Exploring the performance limits of a sulfidogenic UASB during the long-term use of crude glycerol as electron donor

SOx contained in flue gases and S-rich liquid effluents can be valorized to recover elemental sulfur in a two-stage bioscrubbing process. The reduction of sulfate to sulfide is the most crucial stage to be optimized. In this study, the long-term performance of an up-flow anaerobic sludge blanket (UASB) reactor using crude glycerol as electron donor was assessed. The UASB was operated for 400 days with different sulfate and organic loading rates (SLR and OLR, respectively) and a COD/S-SO42− ratio ranging from 3.8 g O2 g−1 S to 5.4 g O2 g−1 S. After inoculation with methanogenic, granular biomass, the competition between sulfate-reducing and methanogenic microorganisms determined to what extent dissolved sulfide and methane were produced. After the complete washout of methanogens, which was revealed by next-generation sequencing analysis, the highest S-EC was reached in the system. The highest average sulfate elimination capacity (S-EC = 4.3 kg S m−3d−1) was obtained at a COD/S-SO42− ratio of 5.4 g O2 g−1 S and an OLR of 24.4 kg O2 m−3d−1 with a sulfate removal efficiency of 94%. The conversion of influent COD to methane decreased from 12% to 2.5% as the SLR increased while a large fraction of acetate (35% of the initial COD) was accumulated. Our data indicate that crude glycerol can promote sulfidogenesis. However, the disappearance of methanogens in the long-term due to the out competition by sulfate reducing bacteria, lead to such large accumulation of acetate

A novel, highly potent nadph-dependent cytochrome P450 reductase from waste liza klunzingeri liver

The use of marine enzymes as catalysts for biotechnological applications is a topical subject. Marine enzymes usually display better operational properties than their animal, plant or bacterial counterparts, enlarging the range of possible biotechnological applications. Due to the fact that cytochrome P450 enzymes can degrade many different toxic environmental compounds, these enzymes have emerged as valuable tools in bioremediation processes. The present work describes the isolation, purification and biochemical characterization of a liver NADPH-dependent cytochrome P450 reductase (CPR) from the marine fish Liza klunzingeri (LkCPR). Experimental results revealed that LkCPR is a monomer of approximately 75 kDa that is active in a wide range of pH values (6–9) and temperatures (40–60 °C), showing the highest catalytic activity at pH 8 and 50 °C. The activation energy of the enzyme reaction was 16.3 kcal mol−1 K−1. The KM values for cytochrome C and NADPH were 8.83 μM and 7.26 μM, and the kcat values were 206.79 s−1 and 202.93 s−1, respectively. LkCPR displayed a specific activity versus cytochrome C of 402.07 µmol min−1 mg1, the highest activity value described for a CPR up to date (3.2–4.7 times higher than the most active reported CPRs) and showed the highest thermostability described for a CPR. Taking into account all these remarkable catalytic features, LkCPR offers great potential to be used as a suitable biocatalyst

Tuning lipase B from Candida antarctica C–C bond promiscuous activity by immobilization on poly-styrene-divinylbenzene beads

Lipase B from Candida antarctica (CALB) is able to catalyze C–C bond formation. After immobilization onto a hydrophobic PS-DVB support, the activity increases when compared to that of the soluble or tan – the commercially available Novozyme 435 (being up to 6 fold more active). Our results show that although this activity is not related to the catalytic group, the promiscuous activity of CALB may be tuned via immobilization. In addition, we have show that the secondary structure of both immobilized enzymes is quite different, using FT-ATR-IR spectroscopy

Enzyme production of D-gluconic acid and glucose oxidase : successful tales of cascade reactions

This review mainly focuses on the use of glucose oxidase in the production of D-gluconic acid, which is a reactant of undoubtable interest in different industrial areas. The enzyme has been used in numerous instances as a model reaction to study the problems of oxygen supply in bioreactors. One of the main topics in this review is the problem of the generated side product, hydrogen peroxide, as it is an enzymeinactivating reagent. Different ways to remove hydrogen peroxide have been used, such as metal catalysts and use of whole cells; however, the preferred method is the coupling glucose oxidase with catalase. The different possibilities of combining these enzymes have been discussed (use of free enzymes, independently immobilized enzymes or co-immobilized enzymes). Curiously, some studies propose the addition of hydrogen peroxide to this co-immobilized enzyme system to produce oxygen in situ. Other cascade reactions directed toward the production of gluconic acid from polymeric substrates will be presented; these will mainly involve the transformation of polysaccharides (amylases, cellulases, etc.) but will not be limited to those (e.g., gluconolactonase). In fact, glucose oxidase is perhaps one of most successful enzymes, and it is involved in a wide range of cascade reactions. Finally, other applications of the enzyme have been reviewed, always based on the production of D-gluconic acid, which produces a decrease in the pH, a decrease in the oxygen availability or the production of hydrogen peroxide; in many instances, cascade reactions are also utilized. Thus, this review presents many different cascade reactions and discusses the advantages/drawbacks of the use of co-immobilized enzymes

RIP2 plays a role in cardiac Ca2+ mishandling prompted by chronic kidney disease

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Production, immobilization and synthesis of pharmacological derivatives of lipase B from Candida antarctica in Pichia pastoris

Lipase B from Candida antarctica (CALB) is widely used because of its excellent enantioselectivity. Producing this recombinant lipase in Pichia pastoris has advantages since it can be cultured in simple media and can reach high cell densities. This capability is especially important when using a constitutive promoter for lipase production, as here. The PPGK promoter is similar to the well-known PGAP promoter and also circumvents the need for inducing production with methanol, which is a hazard when used on a large scale and would increase the downstream production costs, which could be prohibitive for pharmaceutical products. This study tested two main fermentation strategies: continuous and fed-batch. In both cultures, different specific growth rates occurred (0.05, 0.10, 0.15 and 0.18h–1), and process parameters (qP, qS, YX/S, YP/X, YP/S) were evaluated in order to properly compare them. The highest specific production rate achieved with a continuous culture was 57.71 U/gX.h with µ=0.15 h–1 and 16 U/gX.h with µ=0.14 h–1 for a fed-batch culture. Productivity decreased dramatically near the µmax (0.18 h–1) for P. pastoris (57.6% lower). The best strategy for production was calculated over a three-month period. In both cases, the enzyme is secreted to the supernatant and purification is needed to ensure that only LIPB participates in further reactions. The immobilization process is ideal because purification and concentration is achieved in only one step, reusability is made possible, and in certain cases, stability and efficiency are boosted. Hydrophobic core-shell polymeric supports synthesized by a combined suspension and emulsion polymerization process have shown good potential for lipase immobilization procedures and were used in this study, compared to traditional supports such as Accurel, in order to determine their efficiency. After the enzyme was immobilized, the reactions included the resolution of (±)-1,3,5-O-benzyl-myo-inositol (DL-1) via acylation using vinyl acetate in hexane, and resolution of (±)-1,2-O- isopropylidene-3,6-di-O-benzyl-myo-inositol (DL-2) via acylation using vinyl acetate (solvent-free system). The support used directly affected the reaction, but trends were observed. In general, the recombinant lipase produced (LIPB) had higher resolutions than the commercial lipase (CALB, Novozym 435). In the resolution of DL-1 and DL-2 via transesterification (using different media), LIPB immobilized in Accurel or PS-co-DVB/PS-co-DVB showed more activity per enzyme molecule than CALB immobilized in similar supports, while when immobilized in PMMA-co-DVB/ PMMA-co-DVB the activities of the two enzymes were similar. The recombinant LIPB immobilized on PS-co-DVB proved to be the most efficient in the enantioselective resolution of both racemic derivatives, DL-1 and DL-2. The productivity for DL-2 resolution was 50% higher than the commercial Novozym 435, and the new derivative was operationally more stable than Novozym 435. The products obtained had a high level of purity (ee of 99% for both derivatives). Both products of the enantio-selective reaction, L-2 and L-5, obtained from the racemic derivatives (DL-1 and DL-2, respectively), are intermediates from different pharmacological pathways involved in the synthesis of building blocks for drugs that inhibit the etiological agent of Chagas disease, Trypanosoma cruzi

A decision tree to assess short-term mortality after an emergency department visit for an exacerbation of COPD: A cohort study

Background: Creating an easy-to-use instrument to identify predictors of short-term (30/60-day) mortality after an exacerbation of chronic obstructive pulmonary disease (eCOPD) could help clinicians choose specific measures of medical care to decrease mortality in these patients. The objective of this study was to develop and validate a classification and regression tree (CART) to predict short term mortality among patients evaluated in an emergency department (ED) for an eCOPD. Methods: We conducted a prospective cohort study including participants from 16 hospitals in Spain. COPD patients with an exacerbation attending the emergency department (ED) of any of the hospitals between June 2008 and September 2010 were recruited. Patients were randomly divided into derivation (50 %) and validation samples (50 %). A CART based on a recursive partitioning algorithm was created in the derivation sample and applied to the validation sample. Results: Two thousand four hundred eighty-seven patients, 1252 patients in the derivation sample and 1235 in the validation sample, were enrolled in the study. Based on the results of the univariate analysis, five variables (baseline dyspnea, cardiac disease, the presence of paradoxical breathing or use of accessory inspiratory muscles, age, and Glasgow Coma Scale score) were used to build the CART. Mortality rates 30 days after discharge ranged from 0 % to 55 % in the five CART classes. The lowest mortality rate was for the branch composed of low baseline dyspnea and lack of cardiac disease. The highest mortality rate was in the branch with the highest baseline dyspnea level, use of accessory inspiratory muscles or paradoxical breathing upon ED arrival, and Glasgow score <15. The area under the receiver-operating curve (AUC) in the derivation sample was 0.835 (95 % CI: 0.783, 0.888) and 0.794 (95 % CI: 0.723, 0.865) in the validation sample. CART was improved to predict 60-days mortality risk by adding the Charlson Comorbidity Index, reaching an AUC in the derivation sample of 0.817 (95 % CI: 0.776, 0.859) and 0.770 (95 % CI: 0.716, 0.823) in the validation sample. Conclusions: We identified several easy-to-determine variables that allow clinicians to classify eCOPD patients by short term mortality risk, which can provide useful information for establishing appropriate clinical care. Trial registration: NCT02434536

Characterization of Conserved and Promiscuous Human Rhinovirus CD4 T Cell Epitopes

This research was supported by UCM research special funds to P.A.R. and by the CAM research agency through grant IND2020/BMD-17364 to P.A.R.Human rhinovirus (RV) is the most common cause of upper respiratory infections and exacerbations of asthma. In this work, we selected 14 peptides (6 from RV A and 8 from RV C) encompassing potential CD4 T cell epitopes. Peptides were selected for being highly conserved in RV A and C serotypes and predicted to bind to multiple human leukocyte antigen class II (HLA II) molecules. We found positive T cell recall responses by interferon gamma (IFNγ)-ELISPOT assays to eight peptides, validating seven of them (three from RV A and four from RV C) as CD4 T cell epitopes through intracellular cytokine staining assays. Additionally, we verified their promiscuous binding to multiple HLA II molecules by quantitative binding assays. According to their experimental HLA II binding profile, the combination of all these seven epitopes could be recognized by >95% of the world population. We actually determined IFNγ responses to a pool encompassing these CD4 T cell epitopes by intracellular cytokine staining, finding positive responses in 29 out of 30 donors. The CD4 T cell epitopes identified in this study could be key to monitor RV infections and to develop peptide-based vaccines against most RV A and C serotypes.Depto. de Inmunología, Oftalmología y ORLFac. de MedicinaTRUEComunidad de MadridUniversidad Complutense de Madridpu

Novozym 435 : the “perfect” lipase immobilized biocatalyst?

Novozym 435 (N435) is a commercially available immobilized lipase produced by Novozymes. It is based on immobilization via interfacial activation of lipase B from Candida antarctica on a resin, Lewatit VP OC 1600. This resin is a macroporous support formed by polyIJmethyl methacrylate) crosslinked with divinylbenzene. N435 is perhaps the most widely used commercial biocatalyst in both academy and industry. Here, we review some of the success stories of N435 (in chemistry, energy and lipid manipulation), but we focus on some of the problems that the use of this biocatalyst may generate. Some of these problems are just based on the mechanism of immobilization (interfacial activation) that may facilitate enzyme desorption under certain conditions. Other problems are specific to the support: mechanical fragility, moderate hydrophilicity that permits the accumulation of hydrophilic compounds (e.g., water or glycerin) and the most critical one, support dissolution in some organic media. Finally, some solutions (N435 coating with silicone, enzyme physical or chemical crosslinking, and use of alternative supports) are proposed. However, the N435 history, even with these problems, may continue in the coming future due to its very good properties if some simpler alternative biocatalysts are not developed

Aqueous extraction of seed oil from mamey sapote (Pouteria sapota) after Viscozyme L treatment

In this study, aqueous enzymatic extraction (AEE) was evaluated during the process of obtaining oil from mamey sapote seed (OMSS). Viscozyme L enzyme complex was used at pH 4 and 50 ◦C during the optimization of the extraction process by central composite design and response surface methodology. Optimal conditions were: 3.5% (w/w) of enzyme (regarding the seed weight), 5.5 h of incubation time, 235 rpm of agitation rate, and 1:3.5 of solid-to-liquid ratio. These conditions enabled us to obtain an OMSS yield of 66%. No statistically significant differences were found in the fatty acid profile and physicochemical properties, such as the acid and iodine values and the percentage of free fatty acids, between the oil obtained by AEE or by the conventional solvent extraction (SE). However, the oxidative stability of the oil obtained by AEE (11 h) was higher than that obtained by SE (9.33 h), therefore, AEE, in addition to being an environmentally friendly method, produces a superior quality oil in terms of oxidative stability. Finally, the high oil content in mamey sapote seed, and the high percentage of oleic acid (around 50% of the total fatty acid) found in this oil, make it a useful edible vegetable oil