Genetically modified ZIKA virus as a microRNA-sensitive oncolytic virus against central nervous system tumors.

Here we introduce a first-in-class microRNA-sensitive oncolytic Zika virus (ZIKV) for virotherapy application against central nervous system (CNS) tumors. The described methodology produced two synthetic modified ZIKV strains that are safe in normal cells, including neural stem cells, while preserving brain tropism and oncolytic effects in tumor cells. The microRNA-sensitive ZIKV introduces genetic modifications in two different virus sites: first, in the established 3'UTR region, and secondly, in the ZIKV protein coding sequence, demonstrating for the first time that the miRNA inhibition systems can be functional outside the UTR RNA sites. The total tumor remission in mice bearing human CNS tumors, including metastatic tumor growth, after intraventricular and systemic modified ZIKV administration, confirms the promise of this virotherapy as a novel agent against brain tumors-highly deadly diseases in urgent need of effective advanced therapies.

Highly Enantioselective Lewis Acid Catalyzed Conjugate Addition of Imidazo[1,2-a]pyridines to α,ß-Unsaturated 2-Acylimidazoles under Mild Conditions.

A highly enantioselective protocol for the conjugate addition of 2-arylimidazo[1,2-a]pyridines and other imidazo derivatives to α,ß-unsaturated 2-acylimidazoles is described. The method uses a previously reported chiral-at-metal rhodium catalyst and provides the corresponding adducts in yields of 25-98% with enantioselectivities up to er > 99:1. Additionally, the transformation proceeds under mild conditions using ethanol as the solvent at room temperature.

KIO3-catalyzed selective oxidation of thiols to disulfides in water under ambient conditions.

Herein, we report a KIO3-catalyzed oxidative coupling of thiols to their corresponding disulfides in water, in a short time and at ambient temperature. The reaction has a broad scope and exhibits good functional group tolerance, resulting in the desired products in excellent yields. This approach allows the reuse of the reaction system in multiple cycles and scale-up. Furthermore, the current protocol demonstrates compatibility for in situ generation of disulfides and post application in C(sp2)-H bond sulfenylation.

Improving the extraction yield of essential oil from Pimenta dioica (L.) Merr. using Aspergillus niger ATCC 1004 enzyme blend.

The objective of this work was to optimize the application of an enzymatic blend produced by Aspergillus niger ATCC 1004 on the Pimenta dioica fruits for essential oil extraction. The enzyme blend was obtained from the fermentation of cocoa bean shells, an agro-industrial residue. The effects of the enzymatic pre-treatment on the extraction yield, the chemical composition of the oil through gas chromatography, and the fruit structure through scanning electron microscopy (SEM) were assessed. A Doehlert design was used to optimize the process conditions, resulting in an extraction with 117 mL of enzyme during 77 min, which increased the extraction yield by 387.5%. The chemical composition was not altered, which proves that the enzyme blend preserves the quality of the essential oil extracted. The content of eugenol (70%), the major compound in the P. dioica essential oil, had a great increase in its concentration (560%). The enzyme activity analyses showed the presence of endoglucanase (0.4 U/mL), exoglucanase (0.25 U/mL), ß-glucosidase (0.19 U/mL), and invertase (135.08 U/mL). The microscopy analyses revealed changes in the morphology of fruit surface due to the enzymatic action. These results demonstrate the great potential of using enzyme blends produced by filamentous fungi from agro-industrial residues for the essential oils extraction of interest for the pharmaceutical and food industries.

Bacterial Extracellular Vesicles: Potential Therapeutic Applications, Challenges, and Future Prospects.

Almost all cell types naturally secret extracellular vesicles (EVs) in the extracellular space with variable metabolic cargo facilitating intracellular communication, posing immune-modulation capacity. Thus, "bacterial extracellular vesicles" (BEVs), with their great immunoregulatory, immune response stimulation and disease condition-altering potential, have gained importance in the medical and therapeutic industry. Various subtypes of BEVs were observed and reported in the literature, such as exosomes (30-150 nm), microvesicles (100-1000 nm), apoptotic bodies (1000-5000 nm), and oncosomes (1000-10,000 nm). As biological systems are complex entities, inserting BEVs requires extra high purity. Various techniques for BEV isolation have been employed alone or with other strategies, such as ultracentrifugation, precipitation, size-exclusion chromatography, affinity-based separation, ultrafiltration, and field-flow fractionation. But to date, no BEV isolation method is considered perfect as the lack of standard protocols limits their scale-up. Medical research has focused on BEVs to explore their diverse therapeutic potential. This review particularly focused on the recent advancements in the potential medical application of BEVs, current challenges, and prospects associated with their scale-up.

Optimization of enzymatic saccharification of industrial wastes using a thermostable and halotolerant endoglucanase through Box-Behnken experimental design.

This study describes the production, characterization and application of an endoglucanase from Penicillium roqueforti using lignocellulosic agro-industrial wastes as the substrate during solid-state fermentation. The endoglucanase was generated after culturing with different agro-industrial wastes for 96 h without any pretreatment. The highest activity was obtained at 50 °C and pH 4.0. Additionally, the enzyme showed stability in the temperature and pH ranges of 40-80 °C and 4.0-5.0, respectively. The addition of Ca2+, Zn2+, Mg2+, and Cu2+ increased enzymatic activity. Halotolerance as a characteristic of the enzyme was confirmed when its activity increased by 35% on addition of 2 M NaCl. The endoglucanase saccharified sugarcane bagasse, coconut shell, wheat bran, cocoa fruit shell, and cocoa seed husk. The Box-Behnken design was employed to optimize fermentable sugar production by evaluating the following parameters: time, substrate, and enzyme concentration. Under ideal conditions, 253.19 mg/g of fermentable sugars were obtained following the saccharification of wheat bran, which is 41.5 times higher than that obtained without optimizing. This study presents a thermostable, halotolerant endoglucanase that is resistant to metal ions and organic solvents with the potential to be applied in producing fermentable sugars for manufacturing biofuels from agro-industrial wastes.

Slc11a1 gene polymorphism influences dextran sulfate sodium (DSS)-induced colitis in a murine model of acute inflammation.

Ulcerative Colitis (UC) is an inflammatory disease characterized by colonic mucosal lesions associated with an increased risk of carcinogenesis. UC pathogenesis involves environmental and genetic factors. Genetic studies have indicated the association of gene variants coding for the divalent metal ion transporter SLC11A1 protein (formerly NRAMP1) with UC susceptibility in several animal species. Two mouse lines were genetically selected for high (AIRmax) or low (AIRmin) acute inflammatory responses (AIR). AIRmax is susceptible, and AIRmin is resistant to DSS-induced colitis and colon carcinogenesis. Furthermore, AIRmin mice present polymorphism of the Slc11a1 gene. Here we investigated the possible modulating effect of the Slc11a1 R and S variants in DSS-induced colitis by using AIRmin mice homozygous for Slc11a1 R (AIRminRR) or S (AIRminSS) alleles. We evaluated UC by the disease activity index (DAI), considering weight loss, diarrhea, blood in the anus or feces, cytokines, histopathology, and cell populations in the distal colon epithelium. AIRminSS mice have become susceptible to DSS effects, with higher DAI, IL6, G-CSF, and MCP-1 production and morphological and colon histopathological alterations than AIRminRR mice. The results point to a role of the Slc11a1 S allele in DSS colitis induction in the genetic background of AIRmin mice.

The improvement of guava (Psidium guajava) juice quality using crude multi-enzymatic extracts obtained from yeasts.

Guava juice is cloudy and viscous, which hinders filtration, decreases yield, and causes the loss of quality after its processing and during storage. This study aimed to evaluate enzymatic treatment effects using crude multi-enzymatic extracts (CME) obtained from Rhodotorula mucilaginosa, Rhodotorula orizycola, and Pseudozyma sp. produced by submerse fermentation in the extraction of juice guava. Mixtures of 100 ml of guava pulp and multi-enzymatic extracts proposed by Doehlert planning were incubated under constant agitation at 150 rpm and 50°C, and a Doehlert design was applied as a multivariate optimization strategy. The optimal conditions using the multi-enzymatic extract were: 0.4% (v/v) of CME for 131 min for the multi-enzymatic treatment using Pseudozyma sp.; 3.0% (v/v) of CME for 154 min using the R. mucilaginosa CME; and 5.0% (v/v) of CME for 90 min using R. oryzicola. The maximum viscosity reduction values for the juices treated with the CME of yeasts were 10.33%, 86.38%, and 13.33% for the juices treated with the CME of Pseudozyma sp., R. mucilaginosa, and R. orizycola, respectively. The physical-chemical properties were improved after treatment with CMEs, yielding a reduction of clarity, increase of total soluble solids and reducing sugars, and decreasing the acidity (pH) for all treatments with enzymatic extracts of all strains. The yeasts studied showed a potential for CME production to be applied to juice, improving the quality of the juice, and R. mucilaginosa was the most prominent yeast due to most significant reduction of viscosity in guava juice.

Dilute acid pretreatment for enhancing the enzymatic saccharification of agroresidues using a Botrytis ricini endoglucanase.

The enormous amount of agroindustrial residues generated in Brazil can be used as biomass to produce fermentable sugars. This study compared the pretreatments with different proportions of dilute acid. The method involved pretreatment with 0.5%, 1%, and 1.5% (v/v) sulfuric acid, followed by hydrolysis using the halotolerant and thermostable endoglucanase from Botrytis ricini URM 5627. The physicochemical characterization of plant biomass was performed using XRD, FTIR, and SEM. The pretreatment significantly increased the production of fermentable sugars following enzymatic saccharification from wheat bran, sugarcane bagasse, and rice husk: 153.67%, 91.98%, and 253.21% increment in sugar production; 36.39 mg⋅g-1 ± 1.23, 39.55 mg⋅g-1 ± 1.70, and 42.53 mg⋅g-1 ± 7.61 mg⋅L-1 of glucose; and 3.26 ± 0.35 mg⋅g-1 , 3.61mg⋅g-1 ± 0.74 and 3.59 mg⋅g-1 ± 0.80 of fructose were produced, respectively. In conclusion, biomass should preferably be pretreated before the enzymatic saccharification using B. ricini URM 5627 endoglucanase.

Advanced nanomaterials and metal-organic frameworks for catalytic bio-diesel production from microalgal lipids - A review.

Increasing energy demands require exploring renewable, eco-friendly (green), and cost-effective energy resources. Among various sources of biodiesel, microalgal lipids are an excellent resource, owing to their high abundance in microalgal biomass. Transesterification catalyzed by advanced materials, especially nanomaterials and metal-organic frameworks (MOFs), is a revolutionary process for overcoming the energy crisis. This review elaborates on the conversion of microalgal lipids (including genetically modified algae) into biodiesel while primarily focusing on the transesterification of lipids into biodiesel by employing catalysts based on above mentioned advanced materials. Furthermore, current challenges faced by this process for industrial scale upgradation are presented with future perspectives and concluding remarks. These materials offer higher conversion (>90%) of microalgae into biodiesel. Nanocatalytic processes, lack the need for higher pressure and temperature, which simplifies the overall process for industrial-scale application. Green biodiesel production from microalgae offers better fuel than fossil fuels in terms of performance, quality, and less environmental harm. The chemical and thermal stability of advanced materials (particularly MOFs) is the main benefit of the blue recycling of catalysts. Advanced materials-based catalysts are reported to reduce the risk of biodiesel contamination. While purity of glycerin as side product makes it useful skin-related product. However, these aspects should still be controlled in future studies. Further studies should relate to additional aspects of green production, including waste management strategies and quality control of obtained products. Finally, catalysts stability and recycling aspects should be explored.

Structural alteration of cocoa bean shell fibers through biological treatment using Penicillium roqueforti.

Lignocellulosic residues, such as cocoa bean shell (FI), are generated in large quantities during agro-industrial activities. Proper management of residual biomass through solid state fermentation (SSF) can be effective in obtaining value-added products. The hypothesis of the present work is that the bioprocess promoted by P. roqueforti can lead to structural changes in the fibers of the fermented cocoa bean shell (FF) that confer characteristics of industrial interest. To unveil such changes, the techniques of FTIR, SEM, XRD, TGA/TG were used. After SSF, an increase of 36.6% in the crystallinity index was observed, reflecting the reduction of amorphous components such as lignin in the FI residue. Furthermore, an increase in porosity was observed through the reduction of the 2θ angle, which gives the FF a potential candidate for applications of porous products. The FTIR results confirm the reduction in hemicellulose content after SSF. The thermal and thermogravimetric tests showed an increase in the hydrophilicity and thermal stability of FF (15% decomposition) in relation to the by-product FI (40% decomposition). These data provided important information regarding changes in the crystallinity of the residue, existing functional groups and changes in degradation temperatures.

This work presents a new approach for solid state fermentation based on the study of structural changes caused by Penicillium roquefort, which is important to understand the changes in the lignocellulosic matrix after the fungus growth. The results provided important information regarding changes in the crystallinity of the residue, existing functional groups and changes in degradation temperatures. Consequently, they can help in proposals for the total use of the residual solid after fermentation, as well as contribute to reducing the lack of this information in the literature.

Development of the dairy products incorporated with co-product bioactive compounds-rich as an alternative ingredient in the food industry.

The objective was to optimize the phenolic compounds extraction from cocoa shells using the simplex-centroid design with a mixture of solvents (water, methanol, and acetone) as its components, to prove the presence of these compounds and antioxidant activity. Also, the development of dairy products, such as milk beverages and dairy desserts, with bioactive compounds, through the replacement of cocoa powder by cocoa shell was studied and evaluated sensorially. The extraction optimization indicated that a solvent with 56.44% water, 23.77% methanol, and 19.80% acetone are ideal for maximizing the phenolic compounds. In addition, the cocoa shell showed a high antioxidant activity by the methods ß-carotene/linoleic acid, FRAP, and phosphomolybdenum complex. The Check-All-That-Apply, Cochran's Q test, contingency analysis, and hierarchical cluster analysis allowed description characteristics of the dairy products and showed sensory differences between formulations with 100% cocoa shell and others. Both dairy products had good sensory acceptance in all attributes evaluated (appearance, flavor, texture, and overall impression), and their scores did not differ statistically by Tukey's test (p > 0.05). Thus, the cocoa shell is shown as an alternative substitute ingredient to be used in the dairy industry.

Mapping of novel loci involved in lung and colon tumor susceptibility by the use of genetically selected mouse strains.

Two non-inbred mouse lines, phenotypically selected for maximal (AIRmin) and minimal (AIRmax) acute inflammatory response, show differential susceptibility/resistance to the development of several chemically-induced tumor types. An intercross pedigree of these mice was generated and treated with the chemical carcinogen dimethylhydrazine, which induces lung and intestinal tumors. Genome wide high-density genotyping with the Restriction Site-Associated DNA genotyping (2B-RAD) technique was used to map genetic loci modulating individual genetic susceptibility to both lung and intestinal cancer. Our results evidence new common quantitative trait loci (QTL) for those phenotypes and provide an improved understanding of the relationship between genomic variation and individual genetic predisposition to tumorigenesis in different organs.

Gums-based engineered bio-nanostructures for greening the 21st-century biotechnological settings.

Naturally occurring plant-based gums and their engineered bio-nanostructures have gained an immense essence of excellence in several industrial, biotechnological, and biomedical sectors of the modern world. Gums derived from bio-renewable resources that follow green chemistry principles are considered green macromolecules with unique structural and functional attributes. For instance, gum mostly obtained as exudates are bio-renewable, bio-degradable, bio-compatible, sustainable, overall cost-effective, and nontoxic. Gum exudates also offer tunable attributes that play a crucial role in engineering bio-nanostructures of interest for several bio- and non-bio applications, e.g., food-related items, therapeutic molecules, sustained and controlled delivery cues, bio-sensing constructs, and so on. With particular reference to plant gum exudates, this review focuses on applied perspectives of various gums, i.e., gum Arabic, gum albizzia, gum karaya, gum tragacanth, and gum kondagogu. After a brief introduction with problem statement and opportunities, structural and physicochemical attributes of plant-based natural gums are presented. Following that, considerable stress is given to green synthesis and stabilization of gum-based bio-nanostructures. The final part of the review focuses on the bio- and non-bio related applications of various types of gums polysaccharides-oriented bio-nanostructures.

Production of a fermented solid containing lipases from Penicillium roqueforti ATCC 10110 and its direct employment in organic medium in ethyl oleate synthesis.

The production and direct employment in organic medium in the ethyl-oleate synthesis of a fermented solid (FS) containing lipases by Penicillium roqueforti ATCC 10110 (PR10110) was investigated. For the production of this FS, the solid-state fermentation of different agroindustrial waste was used, such as: cocoa shell, sugarcane bagasse, sugarcane bagasse with cocoa shell, and cocoa shell with soybean oil and nutrient solution. The response surface methodology was used to study the effect of independent variables of initial moisture content and inductor concentration, as carbon source and inducer on lipase production. The characterization of the fermented solid in organic medium was also carried out. The highest lipase activity (53 ± 5 U g-1 ) was 16% higher than that obtained with the nonoptimized conditions. The characterization studies observed high stability of the FS in organic solvents for 5 h at 30°C, as well as at different temperatures, and the residual activity was measured against triolein. The FS was also able to catalyze ethyl-oleate synthesis maintaining high relative conversion over five reaction cycles of 96 h at 40°C in n-heptane. These results are promising and highlight the use of the FS containing PR10110 lipases for the first time in biocatalytic processes.

Thermostable trypsin-like protease by Penicillium roqueforti secreted in cocoa shell fermentation: Production optimization, characterization, and application in milk clotting.

The increased demand for cheese and the limited availability of calf rennet justifies the search for milk-clotting enzymes from alternative sources. Trypsin-like protease by Penicillium roqueforti was produced by solid-state fermentation using cocoa shell waste as substrate. The production of a crude enzyme extract that is rich in this enzyme was optimized using a Doehlert-type multivariate experimental design. The biochemical characterization showed that the enzyme has excellent activity and stability at alkaline pH (10-12) and an optimum temperature of 80°C, being stable at temperatures above 60°C. Enzymatic activity was maximized in the presence of Na+ (192%), Co2+ (187%), methanol (153%), ethanol (141%), and hexane (128%). Considering the biochemical characteristics obtained and the milk coagulation activity, trypsin-like protease can be applied in the food industry, such as in milk clotting and in the fabrication of cheeses.

Application crude multienzyme extract from Aspergillus niger as a pretreatment for the extraction of essential oil from Croton argyrophyllus leaves.

Leaves of Croton argyrophyllus contain essential oil with promising active components for the development of drugs and botanical insecticides. In this study, we evaluated the enzymatic pretreatment process to increase the extraction of essential oil from fresh and dried leaves of C. argyrophyllus. Pretreatment was carried out using a crude multienzymatic extract obtained via solid-state fermentation of forage palm by Aspergillus niger, and the extraction was performed by hydrodistillation. A Doehlert matrix was used to optimize the enzymatic pretreatment variables temperature and enzymatic extract. The effect of pretreatment time was also investigated. At optimum experimental conditions, 41.34°C, 140 min, and 130.73 mL of enzyme in 369.27 mL of water, the essential oil yield from fresh leaves subjected to enzymatic pretreatment increased by 9.35% and that from dry leaves by 6.77%. Based on chromatographic analysis (GC-MS), no compound was degraded in the extraction process. Micromorphological analysis confirmed the rupture of the glandular trichomes, favoring essential oil release. Therefore, enzymatic pretreatment associated with hydrodistillation increased the essential oil yield and is a promising application to obtain essential oil for therapeutic purposes without altering its composition.

High-throughput screening for distinguishing nitrilases from nitrile hydratases in Aspergillus and application of a Box-Behnken design for the optimization of nitrilase.

Nitrilases and nitrile hydratases/amidases hydrolyze nitriles into carboxylic acids and/or amides, which are used in industrial chemical processes. In the present study, 26 microorganisms, including yeasts and filamentous fungi, in a minimum solid mineral medium supplemented with glucose and phenylacetonitrile were screened to evaluate their biocatalytic potential. Of these microorganisms, five fungi of the genus Aspergillus were selected and subjected to colorimetry studies to evaluate the production and distinction of nitrilase and nitrile hydratase/amidase enzymes. Aspergillus parasiticus Speare 7967 and A. niger Tiegh. 8285 produced nitrilases and nitrile hydratase, respectively. Nitrilase optimization was performed using a Box-Behnken design (BBD) and fungus A. parasiticus Speare 7967 with phenylacetonitrile volume (µl), pH, and carbohydrate source (starch:glucose; g/g) as independent variables and nitrilase activity (U ml-1 ) as dependent variable. Maximum activity (2.97 × 10-3  U ml-1 ) was obtained at pH 5.5, 80 µl of phenylacetonitrile, and 15 g of glucose. A. parasiticus Speare 7967 showed promise in the biotransformation of nitriles to carboxylic acids.

Candida rugosa lipase immobilized on hydrophobic support Accurel MP 1000 in the synthesis of emollient esters.

OBJECTIVES: To immobilize Candida rugosa lipase in Accurel MP 1000 (CRL-AMP) by physical adsorption in organic medium and apply in the synthesis of wax esters dodecanoyl octadecanoate 1 and hexadecanoyl octadecanoate 2 in a heptane medium, as well as evaluating the stability and recyclability of CRL-AMP in six reaction cycles. RESULTS: The specific activity (Asp) for CRL-AMP was 200 ± 20 U mg-1. Its catalytic activity was 1300 ± 100 U g-1. CRL-AMP was used in the synthesis of esters in heptane medium with a 1:1 acid:alcohol molar ratio at 45 °C and 200 rpm. In synthesis 1, conversion was 62.5 ± 3.9% in 30 min at 10% m v-1 and 56.9 ± 2.8% in 54 min at 5% m v-1; while in synthesis 2, conversion was 79.0 ± 3.9% in 24 min at 10% m v-1, and 46.0 ± 2.4% in 54 min at 5% m v-1. Reuse tests after six consecutive cycles of reaction showed that the biocatalyst retained approximately 50% of its original activity for both reaction systems. CONCLUSIONS: CRL-AMP showed a high potential in the production of wax esters, since it started from low enzymatic load and high specific activities and conversions were obtained, in addition to allowing an increase in stability and recyclability of the prepared biocatalyst.

Optimization of lipase production by Penicillium roqueforti ATCC 10110 through solid-state fermentation using agro-industrial residue based on a univariate analysis.

Lipases (triacylglycerol hydrolases, EC 3.1.1.3) are a class of enzymes with high industrial importance. An option for the production of this enzyme is through fungal growth via solid-state fermentation (SSF). Thus, this research presents a study of lipase production by Penicillium roqueforti ATCC 10110 through SSF using cocoa bran residues (Theobroma cacao) as a substrate. To achieve maximum lipase production, fermentation time (0 to 120 h) and palm oil (PO) percentage (0 to 50%) were optimized through analysis of one factor at a time (OFAT), with lipase activity as the response. The amount of cocoa was fixed (5 g), the incubation temperature was maintained at 27 °C, and the moisture content was established at 70%. For a 72 h incubation, the highest enzyme activity achieved using SSF without adding PO was 14.67 ± 1.47 U g-1, whereas with PO (30%), it was 33.33 ± 3.33 U g-1, thus demonstrating a 44% increase in enzyme activity. Through the OFAT methodology, it was possible to confirm that supplementation with palm residue was efficient and maximized the lipase of P. roqueforti ATCC 10110.