Despite the proven therapeutic role of capsaicin in human health, its usage is still hampered by its high pungency. In this sense, nonpungent capsaicin analogues as olvanil are a feasible alternative ...to the unpleasant sensations produced by capsaicin while maintaining a similar pharmacological profile. Olvanil can be obtained by a lipase‐catalyzed chemoenzymatic process. In the present work, recombinant Candida antarctica lipase B (CALB) was expressed in Pichia pastoris and subsequently immobilized by cross‐linked enzyme aggregate (CLEA) methodology for the synthesis of olvanil. The CALB‐CLEAs were obtained directly from the fermentation broth of P. pastoris without any purification step in order to assess the role of the contaminant proteins of the crude extract as co‐feeders. The CALB‐CLEAs were also bioimprinted to enhance the catalytic performance in olvanil synthesis. When CALB was precipitated with isopropanol, the obtained CALB‐CLEAs exhibited the highest activity in the synthesis of olvanil, regardless of the glutaraldehyde concentration. The maximum product synthesis was found at 72 hr obtaining 6.8 g L−1 of olvanil with a reaction yield of 16%. When CALB was bioimprinted with olvanil, the synthesis was enhanced 1.3 times, reaching 10.7 g L−1 of olvanil at 72 hr of reaction with a reaction yield of 25%. Scanning electron microscopy images indicated different morphologies of the CLEAs depending on the precipitating agent and the template used for bioimprinting. Recombinant CALB‐CLEAs obtained directly from the fermentation broth are a suitable alternative to commercial enzymatic preparations for the synthesis of olvanil in organic medium.
The effects of different concentrations of Hofmeister salts on the hydrolytic activity on triglycerides and the secondary structure of lipase B from Candida antarctica (CALB) were investigated. ...Structural changes after short- and long-time incubation at high salt concentrations were determined using circular dichroism (CD), fluorescence, and RMSD-RMSF simulations. At 5.2 M NaCl, the hydrolytic activity of CALB on tributyrin (TC4) and trioctanoin (TC8) was enhanced by 1.5 (from 817 ± 3.9 to 1228 ± 4.3 U/mg)- and 8.7 (from 25 ± 0.3 to 218 ± 2.3 U/mg)-folds compared with 0.15 M NaCl, respectively at pH 7.0 and 40 °C. An activity activation was seen with other salts tested; however, long-time incubation (24 h) did not result in retention of the activation effect for any of the salts tested. Secondary structure CD and fluorescence spectra showed that long-time incubation with NaCl, KCl, and CsCl provokes a compact structure without loss of native conformation, whereas chaotropic LiCl and CaCl2 induced an increase in the α-helical content, and kosmotropic Na2SO4 provoked a molten globule state with rich β-sheet content. The RMSD-RMSF simulation agreed with the CD analysis, highlighting a principal salt-induced effect at the α-helix 5 region, promoting two different conformational states (open and closed) depending on the type and concentration of salt. Lastly, an increase in the interfacial tension occurred when high salt concentrations were added to the reaction media, affecting the catalytic properties. The results indicate that high-salt environments, such as 2–5.2 M NaCl, can be used to increase the lipolytic activity of CALB on TC4 and TC8.
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•The high concentration of NaCl increased the hydrolytic activity of CALB on triglycerides of short and medium chain length.•CALB can be catalogued as a halotolerant lipase maintaining its activity and native structure at high salt concentrations.•CALB is a robust lipase able to restore its activity after 24h incubation at high salt concentrations.•The major effect of NaCl concentration was at the α-helix 5 of CALB promoting open and closed states (RMSD-RMSF simulation).
Ivermectin inhibits the replication of SARS-CoV-2 in vitro at concentrations not readily achievable with currently approved doses. There is limited evidence to support its clinical use in COVID-19 ...patients. We conducted a Pilot, randomized, double-blind, placebo-controlled trial to evaluate the efficacy of a single dose of ivermectin reduce the transmission of SARS-CoV-2 when administered early after disease onset.
Consecutive patients with non-severe COVID-19 and no risk factors for complicated disease attending the emergency room of the Clínica Universidad de Navarra between July 31, 2020 and September 11, 2020 were enrolled. All enrollments occurred within 72 h of onset of fever or cough. Patients were randomized 1:1 to receive ivermectin, 400 mcg/kg, single dose (n = 12) or placebo (n = 12). The primary outcome measure was the proportion of patients with detectable SARS-CoV-2 RNA by PCR from nasopharyngeal swab at day 7 post-treatment. The primary outcome was supported by determination of the viral load and infectivity of each sample. The differences between ivermectin and placebo were calculated using Fisher's exact test and presented as a relative risk ratio. This study is registered at ClinicalTrials.gov: NCT04390022.
All patients recruited completed the trial (median age, 26 IQR 19–36 in the ivermectin and 21–44 in the controls years; 12 50% women; 100% had symptoms at recruitment, 70% reported headache, 62% reported fever, 50% reported general malaise and 25% reported cough). At day 7, there was no difference in the proportion of PCR positive patients (RR 0·92, 95% CI: 0·77–1·09, p = 1·0). The ivermectin group had non-statistically significant lower viral loads at day 4 (p = 0·24 for gene E; p = 0·18 for gene N) and day 7 (p = 0·16 for gene E; p = 0·18 for gene N) post treatment as well as lower IgG titers at day 21 post treatment (p = 0·24). Patients in the ivermectin group recovered earlier from hyposmia/anosmia (76 vs 158 patient-days; p < 0.001).
Among patients with non-severe COVID-19 and no risk factors for severe disease receiving a single 400 mcg/kg dose of ivermectin within 72 h of fever or cough onset there was no difference in the proportion of PCR positives. There was however a marked reduction of self-reported anosmia/hyposmia, a reduction of cough and a tendency to lower viral loads and lower IgG titers which warrants assessment in larger trials.
ISGlobal, Barcelona Institute for Global Health and Clínica Universidad de Navarra.
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•Bulk compound chocolate products were developed as potential functional food.•Saturated fatty acids were less bioavailable using structured lipids.•The in vitro lipid digestion was ...reduced as well as its caloric impact.•Serum lipids were reduced in healthy C57BL/6J mice.
The bioavailability impact of serum lipids in compound chocolate products based on structured lipids was studied. Compound chocolate products containing fat with and without structured lipids were digested in vitro under simulated gastrointestinal lipolysis conditions and were studied in vivo in healthy C57BL/6J mice. The in vitro digestion results show that products containing structured lipids, milk compound chocolate filling and white compound coating, significantly reduced the release rate of Free Fatty Acids (FFA) and improved the caloric reduction between 12.49% and 13.71% compared to products without structured lipids, suggesting that FFA were not absorbed. Animal feeding studies revealed no adverse effects on the compound products intake; in fact, these products reduced total cholesterol, LDL-c, VLDL-c and triacylglycerols. The present work shows the relevance of developing functional compound chocolate as providing a potential healthy initiative through the biological effect of the bioactive ingredients incorporated.
Capsaicin analogues are typically synthetized by condensation of the amide group of vanillylamine with a fatty acid derivative. The enzyme of choice to perform this reaction is
Candida antarctica
...lipase B; however, this enzyme is unable to react with the phenolic substiruents of the vanillyl ring. So far, this can only be achieved chemically, resulting in capsaicin esters with higher lipophilicity and less irritation than the parent compound. In need of searching for new capsaicin derivatives, we investigated the transesterification of the phenolic OH group of capsaicin and several capsaicin analogues by
Candida antarctica
lipase A (CALA) with vinyl esters. Capsaicin esters were successfully synthetized with a reaction yield of 80.6% and 57.5% with vinyl butyrate and vinyl laurate, respectively. When the reactions were performed with capsaicin analogues of different acyl chain lengths, CALA exhibited a noticeable transesterification preference for medium-length capsaicin analogues. In an attempt to explore the catalytic limits of CALA, we performed the transesterification of capsaicin from
Capsicum
oleoresin. The results showed similar transesterification yields to those obtained with semi-pure capsaicin. Within our knowledge, this is the first report to achieve the synthesis of phenolic OH esters of capsaicin and capsaicin analogues by enzymatic means.
The publication of reports on geo-political risks in the world sponsored by intelligence agencies, university institutes and think tanks are valuable instruments in societies that are being ...increasingly exposed to the effects of globalisation. Although all express mention of geo-political risks of a maritime nature is absent from these documents, it is an interesting exercise to determine: i) Which geo-political risks or threats have a maritime dimension or imply derivations whose occurrence may be linked to maritime space? ii) Which processes or tendencies in the use, occupation and governance of maritime space can fall into the category of geo-political risk? The basic aim is to address the forms that instability and geo-political risks take in the ocean world. If the risks stated in the chosen literature are examined from the maritime perspective, it is possible to perceive ‘secondary’ risks whose size and reach can become major contingencies for international stability. They therefore should not be ignored in the prognosis and evaluation of geo-political risks. In as much as societies' political organisation continues to rest on the nation-State, the dominance of the maritime component in the territorial basis is a permanent source of tensions and conflicts. In parallel with this, the displacement of economic expectations and the supply of traditional and new resources to the marine environment broadens the spectrum of risks and threats.
•Geopolitical risks can lead to restrictions on ocean governance.•The emergence of maritime States is a source of tensions and conflicts.•The blue growth boom shifts the creation of geopolitical risks to the oceans.
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Halophilic archaea are extremophiles, adapted to high-salt environments, showing a big biotechnological potential as enzyme, lipids and pigments producers. Four inert supports (perlite, vermiculite, ...polyurethane foam and glass fiber) were employed for solid-state fermentation (SSF) of the halophilic archaeon
Natronococcus
sp. TC6 to investigate biomass and esterase production. A very low esterase activity and high water activity were observed when perlite, vermiculite and polyurethane were used as supports. When glass fiber was employed, an important moisture loss was observed (8.6 %). Moreover, moisture retention was improved by mixing polyurethane and glass fiber, resulting in maximal biomass and esterase production. Three halophilic archaea:
Natronococcus
sp. TC6,
Halobacterium
sp. NRC-1 and
Haloarcula marismortui
were cultured by submerged fermentation (SmF) and by SSF; an improvement of 1.3- to 6.2-fold was observed in the biomass and esterase production when SSF was used. Growth was not homogeneous in the mixture, but was predominant in the glass fiber thus was probably because the glass fiber provides a holder to the cells, while the polyurethane acts as an impregnation medium reservoir. To the best of our knowledge, this work is the first report on haloarchaea cultivation by SSF aiming biomass and esterase/lipase activity production.
Biologically active peptides produced from fish wastes are gaining attention because their health benefits. Proteases produced by halophilic microorganisms are considered as a source of active ...enzymes in high salt systems like fish residues. Hence, the aim of this study was the bioprospection of halophilic microorganisms for the production of proteases to prove their application for peptide production.
Halophilic microorganisms were isolated from saline soils of Mexico and Bolivia. An enzymatic screening was carried out for the detection of lipases, esterases, pHB depolymerases, chitinases, and proteases. Most of the strains were able to produce lipases, esterases, and proteases, and larger hydrolysis halos were detected for protease activity. Halobacillus andaensis was selected to be studied for proteolytic activity production; the microorganism was able to grow on gelatin, yeast extract, skim milk, casein, peptone, fish muscle (Cyprinus carpio), and soy flour as protein sources, and among these sources, fish muscle protein was the best inducer of proteolytic activity, achieving a protease production of 571 U/mL. The extracellular protease was active at 50°C, pH 8, and 1.4 M NaCl and was inhibited by phenylmethylsulfonyl fluoride. The proteolytic activity of H. andaensis was used to hydrolyze fish muscle protein for peptide production. The peptides obtained showed a MW of 5.3 kDa and a radical scavenging ability of 10 to 30% on 2,2-diphenyl-1-picrylhydrazyl and 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) and a ferric reducing ability of plasma.
The use of noncommercial extracellular protease produced by H. andaensis for biologically active peptide production using fish muscle as the protein source presents a great opportunity for high-value peptide production.
How to cite: Delgado-García M, Flores-Gallegos AC, Kirchmayr M, et al. Bioprospection of proteases from Halobacillus andaensis for bioactive peptide production from fish muscle protein. Electron J Biotechnol 2019;39. https://doi.org/10.1016/j.ejbt.2019.03.001.
The effect of β-blockers on infarct size when used in conjunction with primary percutaneous coronary intervention is unknown. We hypothesize that metoprolol reduces infarct size when administered ...early (intravenously before reperfusion).
Patients with Killip class II or less anterior ST-segment-elevation myocardial infarction (STEMI) undergoing percutaneous coronary intervention within 6 hours of symptoms onset were randomized to receive intravenous metoprolol (n=131) or not (control, n=139) before reperfusion. All patients without contraindications received oral metoprolol within 24 hours. The predefined primary end point was infarct size on magnetic resonance imaging performed 5 to 7 days after STEMI. Magnetic resonance imaging was performed in 220 patients (81%). Mean ± SD infarct size by magnetic resonance imaging was smaller after intravenous metoprolol compared with control (25.6 ± 15.3 versus 32.0 ± 22.2 g; adjusted difference, -6.52; 95% confidence interval, -11.39 to -1.78; P=0.012). In patients with pre-percutaneous coronary intervention Thrombolysis in Myocardial Infarction grade 0 to 1 flow, the adjusted treatment difference in infarct size was -8.13 (95% confidence interval, -13.10 to -3.16; P=0.0024). Infarct size estimated by peak and area under the curve creatine kinase release was measured in all study populations and was significantly reduced by intravenous metoprolol. Left ventricular ejection fraction was higher in the intravenous metoprolol group (adjusted difference, 2.67%; 95% confidence interval, 0.09-5.21; P=0.045). The composite of death, malignant ventricular arrhythmia, cardiogenic shock, atrioventricular block, and reinfarction at 24 hours in the intravenous metoprolol and control groups was 7.1% and 12.3%, respectively (P=0.21).
In patients with anterior Killip class II or less ST-segment-elevation myocardial infarction undergoing primary percutaneous coronary intervention, early intravenous metoprolol before reperfusion reduced infarct size and increased left ventricular ejection fraction with no excess of adverse events during the first 24 hours after STEMI.
http://www.clinicaltrials.gov. Unique identifier: NCT01311700. EUDRACT number: 2010-019939-35.
During the process by which the number of actors was increased manifold as a result of the development of civil society, the fisher community has lost prominence and importance, fading into the wide ...spectrum of interests with which it is competing to make its voice heard in decision-making bodies and in the media. This results in what could be termed the participation paradox—the greater the number of actors, the smaller the role each plays, and the lesser the importance of traditional sectors. Participation and devolution do not bear a linear relationship; greater devolution does not necessarily result in greater participation, a claim that has contributed to the processes of devolution being overvalued. Governance, as interaction between State, civil society and the market, paradoxically might not strengthen the most traditional of the interest groups.
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