Evaluation of the operational conditions of the glucose oxidase and catalase multienzymatic system through enzyme co-immobilization on amino hierarchical porous silica.

Hexaric acids have attracted attention lately because they are platform chemicals for synthesizing pharmaceuticals. In particular, gluconic acid is one of the most studied because it is readily available in nature. In this work, operational conditions like temperature and pH were evaluated for the enzymatic production of gluconic acid. For this purpose, glucose oxidase (GOx) and catalase (CAT) were individually immobilized and co-immobilized using amino-silica as support. The catalytic performance of the enzymes both as separate biocatalysts (GOx or CAT) and as an enzymatic complex (GOx-CAT) was assessed in terms of enzymatic activity and stability at temperatures 45 °C and 50 °C and pH 6 to 8. The results show that CAT is a key enzyme for gluconic acid production as it prevents GOx from being inhibited by H2O2. However, CAT was found to be less stable than GOx. Therefore, different GOx to CAT enzymatic ratios were studied, and a ratio of 1-3 was determined to be the best. The highest glucose conversion conditions were 45 °C and pH 7.0 for 24 h. Regarding the biocatalyst reuse, GOx-CAT retained more than 70% of its activity after 6 reaction cycles. These results contribute to further knowledge and application of oxidases for hexaric acid production and shed greater light on the role of the glucose oxidase/catalase pair in better catalytic performance. Both enzymes were immobilized in one pot, which is relevant for their potential use in industry; an enzyme system was obtained in a single step.

Atypical Hemolytic Uremic Syndrome: A Nationwide Colombian Pediatric Series.

Objectives. Atypical hemolytic uremic syndrome (aHUS) is a rare complement-mediated kidney disease with genetic predisposition and represents up to 10% of pediatric hemolytic uremic syndrome (HUS) cases. Few studies have evaluated aHUS in Latin American population. We studied a Colombian pediatric cohort to delineate disease presentation and outcomes. Methods. A multicenter cohort of 27 Colombian children with aHUS were included. Patients were grouped by age at onset. Clinical features were compared using analysis of variance (ANOVA) and Fisher exact tests. Renal biopsy was performed on 6 patients who were suspected of having other renal diseases before aHUS diagnosis. Results. Most patients were male (70%). The onset of aHUS occurred frequently before age 4 years (60%) and followed gastroenteritis as the main triggering event (52%). Age groups showed comparable clinical presentation, disease severity, treatment, and outcomes. Pulmonary involvement (67%) was the main extrarenal manifestation, particularly in the 1 to 7 age group (P = .01). Renal biopsies were as follows: 3 had membranoproliferative glomerulonephritis (MPGN) type I, one MPGN type III, one C3-glomerulonephritis, and one rapidly progressive GN. Genetic screening was available in 6 patients and identified 2xCFHR5, 2xMCP, 1xADAMTS13/THBD, and 1xDGKE mutations. A total of 15 relapses were seen, of which 8 (72%) occurred in the 1 to 7 age group. The renal outcome was not significantly different regardless of age group. Conclusion. In our cohort, we observed a relatively high frequency of extrarenal involvement at first presentation represented by pulmonary manifestations. The renal prognosis at initial presentation was worse than in previous reports.

Gastrointestinal Foreign Bodies: 14 Years of Experience in a University Clinic

Introduction: Ingesting foreign bodies is a common medical problem, especially in the emergency department. Some small studies describe experiences in this regard. Materials and methods: A descriptive retrospective study included patients with suspected ingestion of foreign bodies admitted to the gastroenterology and GI endoscopy service of the Clínica Universitaria Colombia between January 2007 and August 2020. Results: The age of occurrence of the event was 18 to 95 years, and the average age was 45 years. The foreign bodies ingested and found were variable. The most frequent was fish bones, representing 64.11% of the cases, followed by chicken bones and dietary impaction. Thirty-eight percent of patients required foreign body removal; the most frequently used tool was the foreign body forceps. The primary location was the esophagus in 12.53% of cases, followed by the cricopharynx in 11.18% and the hypopharynx in 10%. Conclusions: The Clínica Universitaria Colombia is a referral site for many gastroenterology emergencies due to its high technological level and extensive human resources. This paper probably describes the largest number of patients with this reason for consultation, which is why this retrospective descriptive study was designed. It shows the demographic characteristics, foreign body types, radiological and endoscopic findings, and associated complications, which help to provide a more accurate knowledge of this pathology.

Introducción: La ingesta de cuerpos extraños es un problema médico frecuente, especialmente en el servicio de urgencias. Existen algunos estudios pequeños que describen las experiencias al respecto. Materiales y métodos: Estudio descriptivo, retrospectivo, en el cual se incluyó a pacientes con sospecha de ingesta de cuerpos extraños, ingresados al servicio de gastroenterología y endoscopia digestiva de La Clínica Universitaria Colombia, entre enero de 2007 y agosto de 2020. Resultados: La edad de ocurrencia del evento se presentó en pacientes desde los 18 hasta los 95 años, y la edad promedio fue de 45 años. Los cuerpos extraños ingeridos y encontrados fueron variables; los más frecuentes fueron la ingesta de espinas de pescado, que representó el 64,11% de los casos, seguido por la ingesta de huesos de pollo y la impactación alimentaria. Un 38% de los pacientes requirieron la extracción de cuerpo extraño y la herramienta usada con mayor frecuencia fue la pinza de cuerpo extraño. La localización principal fue el esófago, en el 12,53% de los casos, seguido por la cricofaringe, en el 11,18%, y la hipofaringe, en el 10%. Conclusiones: La Clínica Universitaria Colombia es un sitio de referencia de una gran cantidad de urgencias en gastroenterología debido a su alto nivel tecnológico y al gran recurso humano que requieren. Este trabajo representa probablemente la cantidad más grande de pacientes con este motivo de consulta, razón por la que se diseñó este estudio descriptivo retrospectivo, que muestra las características demográficas, los tipos de cuerpo extraño, los hallazgos radiológicos y endoscópicos y las complicaciones asociadas, que son de utilidad para tener un conocimiento más real de esta patología.

Modulating Substrate Specificity of Rhizobium sp. Histamine Dehydrogenase through Protein Engineering for Food Quality Applications.

Histamine is a biogenic amine found in fish-derived and fermented food products with physiological relevance since its concentration is proportional to food spoilage and health risk for sensitive consumers. There are various analytical methods for histamine quantification from food samples; however, a simple and quick enzymatic detection and quantification method is highly desirable. Histamine dehydrogenase (HDH) is a candidate for enzymatic histamine detection; however, other biogenic amines can change its activity or produce false positive results with an observed substrate inhibition at higher concentrations. In this work, we studied the effect of site saturation mutagenesis in Rhizobium sp. Histamine Dehydrogenase (Rsp HDH) in nine amino acid positions selected through structural alignment analysis, substrate docking, and proximity to the proposed histamine-binding site. The resulting libraries were screened for histamine and agmatine activity. Variants from two libraries (positions 72 and 110) showed improved histamine/agmatine activity ratio, decreased substrate inhibition, and maintained thermal resistance. In addition, activity characterization of the identified Phe72Thr and Asn110Val HDH variants showed a clear substrate inhibition curve for histamine and modified kinetic parameters. The observed maximum velocity (Vmax) increased for variant Phe72Thr at the cost of an increased value for the Michaelis-Menten constant (Km) for histamine. The increased Km value, decreased substrate inhibition, and biogenic amine interference observed for variant Phe72Thr support a tradeoff between substrate affinity and substrate inhibition in the catalytic mechanism of HDHs. Considering this tradeoff for future enzyme engineering of HDH could lead to breakthroughs in performance increases and understanding of this enzyme class.

Immobilized biocatalyst engineering: Biocatalytic tool to obtain attractive enzymes for industry.

Biocatalysis can improve current bioprocesses by identifying or improving enzymes that withstand harsh and unnatural operating conditions. Immobilized Biocatalyst Engineering (IBE) is a novel strategy integrating protein engineering and enzyme immobilization as a single workflow. Using IBE, it is possible to obtain immobilized biocatalysts whose soluble performance would not be selected. In this work, Bacillus subtilis lipase A (BSLA) variants obtained through IBE were characterized as soluble and immobilized biocatalysts, and how the interactions with the support affect their structure and catalytic performance were analyzed using intrinsic protein fluorescence. Variant P5G3 (Asn89Asp, Gln121Arg) showed a 2.6-fold increased residual activity after incubation at 76 °C compared to immobilized wild-type (wt) BSLA. On the other hand, variant P6C2 (Val149Ile) showed 4.4 times higher activity after incubation in 75 % isopropyl alcohol (36 °C) compared to Wt_BSLA. Furthermore, we studied the advancement of the IBE platform by performing synthesis and immobilizing the BSLA variants using a cell-free protein synthesis (CFPS) approach. The observed differences in immobilization performance, high temperature, and solvent resistance between the in vivo-produced variants and Wt_BSLA were confirmed for the in vitro synthesized enzymes. These results open the door for designing strategies integrating IBE and CFPS to generate and screen improved immobilized enzymes from genetic diversity libraries. Furthermore, it was confirmed that IBE is a platform that can be used to obtain improved biocatalysts, especially those with an unremarkable performance as soluble biocatalysts, which wouldn't be selected for immobilization and further development for specific applications.

Dengue en Barbosa (Santander, Colombia): conocimientos, actitudes y prácticas en la comunidad / Dengue in Barbosa (Santander, Colombia): knowledge, attitudes, and practices in the community

Introducción: El dengue es considerado un problema de la salud pública con un comportamiento endémico-epidémico en Colombia, por lo cual los programas de vigilancia epidemiológica se enfocan en la prevención y control. Objetivo: Evaluar la comprensión y el comportamiento de la población del municipio de Barbosa (Santander, Colombia) acerca del virus del dengue y el vector por medio de un instrumento de conocimientos, actitudes y prácticas (CAP). Métodos: Se realizó un estudio transversal analítico en el que participaron 93 habitantes del municipio de Barbosa, seleccionados por un muestreo no probabilístico por conveniencia. Se empleó un instrumento de CAP validado. La información fue sistematizada y analizada en el programa SPSS v. 26.0. Resultados: El 78,5 por cientode los encuestados mostraron conocimientos deficientes con un puntaje obtenido inferior a 70 puntos. El 73,1 por ciento de los habitantes presentó actitudes negativas con un resultado igual o inferior a 24 puntos. El 67,7 por ciento de la población presentó buenas prácticas, con un resultado mayor o igual a 14 puntos. Se evidencia que, a menor edad, son mayores las actitudes desfavorables frente a la infección por dengue (OR 2,67 IC 95 por ciento 1,03-7,1). Conclusiones: Educar a la población de manera más efectiva acerca de los conocimientos básicos sobre el dengue sigue siendo una estrategia necesaria, además de crear conciencia de que las prácticas en el hogar se deben ejecutar con la finalidad de eliminar el vector y no solo como mecanismo de limpieza. Se deben implementar diferentes estrategias comunicativas para educar a los ciudadanos en este sentido(AU)

Introduction: Dengue is considered a public health problem with endemic-epidemic behavior in Colombia; therefore, epidemiological surveillance programs are aimed at its prevention and control. Objective: To evaluate the understanding and behavior of the population from the municipality of Barbosa (Santander, Colombia) on dengue virus and its vector through a knowledge, attitudes, and practices (KAP) survey. Methods: An analytical cross-sectional study was conducted which included 93 residents of the municipality of Barbosa, selected based on non-probabilistic and convenience sampling A validated KAP instrument was used. The information was systematized and analyzed in SPSS v. 26.0. The information was systematized and analyzed in SPSS v. 26.0. Results: Among the respondents, 78.5 percent showed deficient knowledge with a score below 70 points. Negative attitudes were found in 73.1 percent of the residents with a score equal to or lower than 24 points. Best practices were reported by 67.7 percent of the population, with a score greater than or equal to 14 points. It is evident that at younger ages, unfavorable attitudes towards dengue infection are higher (OR 2.67 CI 95% 1.03-7.1). Conclusions: Educating the population more effectively about basic knowledge of dengue remains a necessary strategy, in addition to raising awareness that household practices should be carried out to eliminate the vector and not only as a cleaning mechanism. Different communicative strategies should be implemented to educate citizens in this regard(AU)

Enzymatic Synthesis of Ascorbyl Palmitate in a Rotating Bed Reactor.

Ascorbyl palmitate, an ascorbic acid ester, is an important amphipathic antioxidant that has several applications in foods, pharmaceuticals, and cosmetics. The enzymatic synthesis of ascorbyl palmitate is very attractive, but few efforts have been made to address its process scale-up and implementation. This study aimed at evaluating the enzymatic synthesis of ascorbyl palmitate in a rotating basket reactor operated in sequential batches. Different commercial immobilized lipases were tested, and the most suitable reaction conditions were established. Among those lipases studied were Amano Lipase PS, Lipozyme® TL IM, Lipozyme® Novo 40086, Lipozyme® RM IM and Lipozyme® 435. Initially, the enzymes were screened based on previously defined synthesis conditions, showing clear differences in behavior. Lipozyme® 435 proved to be the best catalyst, reaching the highest values of initial reaction rate and yield. Therefore, it was selected for the following studies. Among the solvents assayed, 2-methyl-2-butanol and acetone showed the highest yields, but the operational stability of the catalyst was better in 2-methyl-2-butanol. The tests in a basket reactor showed great potential for large-scale application. Yields remained over 80% after four sequential batches, and the basket allowed for easy catalyst recycling. The results obtained in basket reactor are certainly a contribution to the enzymatic synthesis of ascorbyl palmitate as a competitive alternative to chemical synthesis. This may inspire future cost-effectiveness studies of the process to assess its potential as a viable alternative to be implemented.

Replicating endothelial shear stress in organ-on-a-chip for predictive hypericin photodynamic efficiency.

Photodynamic therapy using Hypericin (Hy-PDT) is an alternative non-invasive treatment that enables selective tumor inhibition and angiogenesis derived from the differential recruitment of endothelial cells in the tumor microenvironment. Most PDT studies were performed on in vitro models without vascular biomechanical simulation. Our work strives to develop a microchip that generates a constant shear stress force to investigate the Hy-PDT efficiency on human umbilical vein endothelial cells (HUVECs). The microchip with a single straight microchannel was composed of the bottom layer (polystyrene), the middle layer (double-sided biocompatible adhesive tape), and the top layer (polyester film) and could produce shear stress in the range of 1.4 - 7.0 dyn cm-2. The quantification of vascular endothelial growth factor (VEGF), cell viability, and activities of caspases 3 and 7 were assayed to validate the microchip and Hy-PDT efficacy. After the endothelization, static and dynamic cell incubations with Hy were conducted in microchips. Compared to static systems, the shear stress displayed its effect on the increasing release of VEGF and promoted more cell damage and cell death via necrosis during Hy-PDT. In conclusion, the expressive shear stress-dependent manner during PDT treatments suggests that the microchip could be an essential approach in preclinical tests to evaluate the therapeutic outcome considering the endothelial shear stress microenvironment.

Biomarcadores moleculares del género salmonella aislada en alimentos

Introducción: El género Salmonella está conformado por patógenos alimentarios causantes de salmonelosis, una enfermedad de distribución cosmopolita, que afecta a todos los grupos poblacionales, tanto en los países desarrollados como en los que están en vía de desarrollo, siendo un problema de salud pública. Con relación a esto, los biomarcadores moleculares se convierten en una herramienta útil para su diagnóstico y seguimiento. Objetivo: Describir los marcadores moleculares del género Salmonella y las diferentes técnicas moleculares empleadas para su detección en la actualidad. Metodología: Se realizó una búsqueda bibliográfica en las bases de datos Science direct, PubMed, Proquest y Ovid, utilizando 5 palabras clave, las cuales se combinaron de diferentes maneras para finalmente obtener 50 referencias bibliográficas. Resultados: Se evidencia gran variedad de biomarcadores del género Salmonella, los cuales son detectados por pruebas moleculares, como la técnica de PCR, que permite la detección directa de genes que codifican para los antígenos somáticos O y flagelares H de Salmonella y la determinación de los posibles serovares implicados en las enfermedades transmitidas por alimentos, de una manera rápida y precisa. Cabe resaltar las ventajas de los métodos moleculares frente a la serotipificación en cuanto al costo-beneficio, estandarización óptima, alta sensibilidad, especificidad y rapidez en la obtención de los resultados. Conclusiones: Los biomarcadores moleculares se consideran una herramienta de referencia para la identificación de serovares de Salmonella y su serotipificación, a través de la PCR, con un alto nivel de especificidad y sensibilidad en los resultados, aportando información relevante a nivel epidemiológico.

Introduction: The Salmonella genus is composed of food pathogens that cause salmonellosis, a disease of cosmopolitan distribution that affects all population groups, both in developed and developing countries, being a public health problem. In this regard, molecular biomarkers become a useful tool for its diagnosis and monitoring. Objective: Describe which are the molecular markers of the Salmonella genus and their determination through different molecular techniques currently used. Methodology: A bibliographic search was performed in databases such as Pubmed, Science direct, Elsevier, NCBI, Proquest and Ovid, using 5 keywords, which were combined in different ways to finally obtain 50 bibliographic references from the year 2005 to currently, selecting articles in English and Spanish. Results: A great variety of biomarkers of the Salmonella genus are evident, which are detected by molecular tests such as the PCR technique, this allows the direct detection of the genes that encode the somatic O and flagellar H antigens of Salmonella and quickly allows the determination of the possible serovars involved in food-borne diseases, in a fast and precise way, in addition to highlighting the advantages of the described molecular methods compared to serotyping in terms of cost-benefit, optimal standardization, high sensitivity, specificity and speed in the results. Conclusions: Molecular biomarkers are considered a reference tool for the identification of Salmonella serovars and their serotyping, through molecular techniques such as PCR, with a high level of specificity and sensitivity in the results, which are of epidemiological importance.

IFNL4 genotype influences the rate of HIV-1 seroconversion in men who have sex with men.

Individuals lacking interferon lambda 4 (IFNL4) protein due to a common null mutation (rs368234815) in the IFNL4 gene display higher resistance against several infections. The influence of IFNL4 on HIV-1 infection is still under discussion and conflicting results have been reported. This study intended to corroborate or refute the association of the null allele of IFNL4 and HIV-1 predisposition in a cohort of men who have sex with men (MSM). IFNL4 null genotype was assessed on 619 HIV-1-seronegative MSM who were followed for 36 months during a trial of a prophylactic vaccine against HIV-1. Of those, 257 individuals seroconverted during this period. A logistic regression model was constructed including demographic and IFNL4 genotype. In addition, a meta-analysis using data from the current study and other European populations was conducted. The null IFNL4 genotypes were correlated with lower HIV-1 seroconversion (Adjusted OR = 0.4 [95%CI: 0.2-0.8], P = 0.008) and longer time to seroconversion (889 vs. 938 days, P= 0.01). These results were validated by a meta-analysis incorporating data from other European populations and the result yielded a significant association of the IFNL4 null genotype under a dominant model with a lower probability of HIV-1 infection (OR=0.4 [95% CI: 0.3-0.6]; P= 1.3 x 10E-5).

Effect of process parameters and surfactant additives on the obtained activity of recombinant tryptophan hydroxylase (TPH1) for enzymatic synthesis of 5-hydroxytryptophan (5-HTP).

5-hydroxytryptophan (5-HTP) is an intermediate molecule in the biosynthesis of serotonin, an important neurotransmitter, regulating a series of metabolic and psychological functions in humans. In this work, we studied the heterologous production of Human tryptophan hydroxylase (TPH1) in Escherichia coli, for the synthesis of 5-hydroxytryptophan (5-HTP) from Tryptophan (Trp). To quantify TPH1 activity, a simple fluorescence-based microtiter plate assay was established, based on the changes in fluorescence emission at 340 nm between substrate and product when excited at 310 nm, allowing quick and reliable quantification of released 5-HTP. To increase enzyme production, heterologous TPH1 production was studied in stirred tank bioreactor scale. The effect of rate of aeration (0.25, 0.50 and 0.75 vvm) and agitation (150, 250 and 500 rpm) was evaluated for biomass production, pH, volumetric oxygen transfer coefficient (kLa) and volumetric TPH1 activity. We determined that high agitation and low aeration allowed reaching the maximum measured enzyme activity. Under such conditions, we observed a 90% substrate conversion, obtaining 90 µM (~0.02 g/L) 5-HTP from a 100 µM Tryptophan substrate solution. Finally, we observed that the addition of Tween 20 (0.1%) in the culture broth under production conditions expanded the pH operation range of TPH1. Our results establish a base for a biocatalytic approach as a potential alternative process for the synthesis of 5-HTP using recombinant TPH1.

Biosynthesis of a Novel Antibacterial Dipeptide, Using Proteases From South American Native Fruits, Useful as a Food Preservative.

Antiacanthain and granulosain are the partially purified proteolytic extracts from the South American native fruits of Bromelia antiacantha (Bertol. ) and Solanum granuloso leprosum, respectively. The aim of this work was to compare the ability of both soluble and immobilized antiacanthain and granulosain f or the synthesis of Z-Tyr-Val-OH, a novel antibacterial dipeptide, in different reaction systems formed by almost anhydrous organic solvents (Xw: 1 × 10-5) and several percentages of immiscible organic solvents in 100 mM Tris(hydroxymethyl)aminomethane hydrochloride buffer pH 8.0. Soluble antiacanthain in half of the 24 different organic biphasic media showed higher catalytic potential than in 100 mM Tris(hydroxymethyl)aminomethane hydrolchloride buffer pH 8.0. Soluble granulosain showed lower catalytic potential in all liquid-liquid biphasic media than in the same buffer. However, 50% (v/v) ethyl ethanoate in 100 mM Tris(hydroxymethyl)aminomethane hydrolchloride buffer pH 8.0 allowed to express the highest catalytic potential of both soluble enzymes. In 50% v/v ethyl ethanoate, soluble antiacanthain and granulosain catalyzed the synthesis of Z-Tyr-Val-OH with 72 ± 0.15 and 60 ± 0.10% maximal peptide yields, respectively. Multi-point immobilization in glyoxyl-silica did not lead to better peptide yields than soluble enzymes, in that liquid-liquid biphasic medium under the same reaction conditions. Soluble and glyoxyl-silica immobilized antiacanthain in almost anhydrous ethyl ethanoate (Xw: 1 × 10-5) were able to retain 17.3 and 45% of the initial proteolytic activity of antiacanthain in 100 mM Tris hydrolchloride buffer pH 8.0, respectively, at 40°C under agitation (200 rpm). Soluble and glyoxyl-silica immobilized granulosain were inactivated under the same reaction conditions. Glyoxyl-silica immobilized antiacanthain showed to be a robust biocatalyst in almost anhydrous ethyl ethanoate (Xw: 1 × 10-5), eliciting the best peptide yield (75 ± 0.13%). The synthesis reaction of Z-Tyr-Val-OH could not proceed when soluble antiacanthain was used under the same conditions. Both peptidases only catalyzed the synthesis reaction under kinetic control, using activated acyl donor substrates. Finally, this work reports a novel broad-spectrum antibacterial peptide that significantly decreased (p ≤ 0.05) the specific growth rates of Gram positive and Gram negative microorganisms at very low concentrations (≥15 and 35 µg/ml, respectively); contributing with a new safe food preservative of applying for different food systems.

Immobilized Biocatalyst Engineering: High throughput enzyme immobilization for the integration of biocatalyst improvement strategies.

The increasing use of sustainable manufacturing technologies in the industry presents a constant challenge for the development of suitable biocatalysts. Traditionally, improved biocatalysts are developed either using protein engineering (PE) or enzyme immobilization (EI). However, these approaches are usually not simultaneously applied. In this work, we designed and validated an enzyme improvement platform, Immobilized Biocatalyst Engineering (IBE), which simultaneously integrates PE and EI, with a unique combination of improvement through amino acid substitutions and attachment to a support material, allowing to select variants that would not be found through single or subsequent PE and EI improvement strategies. Our results show that there is a significant difference on the best performing variants identified through IBE, when compared to those that could be identified as soluble enzymes and then immobilized, especially when evaluating variants with low enzyme as soluble enzymes and high activity when immobilized. IBE allows evaluating thousands of variants in a short time through an integrated screening, and selection can be made with more information, resulting in the detection of highly stable and active heterogeneous biocatalysts. This novel approach can translate into a higher probability of finding suitable biocatalysts for highly demanding processes.

Enzymatic synthesis of 4-methoxycinnamoylglycerol Uv filter mediated by immobilized-lipase and nanoparticle formation on N-succinylchitosan / Síntesis enzimática del filtro UV 4-metoxicinamoilglicerol mediado por lipasa inmovilizada y formación de nanopartículas N-succinilquitosano

ABSTRACT The synthesis of 4-methoxycinnamoylglycerol takes advantage of the biodiesel subproduct for obtaining a hydrophilic UV cinnamate derivate filter, useful in sunscreen formulations. The objective here was to demonstrate that esterification of 4-methoxycinnamic acid and glycerol mediated by immobilized-lipase from Thermomyces lanuginosus is selective towards 4-methoxycinnamoylglycerol monoester UV filter, whose chemical characteristics favor the nanoparticles formation by ionotropic gelation on N-Succinyl chitosan. A cinnamic acid conversion (34% in hexane is higher than in other reports, without the presence of other sub-products or degradation products. This eases the purification process by liquid-liquid extraction. The free glyceryl entities favour its incorporation on N-Succinyl chitosan nanoparticles with size around 185(77nm, which are promissory for sunblock products.

RESUMEN En la síntesis de 4-metoxicinamoilglicerol, se aprovecha el subproducto de biodiesel para obtener un filtro UV hidrofílico, derivado de cinamato, útil en formulaciones de bloqueadores solares. El objetivo de este trabajo fue demostrar que la esterificación del ácido 4-metoxicinámico y el glicerol, mediado por la lipasa inmovilizada de Thermomyces lanuginosus, es selectiva hacia el monoester del filtro UV 4-metoxicinamoilglicerol, cuyas características químicas favorecen la formación de nanopartículas, por gelificación ionotrópica en N-succinil-quitosano. Una conversión de ácido cinámico (34% en hexano es mayor que los valores ya reportados, sin la presencia de otros subproductos o productos de degradación. Esto facilita, el proceso de purificación por extracción líquido-líquido. Las entidades de glicerilo libre favorecen su incorporación en nanopartículas de N-succinil-quitosano, con un tamaño de alrededor de 185±77nm, que son promisorias para los productos de protección solar.

Complement C4 Gene Copy Number Variation Genotyping by High Resolution Melting PCR.

BACKGROUND: Complement C4 gene copy number variation plays an important role as a determinant of genetic susceptibility to common diseases, such as systemic lupus erythematosus, schizophrenia, rheumatoid arthritis, and infectious diseases. This study aimed to develop an assay for the quantification of copy number variations in the C4 locus. METHODS: the assay was based on a gene ratio analysis copy enumeration (GRACE) PCR combined with high resolution melting (HRM) PCR. The test was optimized using samples of a known genotype and validated with 72 DNA samples from healthy blood donors. RESULTS: to validate the assay, standard curves were generated by plotting the C4/RP1 ratio values against copy number variation (CNV) for each gene, using genomic DNA with known C4 CNV. The range of copy numbers in control individuals was comparable to distributions observed in previous studies of European descent. CONCLUSIONS: the method herein described significantly simplifies C4 CNV diagnosis to validate the assay.

Development of alginate-pectin microcapsules by the extrusion for encapsulation and controlled release of polyphenols from papaya (Carica papaya L.).

The papaya fruit (Carica papaya L.) contains a wide variety of bioactive compounds with potential applications in the food and nutraceutical industries. The entrapment and release of such bioactive compounds remain a critical step for the development of functional, stable, and cost-effective storage and delivery systems, since the interaction of polymers on capsules and the payload molecules can influence the performance of the capsule system under operational conditions. The present study describes the encapsulation of rutin and trans-ferulic acid-rich extracts from papaya exocarp in a pectin-alginate composite, evaluating the performance of gallic acid encapsulation obtained through in situ and two-step entrapment methods. The best alginate:pectin ratio for gallic acid encapsulation was 55:45 and 61:39, achieving 6.1 mg and 28.1 mg GAE/g capsules when the papaya exocarp extract was encapsulated by in situ and two-step, respectively. We also evaluated the payload release performance of the obtained capsules under in vitro conditions simulating gastrointestinal conditions. Our results indicate an increased protective effect at gastric pH and targeted release of polyphenols when in situ encapsulation is used to encapsulate the extracts. PRACTICAL APPLICATIONS: Currently, adding value to agroindustry processing waste is an important focus to achieve a more economically and environmentally sustainable food industry. The recovery of bioactive molecules such as polyphenols, for food supplements or formulation additives in the form of by-product extracts is gaining importance as novel sustainable processes in the agricultural industry. Thus, the encapsulation of such bioactive extracts for storage and consumption is an active research field, aiming to overcome the low storage stability and lability to gastric conditions, currently hindering their applications in food or pharmaceutical formulations. In this sense, capsule design and the development of efficient encapsulation methods are very important to obtain a suitable carrier and protector system for the capsulated bioactive extracts or molecules. This research aims to add value to papaya waste and potentially to other agroindustry wastes such as pectin and alginate, resulting in a polyphenol carrier with excellent encapsulation and targeted release properties under gastrointestinal conditions. In conclusion, this kind of works could allow to the application of the agroindustry byproducts to obtain high added-value products, in the form of polyphenol-loaded capsules.

Occupational health in the framework of the COVID-19 pandemic: a scoping review / Seguridad y salud en el trabajo en el marco de la pandemia por COVID-19: una revisión exploratoria

ABSTRACT Objective To collect the available evidence related to occupational health in the face of the introduction of the new SARS-CoV-2 coronavirus pandemic. Methods Scoping review developed from the Arksey and O'Malley framework. The search was performed in the databases PubMed, Academic Search Complete, Science Direct, Medline, Scopus, Web of Science and Google Scholar. Documents on COVID-19 and its relationship with occupational health published in English, Portuguese and Spanish were included. The review, selection and characterization of the studies was carried out by five reviewers. Results The search and selection identified 43 documents published between December 2019 and April 2020. The topics covered include occupational exposure, protection measures, psychosocial affectations of workers, particularly health, as well as conditions of work organization that can influence contagion. Conclusions Health workers are the most exposed workforce. Accompaniment, coaching and training in relation to patient care and the use of personal protection equipment are essential to reduce contagion among health personnel. In other work activities, social distancing is the standard measure for the mitigation of transmission, as well as the continuous disinfection of workplaces.(AU)

RESUMEN Objetivo Recopilar la evidencia disponible relacionada con la salud laboral frente a la introducción de la nueva pandemia por el coronavirus SARS-CoV-2. Métodos Revisión exploratoria desarrollada a partir del marco de Arksey y O'Malley. La búsqueda se realizó en las bases de datos PubMed, Academic Search Complete, Science Direct, Medline, Scopus, Web of Science y Google Scholar. Se incluyeron documentos sobre COVID-19 y su relación con la salud ocupacional publicados en inglés, portugués y español. La revisión, selección y caracterización de los estudios fue desarrollada por cinco revisores. Resultados La búsqueda y selección identificó 43 documentos publicados entre diciembre de 2019 y abril de 2020. Las temáticas abordadas incluyen exposición ocupacional, medidas de protección, afectaciones psicosociales de los trabajadores, particularmente de la salud, así como condiciones de la organización del trabajo que pueden influir en el contagio. Conclusiones Los trabajadores de la salud son la fuerza laboral más expuesta. El acompañamiento, entrenamiento y la capacitación en relación con la atención de pacientes y el uso de elementos de protección personal son fundamentales para disminuir el contagio entre personal de salud. En otras actividades laborales el distanciamiento social es la medida estándar para la mitigación de la transmisión, así como la desinfección continua de los lugares de trabajo.(AU)

Deletion and Randomization of Structurally Variable Regions in B. subtilis Lipase A (BSLA) Alter Its Stability and Hydrolytic Performance Against Long Chain Fatty Acid Esters.

The continuous search for novel enzyme backbones and the engineering of already well studied enzymes for biotechnological applications has become an increasing challenge, especially by the increasing potential diversity space provided by directed enzyme evolution approaches and the demands of experimental data generated by rational design of enzymes. In this work, we propose a semi-rational mutational strategy focused on introducing diversity in structurally variable regions in enzymes. The identified sequences are subjected to a progressive deletion of two amino acids and the joining residues are subjected to saturation mutagenesis using NNK degenerate codons. This strategy offers a novel library diversity approach while simultaneously decreasing enzyme size in the variable regions. In this way, we intend to identify and reduce variable regions found in enzymes, probably resulting from neutral drift evolution, and simultaneously studying the functional effect of said regions. This strategy was applied to Bacillus. subtilis lipase A (BSLA), by selecting and deleting six variable enzyme regions (named regions 1 to 6) by the deletion of two amino acids and additionally randomizing the joining amino acid residues. After screening, no active variants were found in libraries 1% and 4%, 15% active variants were found in libraries 2% and 3%, and 25% for libraries 5 and 6 (n = 3000 per library, activity detected using tributyrin agar plates). Active variants were assessed for activity in microtiter plate assay (pNP-butyrate), thermal stability, substrate preference (pNP-butyrate, -palmitate), and compared to wildtype BSLA. From these analyses, variant P5F3 (F41L-ΔW42-ΔD43-K44P), from library 3 was identified, showing increased activity towards longer chain p-nitrophenyl fatty acid esters, when compared to BSLA. This study allowed to propose the targeted region 3 (positions 40-46) as a potential modulator for substrate specificity (fatty acid chain length) in BSLA, which can be further studied to increase its substrate spectrum and selectivity. Additionally, this variant showed a decreased thermal resistance but interestingly, higher isopropanol and Triton X-100 resistance. This deletion-randomization strategy could help to expand and explore sequence diversity, even in already well studied and characterized enzyme backbones such as BSLA. In addition, this strategy can contribute to investigate and identify important non-conserved regions in classic and novel enzymes, as well as generating novel biocatalysts with increased performance in specific processes, such as enzyme immobilization.

Occupational health in the framework of the COVID-19 pandemic: a scoping review.

OBJECTIVE: To collect the available evidence related to occupational health in the face of the introduction of the new SARS-CoV-2 coronavirus pandemic. METHODS: Scoping review developed from the Arksey and O'Malley framework. The search was performed in the databases PubMed, Academic Search Complete, Science Direct, Medline, Scopus, Web of Science and Google Scholar. Documents on COVID-19 and its relationship with occupational health published in English, Portuguese and Spanish were included. The review, selection and characterization of the studies was carried out by five reviewers. RESULTS: The search and selection identified 43 documents published between December 2019 and April 2020. The topics covered include occupational exposure, protection measures, psychosocial affectations of workers, particularly health, as well as conditions of work organization that can influence contagion. CONCLUSIONS: Health workers are the most exposed workforce. Accompaniment, coaching and training in relation to patient care and the use of personal protection equipment are essential to reduce contagion among health personnel. In other work activities, social distancing is the standard measure for the mitigation of transmission, as well as the continuous disinfection of workplaces.

Synthesis with Immobilized Lipases and Downstream Processing of Ascorbyl Palmitate.

Ascorbyl palmitate is a fatty acid ester endowed with antioxidant properties, used as a food additive and cosmetic ingredient, which is presently produced by chemical synthesis. Ascorbyl palmitate was synthesized from ascorbic acid and palmitic acid with a Pseudomonas stutzeri lipase immobilized on octyl silica, and also with the commercial immobilized lipase Novozym 435. The latter was selected for optimizing the reaction conditions because of its high reactivity and stability in the solvent 2-methyl-2-butanol used as reaction medium. The reaction of the synthesis was studied considering temperature and molar ratio of substrates as variables and synthesis yield as response parameter. The highest yield in the synthesis of ascorbyl palmitate was 81%, obtained at 55 °C and an ascorbic acid to palmitic acid molar ratio of 1:8, both variables having a strong effect on yield. The synthesized ascorbyl palmitate was purified to 94.4%, with a purification yield of 84.2%. The use of generally recognized as safe (GRAS) certified solvents with a polarity suitable for the solubilization of the compounds made the process a viable alternative for the synthesis and downstream processing of ascorbyl palmitate.