Oxygen uptake rate (OUR) control strategy for improving avermectin B1a production during fed-batch fermentation on industrial scale (150 m3)

Glucose metabolism plays a crucial role in the process of avermectin B1a biosynthesis. Controlling glucose feeding based on oxygen uptake rate (OUR) was established to improve the efficiency of avermectin B1a  production. The result showed that avermectin B1a production was greatly enhanced by OUR control strategy. In the glucose feeding phase, OUR was maintained at approximate 12 mmol/L/h, which was conducive to avermectin B1a biosynthesis. Using this OUR control strategy, an adequate supply of organic acid precursors produced avermectin B1a 5228 U/mL, which was 22.8% higher than that of the control (batch fermentation, 4256 U/mL) on industrial scale.Key words: Avermectin B1a, glucose feeding, oxygen uptake rate, industrial scale

Engineering cofactor metabolism for improved protein and glucoamylase production in Aspergillus niger

Background: Nicotinamide adenine dinucleotide phosphate (NADPH) is an important cofactor ensuring intracellular redox balance, anabolism and cell growth in all living systems. Our recent multi-omics analyses of glucoamylase (GlaA) biosynthesis in the filamentous fungal cell factory Aspergillus niger indicated that low availability of NADPH might be a limiting factor for GlaA overproduction. Results: We thus employed the Design-Build-Test-Learn cycle for metabolic engineering to identify and prioritize effective cofactor engineering strategies for GlaA overproduction. Based on available metabolomics and 13C metabolic flux analysis data, we individually overexpressed seven predicted genes encoding NADPH generation enzymes under the control of the\ua0Tet-on gene switch in two A. niger recipient strains, one carrying a single and one carrying seven glaA gene copies, respectively, to test their individual effects on GlaA and total protein overproduction. Both strains were selected to understand if a strong pull towards glaA biosynthesis (seven gene copies) mandates a higher NADPH supply compared to the native condition (one gene copy). Detailed analysis of all 14 strains cultivated in shake flask cultures uncovered that overexpression of the gsdA gene (glucose 6-phosphate dehydrogenase), gndA gene (6-phosphogluconate dehydrogenase) and maeA gene (NADP-dependent malic enzyme) supported GlaA production on a subtle (10%) but significant level in the background strain carrying seven glaA gene copies. We thus performed maltose-limited chemostat cultures combining metabolome analysis for these three isolates to characterize metabolic-level fluctuations caused by cofactor engineering. In these cultures, overexpression of either the gndA or maeA gene increased the intracellular NADPH pool by 45% and 66%, and the yield of GlaA by 65% and 30%, respectively. In contrast, overexpression of the gsdA gene had a negative effect on both total protein and glucoamylase production. Conclusions: This data suggests for the first time that increased NADPH availability can indeed underpin protein and especially GlaA production in strains where a strong pull towards GlaA biosynthesis exists. This data also indicates that the highest impact on GlaA production can be engineered on a genetic level by increasing the flux through the pentose phosphate pathway (gndA gene) followed by engineering the flux through the reverse TCA cycle (maeA gene). We thus propose that NADPH cofactor engineering is indeed a valid strategy for metabolic engineering of A. niger to improve GlaA production, a strategy which is certainly also applicable to the rational design of other microbial cell factories.[Figure not available: see fulltext.]

Visualisation of J-type counter-current chromatography: A route to understand hydrodynamic phase distribution and retention

This article can be accessed from the link below - Copyright @ 2012 Elsevier. The article was made available through open access by the Brunel Open Access Publishing Fund.This paper has addressed decade sought-after questions on phase bilateral distribution and stationary phase retention in any J-type high-speed counter-current chromatographic (CCC) centrifuge. Using a 2-D spiral column operated on such a CCC device and an aqueous two-phase system, this work systematically observed the phase interaction during transitional period and at dynamic equilibration under stroboscopic illumination. The experimental results thus obtained were used to examine the effects of the liquid–solid friction force, tangential centrifugal force, and physical properties of the two-phase system on hydrodynamic phase behaviour. We identified that (a) density difference between lower and upper phases is the critical factor to cause unusual phase bilateral distribution in the 2-D spiral column and (b) interfacial tension (manifested primarily as phase settling time) of any two-phase system is the critical factor in explaining inability to retain stationary phase in 3-D helical column and, for certain flow modes, in the 2-D spiral column. This work thus has extended or modified the well-established rule-of-thumb for operating J-type CCC devices and our conclusions can accommodate virtually all the anomalies concerning both hydrophobic and hydrophilic phase systems. To this end, this work has not only documented valuable experimental evidences for directly observing phase behaviour in a CCC column, but also finally resolved fundamentally vital issues on bilateral phase distribution orientation and stationary phase retention in 2-D spiral and 3-D helical CCC columns. Revised recommendations to end users of this technology could thus be derived out of the essence of the present work presumably following further experimental validation and a consensus in the CCC R&D and manufacturing circle.This work is funded by the BBSRC (UK) grant BB/C5063341/1, by MoST (China) as a Major State Basic Research Development Programme grant 2007CB714303 (973 Programme), by M & G Trading, and by Shanghai Science and Technology Board grant 11DZ2292700

Facile one-pot synthesis of three different substituted thiazoles from propargylic alcohols

Three different substituted thiazoles have been successfully synthesized from readily available propargylic alcohols. Various secondary propargylic alcohols or tertiary propargylic alcohols participated well in the reaction, providing the desired products in good yields. This method provides a flexible and rapid route to substituted thiazoles.National Natural Science Foundation of China [20772098]; Program for New Century Excellent Talents in Fujian Province Universit

FeCl3-Catalyzed Propargylation-Cycloisomerization Tandem Reaction: A Facile One-Pot Synthesis of Substituted Furans

An efficient FeCl3-catalyzed tandem propargylation-cycloisomerization reaction of propargylic alcohols or acetates with 1,3-dicarbonyl compounds, leading to the synthesis of substituted furans, has been developed.National Natural Science Foundation of China [20772098, 30572250]; Fujian Province Universit

Malaria incidence from 2005–2013 and its associations with meteorological factors in Guangdong, China

Background: The temporal variation of malaria incidence has been linked to meteorological factors in many studies, but key factors observed and corresponding effect estimates were not consistent. Furthermore, the potential effect modification by individual characteristics is not well documented. This study intends to examine the delayed effects of meteorological factors and the sub-population's susceptibility in Guangdong, China. Methods: The Granger causality Wald test and Spearman correlation analysis were employed to select climatic variables influencing malaria. The distributed lag non-linear model (DLNM) was used to estimate the non-linear and delayed effects of weekly temperature, duration of sunshine, and precipitation on the weekly number of malaria cases after controlling for other confounders. Stratified analyses were conducted to identify the sub-population's susceptibility to meteorological effects by malaria type, gender, and age group. Results: An incidence rate of 1.1 cases per 1,000,000 people was detected in Guangdong from 2005-2013. High temperature was associated with an observed increase in malaria incidence, with the effect lasting for four weeks and a maximum relative risk (RR) of 1.57 (95% confidence interval (CI): 1.06-2.33) by comparing 30°C to the median temperature. The effect of sunshine duration peaked at lag five and the maximum RR was 1.36 (95% CI: 1.08-1.72) by comparing 24 hours/week to 0 hours/week. A J-shaped relationship was found between malaria incidence and precipitation with a threshold of 150 mm/week. Over the threshold, precipitation increased malaria incidence after four weeks with the effect lasting for 15 weeks, and the maximum RR of 1.55 (95% CI: 1.18-2.03) occurring at lag eight by comparing 225 mm/week to 0 mm/week. Plasmodium falciparum was more sensitive to temperature and precipitation than Plasmodium vivax. Females had a higher susceptibility to the effects of sunshine and precipitation, and children and the elderly were more sensitive to the change of temperature, sunshine duration, and precipitation. Conclusion: Temperature, duration of sunshine and precipitation played important roles in malaria incidence with effects delayed and varied across lags. Climatic effects were distinct among sub-groups. This study provided helpful information for predicting malaria incidence and developing the future warning system.School of Nursin

Depressive Neurosis Treated by Acupuncture for Regulating the Liver —A Report of 176 Cases

ObjectiveTo observe therapeutic effect of acupuncture for regulating the liver on depressive neurosis.MethodsIn a multi-center randomized controlled trial, 440 patients were divided into 3 groups: Acupuncture group for regulating the liver (Acup., 176 cases) was treated by acupuncture at Siguan Points, i.e. bilateral Hegu (LI 4) and Taichong (LR 3), Baihui (GV 20) and Yintang (EX-HN3) plus ear-acupuncture, Prozac group (P., 176 cases) by oral administration of Prozac, and Non-acupoint needling group (NAN, 88 cases) by acupuncture at non-acupoints as acupuncture placebo. Self-rating Depression Scale (SDS) was examined before treatment, and one month, two and three months after treatment respectively to evaluate therapeutic effect, and Rating Scale for Side Effects (SERS) was used to evaluate the safety.ResultsAfter one month of treatment, SDS scores in Acup. Group were significantly lower than that in P. Group (P<0.05) and than that in NAN Group (P<0.01), and SDS scores in P. Group were lower than that in NAN Group (P<0.05), showing the SDS scores in Acup. Group <P. Group <NAN Group. After 2 months of treatment, SDS scores in Acup. Group were also significantly lower than that in P. Group (P<0.01) and than that in NAN Group (P<0.01), and SDS scores in P. Group were also lower than that in NAN Group (P<0.05), showing the SDS scores in Acup. Group <P. Group <NAN Group. After 3 months of treatment, SDS scores in Acup. Group were also significantly lower than that in P. Group (P<0.01) and than that in NAN Group (P<0.01), and SDS scores in P. Group were also lower than that in NAN Group (P<0.01), showing the SDS score in Acup. Group <P. Group <NAN Group. After treatment, SERS scores were 0.16±0.95, 6.51±5.09 and 0.23±1.36 in Acup. Group, P. Group and NAN Group respectively. A significant difference existed between Acup. Group and P. Group (P<0.05), but no significant difference between Acup. Group and NAN Group (P>0.05), showing the SERS scores in Acup. Group <NAN Group <P. Group. No side effect was found in Acup. and NAN groups.ConclusionThe therapeutic effect of acupuncture on depressive neurosis is better than or similar to that of Prozac but with less side effect

Unraveling the Novel Protective Effect of Patchouli Alcohol Against Helicobacter pylori-Induced Gastritis: Insights Into the Molecular Mechanism in vitro and in vivo

Patchouli alcohol (PA), a natural tricyclic sesquiterpene extracted from Pogostemon cablin (Blanco) Benth. (Labiatae), has been found to exhibit anti-Helicobacter pylori and anti-inflammatory properties. In this study, we investigated the protective effect of PA against H. pylori-induced gastritis in vitro and in vivo, and determined the underlying mechanism. In the in vivo experiment, a C57BL/6 mouse model of gastritis was established using H. pylori SS1, and treatments with standard triple therapy or 5, 10, and 20 mg/kg PA were performed for 2 weeks. Results indicated that PA effectively attenuated oxidative stress by decreasing contents of intracellular reactive oxygen species (ROS) and malonyldialdehyde (MDA), and increasing levels of non-protein sulfhydryl (NP-SH), catalase and glutathione (GSH)/glutathione disulphide (GSSG). Additionally, treatment with PA significantly attenuated the secretions of interleukin 1 beta (IL-1β), keratinocyte chemoattractant and interleukin 6 (IL-6). PA (20 mg/kg) significantly protected the gastric mucosa from H. pylori-induced damage. In the in vitro experiment, GES-1 cells were cocultured with H. pylori NCTC11637 at MOI = 100:1 and treated with different doses of PA (5, 10, and 20 μg/ml). Results indicated that PA not only significantly increased the cell viability and decreased cellular lactate dehydrogenase (LDH) leakage, but also markedly elevated the mitochondrial membrane potential and remarkably attenuated GES-1 cellular apoptosis, thereby protecting gastric epithelial cells against injuries caused by H. pylori. PA also inhibited the secretions of pro-inflammatory factors, such as monocyte chemotactic protein 1 (MCP-1), tumor necrosis factor-α (TNF-α) and IL-6. Furthermore, after PA treatment, the combination of NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) and cysteine-aspartic proteases 1 (CASPASE-1), the expression levels of NLRP3 inflammasome-related proteins, such as thioredoxin-interacting protein (TXNIP), pro-CASPASE-1, cle-CASPASE-1, and NLRP3 and genes (NLRP3 and CASPASE1) were significantly decreased as compared to the model group. In conclusion, treatment with PA for 2 weeks exhibited highly efficient protective effect against H. pylori-induced gastritis and related damages. The underlying mechanism might involve antioxidant activity, inhibition of pro-inflammatory factor and regulation of NLRP3 inflammasome function. PA exerted anti-H. pylori and anti-gastritis effects and thus had the potential to be a promising candidate for treatment of H. pylori-related diseases

Copper(II)-Catalyzed Synthesis of Pyrimidines from Propargylic Alcohols and Amidine: A Propargylation-Cyclization-Oxidation Tandem Reaction

A new approach to the tandem synthesis of 2,4-disubstituted or 2,4,6-trisubstituted pyrimidines from propargylic alcohols with amidine is described. This reaction is catalyzed by 20 mol% Cu(OTf)(2) to give pyrimidines in moderate to good yields via a propargylation-cyclization-oxidation tandem sequence.National Natural Science Foundation of China[21072159]; Science & Technology Bureau of Xiamen[3502Z20093007]; NFFTBS[J1030415

Classification and hemodynamic characteristics of delayed intracerebral hemorrhage following stent-assisted coil embolism in unruptured intracranial aneurysms

Background and objectiveStent-assisted coil (SAC) embolization is a commonly used endovascular treatment for unruptured intracranial aneurysms (UIAs) but can be associated with symptomatic delayed intracerebral hemorrhage (DICH). Our study aimed to investigate the hemodynamic risk factors contributing to DICH following SAC embolization and to establish a classification for DICH predicated on hemodynamic profiles.MethodsThis retrospective study included patients with UIAs located in the internal carotid artery (ICA) treated with SAC embolization at our institution from January 2021 to January 2022. We focused on eight patients who developed postoperative DICH and matched them with sixteen control patients without DICH. Using computational fluid dynamics, we evaluated the hemodynamic changes in distal arteries [terminal ICA, the anterior cerebral artery (ACA), and middle cerebral artery (MCA)] pre-and post-embolization. We distinguished DICH-related arteries from unrelated ones (ACA or MCA) and compared their hemodynamic alterations. An imbalance index, quantifying the differential in flow velocity changes between ACA and MCA post-embolization, was employed to gauge the flow distribution in distal arteries was used to assess distal arterial flow distribution.ResultsWe identified two types of DICH based on postoperative flow alterations. In type 1, there was a significant lower in the mean velocity increase rate of the DICH-related artery compared to the unrelated artery (−47.25 ± 3.88% vs. 42.85 ± 3.03%; p &lt; 0.001), whereas, in type 2, there was a notable higher (110.58 ± 9.42% vs. 17.60 ± 4.69%; p &lt; 0.001). Both DICH types demonstrated a higher imbalance index than the control group, suggesting an association between altered distal arterial blood flow distribution and DICH occurrence.ConclusionDICH in SAC-treated UIAs can manifest as either a lower (type 1) or higher (type 2) in the rate of velocity in DICH-related arteries. An imbalance in distal arterial blood flow distribution appears to be a significant factor in DICH development