Diseño, arranque y caracterización de un biorreactor de charolas para fermentación en medio sólido

En años recientes ha existido un gran interés en los procesos de fermentación en medio sólido (FMS) por los altos rendimientos que se han obtenido en la producción de metabolitos de alto valor agregado de interés industrial, por lo que se han llevado a cabo investigaciones en el diseño de biorreactores en busca de que sean aplicados a nivel industrial. En este trabajo se diseñó y construyó un biorreactor de charolas a escala de banco y se caracterizaron los distintos mecanismos de transporte y reacción en el sistema mediante métodos experimentales y a través de la dinámica de fluidos computacionales empleando el software COMSOL® Multiphysics. Se construyó un modelo en frío del biorreactor de charolas en acrílico de ½ pulgada de espesor considerándose la adaptación de sensores de gas O2, gas CO2, temperatura y humedad relativa. Se eligió como material de sellado una junta tipo o-ring de neopreno de geometría circular (4.7 mm diámetro) y densidad de 1.45 g/cm3 para sellar bien la unión de los módulos que componen el biorreactor. La charola se construyó con malla (# 40) de acero inoxidable y el difusor de aire tipo serpentín se construyó con tubo de acero inoxidable de ¼ pulgada de diámetro. El medio poroso de empaque del biorreactor consistió en una mezcla de subproductos de frutas y verduras y pasta de soya con un tamaño de partícula: 1.68 > TP > 1.41 mm. Se evaluó el transporte de masa por dispersión por el método de inyección de un pulso o trazador y se calculó el coeficiente de dispersión (D/uL) que caracteriza la dispersión en todo el sistema mediante el análisis de las curvas de distribución de tiempos de residencia (DTR) en donde se obtuvieron valores de (D/uL) de 0.123, 0.196 y 0.220 para los flujos de 200, 300 y 400 mL/min respectivamente. Se evaluó el transporte de calor en el lecho empacado de los mecanismos conductivos y convectivos en la determinaron del perfil radial y axial de temperatura respectivamen a diferentes flujos de aireación con una temperatura del aire de 20 °C observándose gradientes radiales de 0.5, 0.58 y 0.63 °C/cm y gradientes axiales de 2.5, 2.9 y 3.5 °C/cm para flujos de 200, 300 y 400 mL/min respectivamente. Se evaluó la producción de proteasas por Y. lipolytica en fermentación en medio sólido en el biorreactor de charolas, obteniéndose la mayor actividad (40.1 U/gMS) a las 36 h de cultivo utilizando como sustrato subproductos de frutas y verduras-pasta de soya. Posteriormente se realizó un balance estequiométrico a partir de los datos de producción de CO2 obtenidos mediante el monitoreo en línea con el sensor de gas CO2 durante la fermentación. Así mismo, mediante un ajuste de datos de CO2 con el modelo logístico (R2 > 0.99) se determinaron los parámetros cinéticos aparentes: tasa máxima de producción de CO2 (2 = 11.83 2⁄ℎ), tasa específica de producción de CO2 (2 = 0.53 ℎ −1 ), producción total de CO2 (2 154.83 2⁄) y el tiempo de fase “Lag” con un valor de 6.83 h. Finalmente se modeló el biorreactor utilizando la dinámica de fluidos computacionales en el software COMSOL® Multiphysics 4.4. El modelado del biorreactor se realizó considerando un sistema abiótico en donde se determinó el perfil hidrodinámico a diferentes flujos de aireación, así mismo el acoplamiento de las ecuaciones de transporte de masa y calor al modelo hidrodinámico permitieron obtener los perfiles de transferencia de O2 y calor en el biorreactor a diferentes intervalos de tiempo, en donde se obtuvo una diferencia de 2.2 °C para la temperatura comprendida entre la superficie y la base del lecho empacado. Por otro lado se modeló el biorreactor considerando un sistema biótico en dónde se obtuvieron los perfiles de producción de CO2 y consumo de O2, así mismo mediante un análisis de sensibilidad se obtuvieron los perfiles de producción de CO2 a diferentes tasas específicas de crecimiento (μX) a valores de 0.23 a 0.66 h -1 donde se observó que a mayor valor de μX (0.66 h -1 ) la curva del perfil de producción de CO2 es más estrecha y se obtiene una tasa de producción de CO2 mayor (11.83 2⁄ℎ) en comparación con los obtenidos (6.58 2⁄ℎ) a valores más bajos (0.23 h -1 ) de μX

Anti-Inflammatory Effect of Ethanolic Extract from <i>Tabebuia rosea</i> (Bertol.) DC., Quercetin, and Anti-Obesity Drugs in Adipose Tissue in Wistar Rats with Diet-Induced Obesity

Obesity is characterized by the excessive accumulation of fat, which triggers a low-grade chronic inflammatory process. Currently, the search for compounds with anti-obesogenic effects that help reduce body weight, as well as associated comorbidities, continues. Among this group of compounds are plant extracts and flavonoids with a great diversity of action mechanisms associated with their beneficial effects, such as anti-inflammatory effects and/or as signaling molecules. In the bark of Tabebuia rosea tree, there are different classes of metabolites with anti-inflammatory properties, such as quercetin. Therefore, the present work studied the effect of the ethanolic extract of T. rosea and quercetin on the mRNA of inflammation markers in obesity compared to the drugs currently used. Total RNA was extracted from epididymal adipose tissue of high-fat diet-induced obese Wistar rats treated with orlistat, phentermine, T. rosea extract, and quercetin. The rats treated with T. rosea and quercetin showed 36 and 31% reductions in body weight compared to the obese control, and they likewise inhibited pro-inflammatory molecules: Il6, Il1b, Il18, Lep, Hif1a, and Nfkb1 without modifying the expression of Socs1 and Socs3. Additionally, only T. rosea overexpressed Lipe. Both T. rosea and quercetin led to a reduction in the expression of pro-inflammatory genes, modifying signaling pathways, which led to the regulation of the obesity-inflammation state

Evaluating the Performance of <i>Yarrowia lipolytica</i> 2.2ab in Solid-State Fermentation under Bench-Scale Conditions in a Packed-Tray Bioreactor

Solid-State Fermentation (SSF) offers a valuable process for converting agri-food by-products (AFBP) into high-value metabolites, with Yarrowia lipolytica 2.2ab (Yl2.2ab) showing significant potential under laboratory-scale controlled conditions; however, its assessment in larger-scale bioreactor scenarios is needed. This work evaluates Yl2.2ab’s performance in a bench-scale custom-designed packed-tray bioreactor. Key features of this bioreactor design include a short packing length, a wall-cooling system, and forced aeration, enhancing hydrodynamics and heat and mass transfer within the tray. Preliminary studies under both abiotic and biotic conditions assessed Yl2.2ab’s adaptability to extreme temperature variations. The results indicated effective oxygen transport but poor heat transfer within the tray bed, with Yl2.2ab leading to a maximum growth rate of 28.15 mgx gssdb−1 h−1 and maximum production of proteases of 40.10 U gssdb−1 h−1, even when temperatures at the packed-tray outlet were around 49 °C. Hybrid-based modeling, incorporating Computational Fluid Dynamics (CFD) and Pseudo-Continuous Simulations (PCSs), elucidated that the forced-aeration system successfully maintained necessary oxygen levels in the bed. However, the low thermal conductivity of AFBP posed challenges for heat transfer. The bioreactor design presents promising avenues for scaling up SSF to valorize AFBP using Yl2.2ab’s extremophilic capabilities

Sparsentan in patients with IgA nephropathy: a prespecified interim analysis from a randomised, double-blind, active-controlled clinical trial

Background: Sparsentan is a novel, non-immunosuppressive, single-molecule, dual endothelin and angiotensin receptor antagonist being examined in an ongoing phase 3 trial in adults with IgA nephropathy. We report the prespecified interim analysis of the primary proteinuria efficacy endpoint, and safety. Methods: PROTECT is an international, randomised, double-blind, active-controlled study, being conducted in 134 clinical practice sites in 18 countries. The study examines sparsentan versus irbesartan in adults (aged ≥18 years) with biopsy-proven IgA nephropathy and proteinuria of 1·0 g/day or higher despite maximised renin-angiotensin system inhibitor treatment for at least 12 weeks. Participants were randomly assigned in a 1:1 ratio to receive sparsentan 400 mg once daily or irbesartan 300 mg once daily, stratified by estimated glomerular filtration rate at screening (30 to 1·75 g/day). The primary efficacy endpoint was change from baseline to week 36 in urine protein-creatinine ratio based on a 24-h urine sample, assessed using mixed model repeated measures. Treatment-emergent adverse events (TEAEs) were safety endpoints. All endpoints were examined in all participants who received at least one dose of randomised treatment. The study is ongoing and is registered with ClinicalTrials.gov, NCT03762850. Findings: Between Dec 20, 2018, and May 26, 2021, 404 participants were randomly assigned to sparsentan (n=202) or irbesartan (n=202) and received treatment. At week 36, the geometric least squares mean percent change from baseline in urine protein-creatinine ratio was statistically significantly greater in the sparsentan group (-49·8%) than the irbesartan group (-15·1%), resulting in a between-group relative reduction of 41% (least squares mean ratio=0·59; 95% CI 0·51-0·69; p<0·0001). TEAEs with sparsentan were similar to irbesartan. There were no cases of severe oedema, heart failure, hepatotoxicity, or oedema-related discontinuations. Bodyweight changes from baseline were not different between the sparsentan and irbesartan groups. Interpretation: Once-daily treatment with sparsentan produced meaningful reduction in proteinuria compared with irbesartan in adults with IgA nephropathy. Safety of sparsentan was similar to irbesartan. Future analyses after completion of the 2-year double-blind period will show whether these beneficial effects translate into a long-term nephroprotective potential of sparsentan. Funding: Travere Therapeutics

Efficacy and safety of sparsentan versus irbesartan in patients with IgA nephropathy (PROTECT): 2-year results from a randomised, active-controlled, phase 3 trial

BackgroundSparsentan, a novel, non-immunosuppressive, single-molecule, dual endothelin angiotensin receptor antagonist, significantly reduced proteinuria versus irbesartan, an angiotensin II receptor blocker, at 36 weeks (primary endpoint) in patients with immunoglobulin A nephropathy in the phase 3 PROTECT trial's previously reported interim analysis. Here, we report kidney function and outcomes over 110 weeks from the double-blind final analysis.MethodsPROTECT, a double-blind, randomised, active-controlled, phase 3 study, was done across 134 clinical practice sites in 18 countries throughout the Americas, Asia, and Europe. Patients aged 18 years or older with biopsy-proven primary IgA nephropathy and proteinuria of at least 1·0 g per day despite maximised renin–angiotensin system inhibition for at least 12 weeks were randomly assigned (1:1) to receive sparsentan (target dose 400 mg oral sparsentan once daily) or irbesartan (target dose 300 mg oral irbesartan once daily) based on a permuted-block randomisation method. The primary endpoint was proteinuria change between treatment groups at 36 weeks. Secondary endpoints included rate of change (slope) of the estimated glomerular filtration rate (eGFR), changes in proteinuria, a composite of kidney failure (confirmed 40% eGFR reduction, end-stage kidney disease, or all-cause mortality), and safety and tolerability up to 110 weeks from randomisation. Secondary efficacy outcomes were assessed in the full analysis set and safety was assessed in the safety set, both of which were defined as all patients who were randomly assigned and received at least one dose of randomly assigned study drug. This trial is registered with ClinicalTrials.gov, NCT03762850.FindingsBetween Dec 20, 2018, and May 26, 2021, 203 patients were randomly assigned to the sparsentan group and 203 to the irbesartan group. One patient from each group did not receive the study drug and was excluded from the efficacy and safety analyses (282 [70%] of 404 included patients were male and 272 [67%] were White) . Patients in the sparsentan group had a slower rate of eGFR decline than those in the irbesartan group. eGFR chronic 2-year slope (weeks 6–110) was −2·7 mL/min per 1·73 m2 per year versus −3·8 mL/min per 1·73 m2 per year (difference 1·1 mL/min per 1·73 m2 per year, 95% CI 0·1 to 2·1; p=0·037); total 2-year slope (day 1–week 110) was −2·9 mL/min per 1·73 m2 per year versus −3·9 mL/min per 1·73 m2 per year (difference 1·0 mL/min per 1·73 m2 per year, 95% CI −0·03 to 1·94; p=0·058). The significant reduction in proteinuria at 36 weeks with sparsentan was maintained throughout the study period; at 110 weeks, proteinuria, as determined by the change from baseline in urine protein-to-creatinine ratio, was 40% lower in the sparsentan group than in the irbesartan group (−42·8%, 95% CI −49·8 to −35·0, with sparsentan versus −4·4%, −15·8 to 8·7, with irbesartan; geometric least-squares mean ratio 0·60, 95% CI 0·50 to 0·72). The composite kidney failure endpoint was reached by 18 (9%) of 202 patients in the sparsentan group versus 26 (13%) of 202 patients in the irbesartan group (relative risk 0·7, 95% CI 0·4 to 1·2). Treatment-emergent adverse events were well balanced between sparsentan and irbesartan, with no new safety signals.InterpretationOver 110 weeks, treatment with sparsentan versus maximally titrated irbesartan in patients with IgA nephropathy resulted in significant reductions in proteinuria and preservation of kidney function

Efficacy and safety of sparsentan versus irbesartan in patients with IgA nephropathy (PROTECT): 2-year results from a randomised, active-controlled, phase 3 trial

Background Sparsentan, a novel, non-immunosuppressive, single-molecule, dual endothelin angiotensin receptor antagonist, significantly reduced proteinuria versus irbesartan, an angiotensin II receptor blocker, at 36 weeks (primary endpoint) in patients with immunoglobulin A nephropathy in the phase 3 PROTECT trial's previously reported interim analysis. Here, we report kidney function and outcomes over 110 weeks from the double-blind final analysis. Methods PROTECT, a double-blind, randomised, active-controlled, phase 3 study, was done across 134 clinical practice sites in 18 countries throughout the Americas, Asia, and Europe. Patients aged 18 years or older with biopsy-proven primary IgA nephropathy and proteinuria of at least 1·0 g per day despite maximised renin–angiotensin system inhibition for at least 12 weeks were randomly assigned (1:1) to receive sparsentan (target dose 400 mg oral sparsentan once daily) or irbesartan (target dose 300 mg oral irbesartan once daily) based on a permuted-block randomisation method. The primary endpoint was proteinuria change between treatment groups at 36 weeks. Secondary endpoints included rate of change (slope) of the estimated glomerular filtration rate (eGFR), changes in proteinuria, a composite of kidney failure (confirmed 40% eGFR reduction, end-stage kidney disease, or all-cause mortality), and safety and tolerability up to 110 weeks from randomisation. Secondary efficacy outcomes were assessed in the full analysis set and safety was assessed in the safety set, both of which were defined as all patients who were randomly assigned and received at least one dose of randomly assigned study drug. This trial is registered with ClinicalTrials.gov, NCT03762850. Findings Between Dec 20, 2018, and May 26, 2021, 203 patients were randomly assigned to the sparsentan group and 203 to the irbesartan group. One patient from each group did not receive the study drug and was excluded from the efficacy and safety analyses (282 [70%] of 404 included patients were male and 272 [67%] were White) . Patients in the sparsentan group had a slower rate of eGFR decline than those in the irbesartan group. eGFR chronic 2-year slope (weeks 6–110) was −2·7 mL/min per 1·73 m2 per year versus −3·8 mL/min per 1·73 m2 per year (difference 1·1 mL/min per 1·73 m2 per year, 95% CI 0·1 to 2·1; p=0·037); total 2-year slope (day 1–week 110) was −2·9 mL/min per 1·73 m2 per year versus −3·9 mL/min per 1·73 m2 per year (difference 1·0 mL/min per 1·73 m2 per year, 95% CI −0·03 to 1·94; p=0·058). The significant reduction in proteinuria at 36 weeks with sparsentan was maintained throughout the study period; at 110 weeks, proteinuria, as determined by the change from baseline in urine protein-to-creatinine ratio, was 40% lower in the sparsentan group than in the irbesartan group (−42·8%, 95% CI −49·8 to −35·0, with sparsentan versus −4·4%, −15·8 to 8·7, with irbesartan; geometric least-squares mean ratio 0·60, 95% CI 0·50 to 0·72). The composite kidney failure endpoint was reached by 18 (9%) of 202 patients in the sparsentan group versus 26 (13%) of 202 patients in the irbesartan group (relative risk 0·7, 95% CI 0·4 to 1·2). Treatment-emergent adverse events were well balanced between sparsentan and irbesartan, with no new safety signals. Interpretation Over 110 weeks, treatment with sparsentan versus maximally titrated irbesartan in patients with IgA nephropathy resulted in significant reductions in proteinuria and preservation of kidney function.</p