, where the residual activity was 50%. Caseinolytic activity was inhibited only by PMSF, while keratinolytic activity was inhibited by PMSF and EDTA. When investigating the application of C. byrsina peptidases as an additive to commercial detergent, we observed an egg stain removal performance that was similar to that demonstrated by the commercial detergent. CONCLUSIONS: Based on their activity and stability at alkaline pH, these enzymes appear to be attractive candidates for use in the detergent industry. Additionally, the collagenolytic activity of these enzymes potentially allows for their use in a wide array of industrial sectors that require collagenolytic enzymes, such as for the production of collagen hydrolysates from residues derived from the meat industry.
Subject(s)
Feathers/chemistry , Peptide Hydrolases/chemistry , Peptide Hydrolases/metabolism , Polyporaceae/growth & development , Animals , Batch Cell Culture Techniques , Caseins/chemistry , Enzyme Stability , Fermentation , Fungal Proteins/metabolism , Hot Temperature , Hydrogen-Ion Concentration , Polyporaceae/enzymology , TextilesABSTRACT
Lipase stability in organic solvent is crucial for its application in many biotechnological processes as biocatalyst. One way to improve lipase's activity and stability in unusual reaction medium is its immobilization on inert supports. Here, lipases from different sources and immobilized through weak chemical interactions on hydrophobic and ionic supports had their transesterification ability dramatically dependent on the support and also on the solvent that had been used. The ethanolysis of sardine oil was carried out at the presence of cyclohexane and tert-amyl alcohol, in which Duolite A568-Thermomyces lanuginosa lipase derivative achieved 49% of ethyl esters production after 24 h in cyclohexane. The selectivity of immobilized lipases was also studied and, after 3 h of synthesis, the reaction with Duolite A568-Thermomyces lanuginosa derivative in cyclohexane produced 24% ethyl ester of eicosapentaenoic acid and 1.2% ethyl ester of docosahexaenoic acid, displaying a selectivity index of 20 times the ethyl ester of eicosapentaenoic acid. Different derivatives of Candida antarctica lipases fraction B (CALB) and phospholipase Lecitase® Ultra (Lecitase) were also investigated. Along these lines, a combination between these factors may be applied to improve the activity and selectivity of immobilized lipases, decreasing the total cost of the process.
Subject(s)
Alcohols/chemistry , Esters/chemistry , Fungal Proteins/chemistry , Hexanes/chemistry , Lipase/chemistry , Organic Chemicals/chemistry , Solvents/chemistry , Adsorption , Animals , Biocatalysis , Candida/metabolism , Catalysis , Colorimetry/methods , Cyclohexanes/chemistry , Enzymes, Immobilized/chemistry , Esterification , Ethane/chemistry , Ethanol/chemistry , Fishes , Hydrophobic and Hydrophilic Interactions , Ions , PentanolsABSTRACT
Currently, poultry farming is one of the sectors that have a significant impact on the global economy. In recent years, there has been an increase in the production of broilers, inflicting this segment of the industry to generate tons of keratin due to huge disposal of chicken feathers. This points to the need to degrade these chicken feathers, as they have emerged as a major threat to the environment. Thus, in this study we aimed to identify keratinases that are produced by the bacterium Citrobacter diversus and further investigate the biochemical characteristics of these keratin-degrading enzymes. In a submerged medium, the bacterium was capable of degrading chicken feathers almost completely after 36 h of fermentation. We found a maximum caseinolytic activity at pH 9-10.5 and 50-55 °C, and keratinolytic activity at pH 8.5-9.5 and 50 °C. Thus, given its stability at higher temperatures, upon incubation of this enzyme extract for 1 h at 50 °C, it showed approximately 50% of the keratinolytic and 100% of the caseinolytic activity. Further, under pH stability for 48 h at 4 °C, the enzyme extract maintained greater residual activity in the pH range 6-8. Caseinolytic activity was inhibited by EDTA and PMSF, whereas the keratinolytic activity was inhibited only by EDTA. Additionally, due to its alkaline activity and detergent compatibility, this enzyme extract could improve washing performance when added to a commercial detergent formulation. Using application tests, we could demonstrate a potential use of this bacterial enzyme extract as an additive in detergents to remove egg stains from cloth.
Subject(s)
Bacterial Proteins/metabolism , Citrobacter koseri/enzymology , Detergents/metabolism , Peptide Hydrolases/metabolism , Animals , Bacterial Proteins/isolation & purification , Biodegradation, Environmental , Caseins/metabolism , Chickens , Citrobacter koseri/metabolism , Culture Media/metabolism , Detergents/chemistry , Feathers/metabolism , Fermentation , Hydrogen-Ion Concentration , Keratins/metabolism , Peptide Hydrolases/isolation & purification , TemperatureABSTRACT
The effect of pectinolytic enzyme preparation (PEP) produced by the fungus Thermomucor indicae-seudaticae-N31 (PEP-N31) on total phenolic content, concentrations of methanol and color of grape juice was studied. Positive results were found when PEP-N31 was used to extract phenolic compounds after the grapes had been blanched for 3 min and macerated for 1 h. The resulting juice had better yield, color characteristics and higher phenolic content (1637.21 mg.L-1, as gallic acid equivalent, or GAE) than the conventionally prepared juice (1422.59 mg GAE.L-1), and it was very similar to the juice obtained through the treatment with a commercial enzyme (1682.10 mg GAE.L-1). The concentration of methanol in the juice produced with the PEP-N31 was less than 200 mg.L-1. These results encourage the use of PEP produced by Thermomucor indicae-seudaticae-N31 by the grape-processing industry.