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El uso de enzimas como α-amilasas, xilanasas y glucosidasas es una buena alternativa para obtener una mayor eficiencia en los productos de la industria panificadora, sin dejar de mencionar la lacasa que ha mostrado un potencial prometedor. Así, el objetivo de este estudio fue obtener concentrados enzimáticos de xilanasas, amilasas, celulasas y lacasas a partir de cepas nativas de Pycnoporus sanguineus. Para lo cual, se evaluó el efecto del medio de cultivo sobre la producción de lacasa juntamente con el escrutinio en medio sólido para la detección de xilanasas, amilasas y celulasas. A los concentrados enzimáticos obtenidos se les determinaron las unidades de lacasa y carbohidrolasas. Los títulos más altos de lacasa se presentaron en el medio Tx2. Con respecto a las carbohidrolasas, se detectaron en diferentes niveles de actividad y combinaciones. El concentrado con altos niveles de amilasa y lacasa fue CH116, mientras que LE90 tuvo la mayor actividad de xilanasa y LE133 de celulasa. La cepa CH116 fue productora de amilasa, celulasa, xilanasa y lacasa, siendo la de mayor potencial en la industria panificadora, mientras que LE90 puede ser utilizada en la industria papelera por su combinación de xilanasas y lacasa.

Sweet sorghum Sorghum bicolor (L.) Moench) is a promising crop for bioethanol production characterized by its high biomass yield, sugar-rich stalks, facile juice extraction, and ability to adapt to different production systems. The objective of this study was to assess the effects of cultivation practices (soil treatments) on the juice, sugar, and ethanol yields of sweet sorghum variety Roger. Three methods of tillage (minimum, traditional, and traditional tillage with rupture of the plow layer) and three types of fertilizers (inorganic, organic, and without fertilizer) were assayed. The best results of stem yield (44.3 tons/ha), juice production (25.6 m3/ha), concentration of soluble solids in the juice (16.9 °Brix), and juice ethanol yield (2.1 m3/ha) were obtained under a mixture of traditional plowing with plow layer breaking, applying an organic fertilizer. In addition, the extracted juice allowed yeast (industrial Saccharomyces cerevisiae PE-2) growth and conversion to ethanol without the need for nutrients supplementation. Therefore, sweet sorghum continues to be an interesting bioenergetic crop for production of first-generation biofuels.

Tuberculosis drug resistance (DR) is a global problem that is not fully elucidated. Previously, overexpression of esxG and esxH genes was reported in a multidrug-resistant (MDR) Mycobacterium tuber...

The use of lignocellulosic wastes reduces dependence on fossil fuel resources, contributes to sustainable waste management, and reinforces the circular economy model of continual use of resources. Typically, the second generation of bioethanol production involves several steps to transform lignocellulosic material into bioethanol. The more complicated step of the overall process is to define a tailor-made to each lignocellulosic material available with a wide variety of complex structures of the lignin, hemicellulose, and cellulose. However, the thermochemicals are the most frequently reported pretreatments, and they have the bottleneck of producing an additional waste stream with a high charge of pollution owing to chemical products implicated. This problem harms on the zero waste policy that we must include in our technological process to be considered sustainable and ecological processes. Consolidated bioprocessing (CBP) is a viable alternative to produce bioethanol from lignocellulosic materials, using a single microorganism, in one-step, and no chemical products are implied. Three agro-industrial wastes with lignocellulosic characteristics were evaluated as a substrate for bioethanol production with a Mexican native white-rot fungus Trametes hirsuta CS5 in a one-step process. Qualitative and quantitative analyses of lignocellulolytic enzymes produced by native fungus were carried out. Instead, Trametes hirsuta is a lignin degrader white-rot fungus; it was capable to produce until 500 U/L of cellulase titers and a maximum xylanase activity of 45 U/mL when it was cultivated in orange peel substrate. Substantial ethanol yields were achieved using lignocellulosic materials like brewer’s spent grain (BSG), orange peel (OP), and wheat bran (WB) as a carbon source in fermentation with no chemicals, which represents a zero waste environment-friendly ethanol production system. Ethanol yield on wheat bran was the highest of all evaluated substrates, reaching value of 34.9% at 7 days, being T. hirsuta able to degrade other hexoses and pentoses present in the structural polymers of cellulose and hemicellulose.

The aim of this study was to identify a Mexican native basidiomycete strain with antagonistic potential against phytopathogenic fungi. The antifungal activity of fifteen isolates against Fusarium oxysporum, Fusarium solani, Colletotrichum truncatum, Colletotrichum coccodes, Colletotrichum sp., Stemphylium solani, Botrytis cinerea, Epicoccum nigrum, Phomopsis sp. and Alternaria solani was studied in vitro using dual culture assay. An isolate of Macrolepiota sp (CS185) exhibited the strongest ability to compete on potato dextrose agar medium (PDA) causing 22.2 up to 46.6% inhibition in seven out of ten pathogens evaluated. The addition of 30% Macrolepiota sp. liquid filtrates (from 7 day-cultures in potato dextrose broth) to PDA plates inhibited 66% of A. solani growth. Application of Macrolepiota sp. CS185 culture fluids to control early blight disease caused in tomato plants by A. solani under greenhouse conditions diminished disease incidence and percent of damage. The length of the stems, the roots and the foliar area increased in the treated plants and, above all, the foliar symptoms decreased by 73%. Sesquiterpene lactones and quinones identified in culture fluids might be responsible for Macrolepiota antifungal activity. This study proves that Macrolepiota sp. CS185 culture filtrates have potential in biocontrol of early blight disease.

One primary drawback of enzyme catalysis at industrial scale is the short-term service life of the enzymes, they lose their activity due to oxidation or other processes which results in less stability and a shorter lifetime thereby rendering them less efficient. An effective way to increase the stability of the enzymes is to attach them to nanoparticles. In this work, the polymer Eudragit&reg, L 100-55 sensitive to pH was used to prepare laccase polymeric nanoparticles by the double-emulsion solvent evaporation approach. The size and morphology of the nanoparticles obtained were evaluated&mdash, as well as the encapsulation efficiency and zeta potential. pH effect on activity and stability was compared between free and immobilized laccase. Their stability was also determined in a sequential assay involving acidic pHs up to alkaline ones. The nanoparticles had a spherical shape with a mean size of 147 nm, zeta potential of &minus, 22.7 mV at pH 7.0 and load efficiency of 87%. The optimum pH of both free and immobilized laccases was 3.0, being the nanoparticles more stable at acidic pHs. Thus, this would be the first report of obtaining laccase nanoparticles with potential application in animal feed due to the stability conferred to enzymatic activity at pHs similar to those present in the gastrointestinal tract of rabbits, which would allow their potential use in animal feed.

One primary drawback of enzyme catalysis at industrial scale is the short term service life of the enzymes. Enzymes lose their activity due to oxidation or other processes which results in less stability and a shorter lifetime thereby rendering them less efficient. An effective way to increase the stability, longevity and reusability of the enzymes is to attach them to nanoparticles by applying the double-emulsion technique. In this work the polymer Eudragit® L 100-55 sensitive to pH was used to prepare laccase polymeric nanoparticles by the double-emulsion solvent evaporation approach. The size and morphology of the nanoparticles obtained was evaluated by Scanning Electron Microscope and particle size distribution was assessed by Photon Correlation Spectroscopy. Encapsulation efficiency and zeta potential were calculated. The effect of pH on laccase activity and stability was compared between free laccase and the immobilized one. Their stability was also determined in a sequential assay involving acidic pHs up to alkaline ones. The nanoparticles had a spherical shape with a mean size of 289 nm, zeta potential of -22.7 mV at pH 7.0, and load efficiency of 87%. The optimum pH of both free and immobilized laccases was 3.0, being the nanoparticles more stable at acidic pHs (2.0-4.0). Howevwe, this last kept 80% of enzyme activity at pH 2.0 approx., after 24 h. The polymer Eudragit® L 100-55 also conferred them resistance towards the pHs usually found at the gastrointestinal tract. These results suggest the potential use of nanoparticles as adjuvants in animal feed, serving as carriers for oral laccase delivery.

IMMUNEPOTENT CRP is a dialyzable leukocyte extract obtained from bovine spleen with immunomodulatory and antitumour properties; therefore, when administrated as an adjuvant therapy for cancer patients, it has increased their survival and quality of life. The bioavailability of any orally administered compound can be reduced due to gastrointestinal enzymes. In this study, we evaluated if IMMUNEPOTENT CRP is resistant to the treatment with different enzymes (proteases, nucleases, polysaccharide-degrading enzymes or lipase), using as parameters for biological activity measurement its in vitro antitumour and antioxidant properties and in vivo the antitumour effect of IMMUNEPOTENT CRP treated with proteinase K. In conclusion, we consider necessary to include the antioxidant and cytotoxic activity on the MCF-7 cancer cell line as parameters for the quantitative determination of biological activity or potency tests for batch release. Additionally, the results showed that different enzymatic treatments do not affect the antitumour and antioxidant activities of IMMUNEPOTENT CRP in vitro, suggesting that this product can be administered orally without any loss of biological activity. Furthermore, IMMUNEPOTENT CRP treatment with proteinase K increases the antitumour activity in vivo.

Pleurotus ostreatus is an edible mushroom with many functional compounds. In previous studies, it has been shown that solid-state fermentation of Phaseolus vulgaris and Avena sativa by this fungus increases essential amino acids, diminishes antinutrients as well as improves the bioaccessibility of different compounds. The objective was to obtain a functional food based on biotransformed ingredients by solid-state fermentation with P. ostreatus and evaluate its properties and the effect the food processing has on them. Results showed that cookies made with fermented flours provide a higher antioxidant capacity, when compared with cookies made without fermented flours, while maintaining its antioxidant capacity after simulated digestion assay. They also showed higher content of bioavailable protein compared to wheat control cookie, higher fiber content and lower sugars compared to commercial cookies. Cookies had good sensorial acceptability. This study shows the potential use of biotransformed cereals and legumes by P. ostreatus in functional foods.

Functionalization of thin films with organic ligands has been the subject of intense research due to their potential application as heterogeneous molecular nanosystems. In this work, self-assembled monolayers of thiols (16-mercaptohexadecanoic acid and 11-mercaptoundecanol) were used to bind laccase (from Pycnoporus sanguineus CS43) to nanostructured gold thin films obtained by DC sputtering. Sputtering power, sputtering pressure and substrate temperature were optimized to enhance the activity of the immobilized biomolecules. Scanning electron microscopy, confocal microscopy, X-ray diffraction and UV-vis spectroscopy were used to characterize the SAM-functionalized gold substrates. Our results demonstrate that the highest immobilized enzyme activity values can be achieved on substrates of surface roughness ˜200 nm and Au particle size of about 14 nm. The outstanding quality of the as-prepared substrates makes them particularly attractive as bionanosensors.

Background Mortality due to tuberculosis (TB) has increased due to the development of drug resistance, the mechanisms of which have not been fully elucidated. Our research group identified a low expression of lipF gene in Mycobacterium tuberculosis clinical isolates with drug resistance. The aim of this work was to evaluate the effect of lipase F (LipF) expression on mycobacterial drug resistance. Results The effects of expressing lipF from Mycobacterium tuberculosis in Mycobacterium smegmatis on resistance to antituberculosis drugs were determined with resazurin microtiter assay plate and growth kinetics. Functionality of ectopic LipF was confirmed. LipF expression reduced the rifampicin (RIF) and streptomycin (STR) minimum inhibitory concentration (MIC) from 3.12 μg/mL to 1.6 μg/mL and 0.25 μg/mL to 0.06 μg/mL respectively, moreover a reduced M. smegmatis growth in presence of RIF and STR compared with that of a control strain without LipF expression (p p Conclusions LipF expression was associated with increased RIF and STR sensitivity in mycobacteria. Reduced LipF expression may contribute to the development of RIF and STR resistance in Mycobacterium species. Our findings provide information pertinent to understanding mycobacterial drug resistance mechanisms.

Laccases have attracted a great deal of interest because of their remarkable ability for the degradation of synthetic dyes present in wastewaters. New laccase producing sources with robust operational and functional properties are being continuously explored. In this work, the potential for the decolorization and detoxification of synthetic dyes was evaluated in two Mexican strains of the genus Trametes. The decolorization capacity of Trametes maxima LE130 and Trametes sp. LA1 was tested in solid and liquid media. The phytotoxicity of the degradation products was determined using Raphanus sativus and Pisum sativum seeds. In solid media, both strains showed a higher decolorization capacity (p ≤ 0.05) than Phanerochaete chrysosporium ATCC 24725, which is known to be very efficient in lignin and dye-degradation. They produced laccase as the main ligninolytic enzyme; T. maxima LE130 secreted a single isoform of 43.9 kDa, while Trametes sp. LA1 produced three isoforms of 67.3, 58.6 and 52.7 kDa, respectively. Trametes sp. LA1 culture fluids were capable of decolorizing and detoxifying chemically diverse dyes (anthraquinonic dye Remazol Brilliant Blue R, azoic Reactive Black 5 and triphenylmethane Crystal Violet) without the addition of redox mediators. Therefore, this could be considered as a new laccase source which could be potentially competitive in the bioremediation of dye-containing wastewaters.

The global rise in urbanization and industrial activity has led to the production and incorporation of foreign con‑ taminant molecules into ecosystems, distorting them and impacting human and animal health. Physical, chemical, and biological strategies have been adopted to eliminate these contaminants from water bodies under anthropo‑ genic stress. Biotechnological processes involving microorganisms and enzymes have been used for this purpose; specifcally, laccases, which are broad spectrum biocatalysts, have been used to degrade several compounds, such as those that can be found in the efuents from industries and hospitals. Laccases have shown high potential in the biotransformation of diverse pollutants using crude enzyme extracts or free enzymes. However, their application in bioremediation and water treatment at a large scale is limited by the complex composition and high salt concentra‑ tion and pH values of contaminated media that afect protein stability, recovery and recycling. These issues are also associated with operational problems and the necessity of large-scale production of laccase. Hence, more knowledge on the molecular characteristics of water bodies is required to identify and develop new laccases that can be used under complex conditions and to develop novel strategies and processes to achieve their efcient application in treating contaminated water. Recently, stability, efciency, separation and reuse issues have been overcome by the immobilization of enzymes and development of novel biocatalytic materials. This review provides recent information on laccases from diferent sources, their structures and biochemical properties, mechanisms of action, and applica‑ tion in the bioremediation and biotransformation of contaminant molecules in water. Moreover, we discuss a series of improvements that have been attempted for better organic solvent tolerance, thermo-tolerance, and operational stability of laccases, as per process requirements., Instituto de Fisiología Vegetal

[EN] The aim of the research was to determine the impact of fermentation with Pleurotus ostreatus on kidney beans, black beans, and oats. The results indicate that the fungus has a positive effect on the substrates when compared to the controls. The antioxidant activity (39.5% on kidney beans and 225% on oats in relation to the controls) and content of total polyphenols (kidney beans three times higher regarding the controls) increased significantly by the presence of the fungus mycelium, even after simulated digestion. There was a significant increase in protein digestibility (from 39.99 to 48.13% in black beans, 44.06 to 69.01% in kidney beans, and 63.25 to 70.01% in oats) and a decrease of antinutrient tannins (from 65.21 to 22.07 mg in black beans, 35.54 to 23.37 in kidney beans, and 55.67 to 28.11 in oats) as well as an increase in the contents of some essential amino acids. Overall, this fermentation treatment with Pleurotus ostreatus improved the nutritional quality of cereals and legumes, making them potential ingredients for the elaboration and/or fortification of foods for human nutrition., We would like to thank Consejo Nacional de Ciencia y Tecnologa (CONACyT) for financially supporting E.E.-P to obtain her Ph.D. (scholarship 446871).

In this study, in-house isolated laccase isoforms, i.e., Lac-I and Lac-II of the basidiomycete Pycnoporus sanguineus (CS43), were evaluated in relation to their Remazol Brilliant Blue R (RBBR) dye degradation capacity. A modified Dhouib medium additionally supplemented with 3% ethanol as a secondary inducer was used to propagate P. sanguineus CS43 for enhanced production of laccase under liquid state fermentation. The crude laccase extract was purified by passing through ion exchange diethylaminoethanol (DEAE)-Sepharose and gel filtration-based Sephadex G-200 column chromatography. The purified laccase fractions were subjected to the electrophoresis, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed two laccase isoforms Lac-I and Lac-II with 66 and 68 kDa, respectively. To explore the industrial applicability, for RBBR dye, degradation efficiencies ranged from 82 to 88% after 3 h of incubation for both; Lac-I and Lac-II at both concentrations were recorded. However, with 8 U/mL, the degradation ranged between 70 to 80% during the first 5 min of incubation. Enhanced degradation of RBBR dye was obtained in the presence of violuric acid and N-hydroxypthalamide as laccase mediators. Finally, using RBBR as a substrate kinetic characterization of both Lac-I and Lac-II isoforms was performed that revealed K m (0.243 and 0.117 mM for Lac-I and Lac-II) and V max (1.233 and 1.012 mM/Sec for Lac-I and Lac-II) values, respectively.

Laccases are enzymes that have the ability to catalyze the oxidation of a wide spectrum of phenolic compounds with the four-electron reduction of molecular oxygen to water. The active site of those proteins contains four copper ions, classified into three types. Laccases are interesting enzymes for study from the point of view of their structure, function and application because of their role in lignin degradation. Structural studies of two thermostable laccases produced by the strain Pycnoporus sanguineus CS43 (PsLacI and PsLacII) were performed. Both isoforms of PsLac show high thermal stability, at 60°C and 50°C, respectively, and they remained active at a high concentration of organic solvents. However, PsLacI has a higher thermal and pH stability and tolerance against inhibitors, and is a more efficient catalyst for ABTS and DMP (laccases substrate) than PsLacII. Based on the determined crystal structures we achieved insights into the structural factors relevant for the enzymatic properties of PsLacI and PsLacII. N-glycosylation site Asn354, which is very often present in structures of fungal laccases from other species, was not present in PsLac. This observation may be of particular significance due to the close distance between Asn354 and the substrate-binding pocket. This results in better access to the hydrophobic cavity for a particular substrate. Furthermore, we identified significant differences in the region of substrate-binding pocket, which confer PsLacI a markedly better performance than PsLacII.

Laccases have attracted a great deal of interest because of their remarkable ability for the degradation of synthetic dyes present in wastewaters. New laccase producing sources with robust operational and functional properties are being continuously explored. In this work, the potential for the decolorization and detoxification of synthetic dyes was evaluated in two Mexican strains of the genus Trametes. The decolorization capacity of Trametes maxima LE130 and Trametes sp. LA1 was tested in solid and liquid media. The phytotoxicity of the degradation products was determined using Raphanus sativus and Pisum sativum seeds. In solid media, both strains showed a higher decolorization capacity (p &le, 0.05) than Phanerochaete chrysosporium ATCC 24725, which is known to be very efficient in lignin and dye-degradation. They produced laccase as the main ligninolytic enzyme, T. maxima LE130 secreted a single isoform of 43.9 kDa, while Trametes sp. LA1 produced three isoforms of 67.3, 58.6 and 52.7 kDa, respectively. Trametes sp. LA1 culture fluids were capable of decolorizing and detoxifying chemically diverse dyes (anthraquinonic dye Remazol Brilliant Blue R, azoic Reactive Black 5 and triphenylmethane Crystal Violet) without the addition of redox mediators. Therefore, this could be considered as a new laccase source which could be potentially competitive in the bioremediation of dye-containing wastewaters.

Laccase from Pycnoporus sanguineus CS43 was successfully immobilized onto Immobead-150 and Eupergit-C by covalent binding and by entrapment in LentiKats. The highest immobilization was onto Immobead-150 (97.1±1.2%) compared to the other supports, LentiKats (89±1.1%) and Eupergit-C (83.2±1.4%). All three immobilized enzyme systems showed increased thermostability and better mechanical properties than free laccase. Moreover, after 5 cycles of reuse of these systems, 90% of initial laccase activity was retained. Immobead-150 and LentiKats systems exhibited the highest efficiencies in removal of m-cresol under the combined actions of biodegradation and adsorption, while laccase entrapped in LentiKats showed a high ability for degradation of m-cresol within 24h. In addition, the typical Michaelis-Menten enzymatic model effectively described the kinetic profile of m-cresol degradation by the enzyme entrapped in LentiKats. Based on the results obtained in the present study, it can be established that the immobilized biocatalysts developed here possess significant potential for wastewater treatment.

Rotavirus vaccine was developed using the most prominent G and P genotypes circulating in children population. Therefore, severe gastroenteritis has been reduced around the world. This study investigated the G and P rotavirus genotypes circulating in children from two hospitals in the city of Chihuahua, Mexico. Additionally, polyclonal antibodies against Rotavirus Wa strain were used to determine their homotypic and heterotypic reactivity to both P[8] and P[4] genotypes. G1, G2, and G3 VP7 genotypes and P[8] and P[4] VP4 genotypes were detected in common and uncommon combinations as well as mixed infectious. The predominant combination was G1P[8]. Phylogenetic analysis of VP4 gene revealed the presence of P[8]-1 and P[8]-3 lineages of P[8] genotype and P[4]-5 lineage of P[4] genotype. All but five G1P[8] rotavirus were detected by polyclonal anti-Rotavirus Wa strain. Mutation analysis revealed differences in three of the four neutralizing epitopes previously reported to VP8* subunit of VP4 protein. Results of this study offer insights over genetic variants of field rotavirus that could be detected in a homotypic and heterotypic way by antibodies elicited to rotavirus with P[8] genotype. [Int Microbiol 2016; 19(1):27-32].

Sphingomyelinase (SMase) activity was measured in Entamoeba histolytica particulate and soluble subcellular fractions. The effects on SMase of incubation time, total protein concentration, pH, and several divalent cations were determined. SMase-C and other unidentified esterase activity were detected in soluble and particulate fractions. SMase-C was 94.5-96.0% higher than the unidentified esterase activity. Soluble and insoluble SMase-C specific activities increased with protein dose and incubation time. Soluble and insoluble SMase-C activities were maximum at pH 7.5 and were dependent on Mg(2+), Mn(2+), or Co(2+), and inhibited by Zn(2+), Hg(2+), Ca(2+), and EDTA. SMase-C was active in the pH range of 3-10 and its maximum activity was at pH 7.5. The soluble and insoluble SMases have remarkably similar physicochemical properties, strongly suggesting that E. histolytica has just one isoform of neutral SMase-C that had not been described before and might be essential for E. histolytica metabolism or virulence.

Flocculation is an important ability of some yeasts used in the alcohol fermentation industry. This characteristic depends on the presence and activity of FLO genes, which encode for several proteins called zymolectins. To identify the type of FLO genes present in some lager and wild yeasts, we designed specific primers for PCR amplification of seven FLO genes. DNA samples were obtained from six lager, one baker's, and three wild Saccharomyces strains. Our results showed that the incidence of flocculation genes is variable. Only FLO11 was present in all yeast strains. In contrast, Lg-FLO was detected exclusively in lager strains, but they lacked the FLO5 gene. FLO8 was present in all except the baker's strain, whereas FLO1, -9, and -10 showed a polymorphic distribution. Analysis of FLO gene content and flocculation ability indicated that the number of different FLO genes is related to the intensity of flocculation. Therefore, detection of FLO genes could provide a valuable tool for yeast strain selection.

A survey to isolate native white rot basidiomycetes from Northeast Mexico was conducted in the forests of the Sierra Madre Oriental in the state of Nuevo Leon. A total of 92 isolates from at least 20 different genera, were screened on Bran-Flakes solid plate cultures for the production of ligninolytic oxidases and/or peroxidases with guaiacol and o-anisidine as substrates; their lignin depolymerizing potential using the polymeric dye Poly R 478; their ability to decolorize anthraquinonic (Remazol Brilliant Blue Reactive), azo (Acid Red 44) and triphenylmethane (Crystal Violet) dyes. Among all fungi tested, 15 isolates showed extensive decolorization of the three dyes within a week and gave a positive reaction in guaiacol and o-anisidine tests. Nine of them were also efficient degraders of Poly R-478. Two isolates (CS5 and CU1) showed decolorization of all dyes within 5 days, comparing favorably with reference strains of P. chrysosporium, Pleurotus ostreatus, and Bjerkandera adusta. Decolorization was associated with laccase activity in both isolates and reached 90% or more for all dyes within 24 h in 8-day-old liquid cultures. The coupling of pairs 2,4-dichlorophenol + 4-aminoantipyrine and 3-dimethylaminobenzoic acid + 3-methyl-2-benzothiazolinone hydrazone, strongly suggest that the laccases of both strains correspond to those considered of high redox potential. These strains are considered good candidates for bioremediation of dye polluted effluents due to their ligninolytic potential and decolorizing performance.

With the aim of identifying and exploiting the mycological resources available in the Mexican Sierra Madre Oriental, the lignocellulolytic and pectinolytic potential of autochthonous fungi were evaluated in the present work. A solid media selection system was established in which 74 isolated strains were tested and compared to six international reference strains. The macrofungi Xylaria sp CS121, Inonotus sp CU7, Basidiomycete CH32, Basidiomycete CH23, Xylaria poitei, and Trametes maxima CU1 showed the highest cellulolytic and pectinolytic potential. The greatest lignolytic capability was exhibited by T. maxima CU1 and Pycnoporus sanguineus CS43. Under stirred submerged culture, T. maxima CU1 (cellulases, cellobiose dehydrogenase, manganese peroxidase (MnP), and laccase, with 200, 359, 51, and 267 U/L, respectively) and Xylaria sp CS121 (198 U/L of xylanases) were the highest enzymatic producers. Under stationary conditions, the best producers were Inonotus sp CU7 for cellulases, P. sanguineus CS43 for cellobiose dehydrogenase and laccase, and T. maxima CU1 for xylanases and MnP (242, 467, 35, 165, and 31 U/L, respectively). These results demonstrate the efficiency of enzymatic profiling as a tool for enzyme discovery with Mexican native fungi.

A native strain of Pycnoporus sanguineus (CS2) was selected because of its lignocellulolytic potential on a color-based agar plate screening system; pectinolytic and amylolytic capabilities were also assessed. The effects of different lignocellulosic substrates under submerged cultures of selected fungi on the enzymatic production of cellulases, xylanases, amylases, and laccases were evaluated. Seven different treatments, in which different combinations of wheat straw (WS), wheat bran (WB), orange peels (OP), grapefruit peels (GP), and apple peels (AP) were used, were established. Controls of 2% (w/v) carboxymethyl-cellulose (CMC) and xylan were used in a rich medium for basidiomycetes (RMB). The highest titers were achieved using OP-based cultures, with large titers of CMCases (33.5 U), avicelases (15.7 U), ß-D-glucosidase (72.9U), and xylanases (18.3 U). The best levels of amylase and laccase activity were obtained in the RMB plus CMC (RMB-CMC) (7.2 U) and in the medium OF/AP/GP (6.4 U), respectively. The relative molecular sizes of cellulase, xylanase, and amylase were 66.2, 56.5, and 90.8 kDa, respectively. Laccase and amylase had maximum activities at 60 °C whereas cellulase and hemicellulase had maximum activities at 70 °C. The optimal pH for cellulases, xylanases, and amylases was 5.0 in every case, and more than 95% activity was observed at pH 6. These results reveal some efficient operating parameters for the application of these enzymatic extracts as adjuvants to improve animal nutrition.

The production of thermostable laccases from a native strain of the white-rot fungus Pycnoporus sanguineus isolated in Mexico was enhanced by testing different media and a combination of inducers including copper sulfate (CuSO4). The best conditions obtained from screening experiments in shaken flasks using tomato juice, CuSO4, and soybean oil were integrated in an experimental design. Enhanced levels of tomato juice as the medium, CuSO4 and soybean oil as inducers (36.8% (v/v), 3 mmol/L, and 1% (v/v), respectively) were determined for 10 L stirred tank bioreactor runs. This combination resulted in laccase titer of 143,000 IU/L (2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid), pH 3.0), which represents the highest activity so far reported for P. sanguineus in a 10-L fermentor. Other interesting media resulting from the screening included glucose-bactopeptone which increased laccase activity up to 20,000 IU/L, whereas the inducers Acid Blue 62 and Reactive Blue 19 enhanced enzyme production in this medium 10 times. Based on a partial characterization, the laccases of this strain are especially promising in terms of thermostability (half-life of 6.1 h at 60 °C) and activity titers.

In this paper we study the calcium uptake in the erythrocyte, a non-excitable cell. This uptake is performed through a passive transport system with two kinetic components (Michaelis-Menten and Hill). The uptake of calcium seems to be driven by voltage through its electrophoretical effect. Lead is capable of inhibiting calcium uptake in a non-competitive manner. As it has been described in other systems, lead is also capable of inhibiting calcium efflux by inhibiting Ca(Mg)-ATPase. Under physiological conditions, the function of ATPase reduces the effect of lead on calcium influx. However, in chronic intoxication a small increment of intracellular calcium is observed, indicating that lead is affecting calcium efflux mainly. We discuss the effects of lead on calcium equilibrium in erythrocytes.

A 3 years study was conducted to determine the evolution of blood lead concentrations (PbB) and free erythrocyte protoporphyrins (FEP) as indicators of absorption and biochemical damage in children (7-12 years old) living within one kilometer of a metallurgic factory that processes lead in Torreón, Coahuila, México. In addition, neuromuscular conduction velocity, motor coordination and IQ where determined as indicators of physiological damage. During this period of time the children showed increased lead blood concentrations of 19.2 ± 4.5 μg dl -1 at the beginning of the study to 27.5±4.9 μg dl -1 at the end. Likewise, FEP increased from 56.6±20.0 μg dl-1 to 92.9 ± 28.9 μg dl-1. Physiological damage was evident since motor coefficients as well as IQ tests were reduced significantly when compared to children (7 -12 years old) living 4.5 km away from the factory who showed PbB=8.9 ± 1.3 μg dl-1 and FEP=16.9 ± 4.7 μg dl-1 throughout the study. Based on these data it was possible to define three different groups with regards to lead toxicity sensitivity.

The effect of laccase from Trametes maxima CU1 on physicochemical quality of bread was evaluated. Laccase treatment was 0.05% lower than the control in height, 0.33% higher in weight loss and reduced 17.71% the hardness of bread. In color, treatment with laccase was 18.72, 8.51 and 0.61% higher in L*, h, and C*, respectively. Chemical parameters of laccase treatment 12.00, 14.10 and 41.62% were higher than control in soluble arabinoxylans, protein and total phenols content, respectively. Based on the results obtained in the present study, laccase from T. maxima CU1 can be considered a good option for extraction and application as improver of the physicochemical quality of bread at industrial level.

We recently reported that DAG (diacylglycerol) generated during sphingomyelin synthesis plays an important role in protein kinase C activation and cell proliferation in Madin–Darby canine kidney cells [Cerbon and Lopez-Sanchez (2003) Biochem. J. 373, 917–924]. In yeast cells, IPC (inositol phosphoceramide) synthase catalyses the transfer of phosphoinositol from phosphatidylinositol to ceramide to form IPC and generates DAG. In the present study, we found that, during the G1 to S transition after N2-starvation, there was a significant increase in the synthesis of IPC accompanied by a progressive increase (up to 6-fold) in the level of DAG. The increased DAG levels coincided with decrements in ceramide and sphingoid base levels, conditions that are adequate for the activation of putative protein kinase C required for the G1 to S transition and proliferation of yeast cells. To separate the role of DAG generated during IPC synthesis from that originating from other sources, we utilized β-chloroalanine and myriocin, inhibitors of serine:palmitoyl-CoA transferase, the first committed step in sphingolipid synthesis, to avoid accumulation of sphingolipid intermediates. When the synthesis of sphingolipids was inhibited, DAG accumulation was significantly decreased and the G1 to S transition was blocked; such blockage was avoided by metabolic complementation with phytosphingosine. The DAG/ceramide ratio was 0.27 and it changed to 2.0 during growth re-initiation, suggesting that the synthesis of phosphosphingolipids could act to switch growth arrest (increased ceramide) to a mitogenic signal (increased DAG), and that this signalling process is preserved in yeast and mammalian cells.

The aim of this study was to examine lead (Pb) intestinal absorption, its mobilization and redistribution during lactation in rats chronically exposed to lead. Lead and calcium (Ca) concentrations were measured in blood, milk, femurs, liver and kidney samples obtained from pregnant and lactating mother rats which were subjected to different schedules of exposure to Pb: 158 days before and during lactation (group A), 144 days before lactation (group B) and 14 days only during lactation (group C). Results were compared to those of non-pregnant but Pb- exposed matched rats and non-exposed control rats. In groups A and B during lactation, Pb in blood (PbB), liver and kidney increased while Pb in bone decreased. Since there was not an external source of Pb in group B during lactation, the results indicate resorption of Pb in bone as the main source of Pb in the organism. In group A, there was an additional increase of PbB when compared to group B as a result of Pb intestinal absorption. In group C a significant increase in PbB due to intestinal absorption and deposit in bone was found when compared to non- pregnant 144 days old rats, suggesting that the three processes intestinal absorption, bone resorption and bone absorption were taking place. These data indicate that Pb stored in bone as a result of prior maternal exposure, should be considered as a major source of self intoxication and of Pb in milk available to suckling pups.

In this work, the biocatalytic ability of laccases from filtered culture supernatant of Pycnoporus sanguineus was evaluated without mediators and under mild reaction conditions. This 100 U L -1 laccase cocktail removed 50% Diclofenac, 97% β-Naphthol and 71% 2,4 Dichlorophenol within 8 h of reaction and 78% for 5,7-Diiodo-8-hydroxyquinoline within 3.5 h; at initial concentrations of 10 mg L-1 and at 25 °C. Furthermore, this enzyme cocktail also removed in excess of 53% all tested compounds in a real groundwater sample from northwestern Mexico. In comparison with purified laccases, the use of cocktail offers operational advantages since additional purification steps can be avoided. © 2015 The Authors.

In this study, the purification and characterization of two thermostable laccases produced by Pycnoporus sanguineus CS43 (LacI and LacII) were performed. Also, their biotechnological potential was assessed through the degradation of endocrine disrupting chemicals (EDCs). Laccases were purified by ultrafiltration, ion exchange (IEX) and hydrophobic interaction chromatography achieving specific activities close to 285 U mg -1 . The molecular weights of LacI and LacII, determined by SDS-electrophoresis, were 68 and 66 kDa, respectively. Both laccases showed high amino acid sequence similarity (91%) between them and high thermostability, at 50 and 60 °C (half-lives of 277.7 and 18 h for LacI, 35.8 and 2.25 h for LacII). The isoforms oxidized common laccase substrates such as 2,2′-azino-bis (3-ethylbenzthiazoline-6- sulfonate (ABTS), 2,6-dimethoxyphenol (DMP) and guaiacol at acidic pH conditions. ABTS was the most efficient substrate, showing high specificity constants of 74,816 and 36,746 mM -1 s -1 for Lac I and LacII, and Michaelis constants (K m ) of 6.9 and 12.2 μM respectively at pH 3. Both purified laccases remained active at high concentrations of organic solvents (acetonitrile, ethanol and acetone), with an IC 50 (v/v) of >64%, 55% and 47% for LacI, and 33%, 52% and 31% for LacII, respectively. LacI and LacII were tested to degrade EDCs, nonylphenol and triclosan, with more than 95% removal after 8 h of treatment with 100 U/L at pH 5 as determined by means of HPLC. The high thermostability, unique Michaelis-Menten kinetic parameters, and organic solvent tolerance demonstrated for the isoforms produced by P. sanguineus CS43 render them promising candidates for industrial applications. LacI exerted a higher thermal and pH stability, tolerance against inhibitors and was a more efficient catalyst for ABTS and DMP than LacII. © 2014 The Authors.

The biodegradation of organic compounds present in water at trace concentration has become a critical environmental problem. In particular, enzymatic oxidation by fungal laccases offers a promising alternative for efficient and sustainable removal of organic pollutants in water. In this work, the biocatalytic ability of laccases from the Pycnoporus sanguineus CS43 fungus was evaluated. A filtered culture supernatant (laccase cocktail) evidenced an enhanced biotransformation capability to remove common endocrine-disruptor compounds (EDCs), such as bisphenol A, 4-nonylphenol, 17-α-ethynylestradiol and triclosan. A biodegradation of around 89–100 % was achieved for all EDCs using synthetic samples (10 mg L−1) and after the enzymatic treatment with 100 U L−1 (50.3 U mg −1). The biodegradation rates obtained were fitted to a first order reaction. Furthermore, enzymatic biocatalytic activity was also evaluated in groundwater samples coming from northwestern Mexico, reaching biotransformation percentages between 55 and 93 % for all tested compounds. As far as we know this is the first study on real groundwater samples in which the enzymatic degradation of target EDCs by a laccase cocktail from any strain of Pycnoporus sanguineus was evaluated. In comparison with purified laccases, the use of cocktail offers operational advantages since additional purification steps can be avoided.