Engineering the ß-galactosidase from Aspergillus oryzae for making lactose-free and no-sugar-added yogurt.

Yogurt usually contains 5% to 7% sugar and 3% to 5% lactose. As ß-galactosidases can hydrolyze lactose and improve sweetness, they have the potential to produce lactose-free (LF) and no-sugar-added (NSA) yogurt. In this study, the ß-galactosidase AoBgal35A from Aspergillus oryzae was engineered by site-saturation mutagenesis. Results of 19 variants of T955 residue showed that the lactose hydrolysis rate of T955R-AoBgal35A was up to 90.7%, which is much higher than the 78.5% of the wild type. Moreover, the optimal pH of T955R-AoBgal35A was shifted from pH 4.5 to pH 5.5, and the optimal temperature decreased from 60°C to 50°C. The mutant T955R-AoBgal35A was successfully expressed in Komagataella pastoris, which produced extracellularly 4,528 U/mL of ß-galactosidase activity. The mutant T955R-AoBgal35A was used to produce LF yogurt. The Streptococcus thermophilus count of LF yogurt increased from 7.9 to 9.5 log cfu/g, which is significantly higher than that of the control group (8.9 log cfu/g). The residual lactose content of LF yogurt was 0.13%, meeting the requirements of the national standard in China for the "lactose-free" label (<0.5%). Furthermore, sugar in yogurt was replaced by whey powder to produce LF-NSA yogurt. The optimal addition content of whey powder was 7.5%. The texture, water-holding capacity, and titratable acidity of LF and LF-NSA yogurt achieved good shelf life stability. Therefore, this study provides an insight for technological implications of ß-galactosidases in the dairy industry.

Cu(I)/Chiral Bisoxazoline-Catalyzed Enantioselective Doyle-Kirmse Reaction of Allenyl Sulfides with α-Diazoesters.

Herein, a Cu(I)-catalyzed enantioselective Doyle-Kirmse reaction of allenyl sulfides and α-diazoesters is reported, which provides an efficient synthetic route to enantio-enriched chiral tertiary homopropargylic sulfides. This reaction features high enantioselectivities (up to 96 % ee) and good functional group tolerance. The alkyl substituted α-diazoester has also been demonstrated as the efficient substrates in the asymmetric Doyle-Kirmse reaction. Mechanistic studies, including kinetic experiments, were conducted to gain insights into the details of the reaction pathway. The potential synthetic utility of this protocol has also been demonstrated.

Catalytic Asymmetric Homologation of 4-Substituted Cyclohexanones with CF3 CHN2 : Enantioselective Synthesis of α-Trifluoromethyl Cycloheptanones.

Introduction of the trifluoromethyl group (CF3 ) into organic molecules in an enantioselective manner has attracted significant attention, but still remains a challenging problem. We herein report a catalytic asymmetric trifluoromethylation of cyclic ketones via a ScIII /chiral bisoxazoline-catalyzed homologation reaction by employing 2,2,2-trifluorodiazoethane (CF3 CHN2 ) as the CF3 source. This desymmetrization process is highly efficient and generates two chiral centers with excellent diastereoselectivity and enantioselectivity, affording chiral α-trifluoromethyl cyclic ketones in a straightforward manner.

Design and Experimentation of a Self-Sensing Actuator for Active Vibration Isolation System with Adjustable Anti-Resonance Frequency Controller.

The vibration isolation system is now indispensable to high-precision instruments and equipment, which can provide a low vibration environment to ensure performance. However, the disturbance with variable frequency poses a challenge to the vibration isolation system, resulting in precision reduction of dynamic modeling. This paper presents a velocity self-sensing method and experimental verification of a vibration isolation system. A self-sensing actuator is designed to isolate the vibration with varying frequencies according to the dynamic vibration absorber structure. The mechanical structure of the actuator is illustrated, and the dynamic model is derived. Then a self-sensing method is proposed to adjust the anti-resonance frequency of the system without velocity sensors, which can also reduce the complexity of the system and prevent the disturbance transmitting along the cables. The self-sensing controller is constructed to track the variable frequency of the disturbance. A prototype of the isolation system equipped with velocity sensors is developed for the experiment. The experiment results show that the closed-loop transmissibility is less than -5 dB in the whole frequency rand and is less than -40 dB around, adding anti-resonance frequency which can be adjusted from 0 Hz to initial anti-resonance frequency. The disturbance amplitude of the payload can be suppressed to 10%.

Cu(I)/Chiral Bisoxazoline-Catalyzed Enantioselective Sommelet-Hauser Rearrangement of Sulfonium Ylides.

Catalytic asymmetric thia-Sommelet-Hauser rearrangement of sulfonium ylides remains a great challenge due to its multistep reaction mechanism involving metal carbene formation, proton transfer, and [2,3]-sigmatropic rearrangement. In particular, the key problem of such reactions is the differentiation of the enantiotopic lone pair electrons of sulfur, which generates the sulfonium ylide intermediate bearing chirality on the sulfur atom. With a modified chiral bisoxazoline ligand, we developed a Cu(I)-catalyzed asymmetric thia-Sommelet-Hauser rearrangement with good to excellent enantioselectivities. Mechanistic studies provide insights into the details of the reaction mechanism.

Palladium-Catalyzed Cycloaromatization/Alkylation of o-(Alkynyl)styrenes.

A Pd(II)-catalyzed mild and highly regioselective 6-endo cyclization/alkylation reaction of o-(alkynyl)styrenes with simple allylic alcohols has been developed. Under mild reaction conditions, the vinyl palladium species generated in situ after cyclization could insert a C-C double bond of allylic alcohol through a cross-coupling reaction and led to the formation of (alkyl)naphthalenes. This cascade cross-coupling reaction represents a direct and atom economic method for the construction of functionalized naphthalene derivatives in moderate to good yields.

Macrolide Synthesis through Intramolecular Oxidative Cross-Coupling of Alkenes.

A RhIII -catalyzed intramolecular oxidative cross-coupling between double bonds for the synthesis of macrolides is described. Under the optimized reaction conditions, macrocycles containing a diene moiety can be formed in reasonable yields and with excellent chemo- and stereoselectivity. This method provides an efficient approach to synthesize macrocyclic compounds containing a 1,3-conjugated diene structure.

Spatiotemporal distribution of nitrogen and phosphorus in alpine lakes in the Sanjiangyuan Region of the Tibetan Plateau.

To provide a theoretical basis for alpine source lake protection, ten samples were taken from each lake annually from 2012 to 2015. Each year, the various species of nitrogen and phosphorus nutrients were measured. The average contents of nitrate nitrogen, ammonia nitrogen, nitrite nitrogen, total phosphorus, and total nitrogen in the four lakes are 0.195-0.0 mg/L, 0.038-0.143 mg/L, 0.004-0.168 mg/L, 0.006-0.740 mg/L, and 0.050-0.547 mg/L, respectively. The total phosphorus contents in Eling Lake, Longbao Lake and Sea Star were higher than Class I water quality standards, and the total nitrogen contents in Eling Lake, Sea Star and Zhaling Lake were higher than Class I water quality standards as well. The concentration contour maps of the nitrate nitrogen, ammonia nitrogen, nitrite nitrogen, total phosphorus and total nitrogen showed that the indicators of the four lakes in the east, the west, and the center of the lake did not have the same trend. From 2012 to 2015, each of the measured nutrients showed a rising trend year by year. The four lakes are polluted by both endogenous and exogenous pollution, and it is necessary to limit the exogenous pollution and protect the alpine lakes immediately.

The Effect of Bifidobacterium animalis subsp. lactis MN-Gup on Glucose Metabolism, Gut Microbiota, and Their Metabolites in Type 2 Diabetic Mice.

Probiotics have garnered increasing attention as a potential therapeutic approach for type 2 diabetes mellitus (T2DM). Previous studies have confirmed that Bifidobacterium animalis subsp. lactis MN-Gup (MN-Gup) could stimulate the secretion of glucagon-like peptide-1 (GLP-1) in NCI-H716 cells, but whether MN-Gup has a hypoglycemic effect on T2DM in vivo remains unclear. In this study, a T2DM mouse model was constructed, with a high-fat diet and streptozotocin in mice, to investigate the effect of MN-Gup on diabetes. Then, different doses of MN-Gup (2 × 109 CFU/kg, 1 × 1010 CFU/kg) were gavaged for 6 weeks to investigate the effect of MN-Gup on glucose metabolism and its potential mechanisms. The results showed that a high-dose of MN-Gup significantly reduced the fasting blood glucose (FBG) levels and homeostasis model assessment-insulin resistance (HOMA-IR) of T2DM mice compared to the other groups. In addition, there were significant increases in the short-chain fatty acids (SCFAs), especially acetate, and GLP-1 levels in the MN-Gup group. MN-Gup increased the relative abundance of Bifidobacterium and decreased the number of Escherichia-Shigella and Staphylococcus. Moreover, the correlation analysis revealed that Bifidobacterium demonstrated a significant positive correlation with GLP-1 and a negative correlation with the incremental AUC. In summary, this study demonstrates that Bifidobacterium animalis subsp. lactis MN-Gup has significant hypoglycemic effects in T2DM mice and can modulate the gut microbiota, promoting the secretion of SCFAs and GLP-1.

Predicting protein-ATP binding sites from primary sequence through fusing bi-profile sampling of multi-view features.

BACKGROUND: Adenosine-5'-triphosphate (ATP) is one of multifunctional nucleotides and plays an important role in cell biology as a coenzyme interacting with proteins. Revealing the binding sites between protein and ATP is significantly important to understand the functionality of the proteins and the mechanisms of protein-ATP complex. RESULTS: In this paper, we propose a novel framework for predicting the proteins' functional residues, through which they can bind with ATP molecules. The new prediction protocol is achieved by combination of sequence evolutional information and bi-profile sampling of multi-view sequential features and the sequence derived structural features. The hypothesis for this strategy is single-view feature can only represent partial target's knowledge and multiple sources of descriptors can be complementary. CONCLUSIONS: Prediction performances evaluated by both 5-fold and leave-one-out jackknife cross-validation tests on two benchmark datasets consisting of 168 and 227 non-homologous ATP binding proteins respectively demonstrate the efficacy of the proposed protocol. Our experimental results also reveal that the residue structural characteristics of real protein-ATP binding sites are significant different from those normal ones, for example the binding residues do not show high solvent accessibility propensities, and the bindings prefer to occur at the conjoint points between different secondary structure segments. Furthermore, results also show that performance is affected by the imbalanced training datasets by testing multiple ratios between positive and negative samples in the experiments. Increasing the dataset scale is also demonstrated useful for improving the prediction performances.

Pd-Catalyzed coupling of benzyl bromides with BMIDA-substituted N-tosylhydrazones: synthesis of trans-alkenyl MIDA boronates.

A palladium-catalyzed stereoselective synthesis of alkenyl boronates from N-methyliminodiacetyl boronate (BMIDA)-substituted N-tosylhydrazone and benzyl bromides is developed. A range of trans-alkenyl MIDA boronates as single stereoisomers were obtained in moderate yields with good functional group compatibility. The resultant boronate products may be transformed to other boron-containing compounds and may also be directly used in cross-coupling reactions.

The Removal of Erythromycin and Its Effects on Anaerobic Fermentation.

In view of the problems of antibiotic pollution, anaerobic fermentation technology was adopted to remove erythromycin in this study. The removal of erythromycin and its effects mechanism on anaerobic fermentation were studied, including biogas performance, process stability, substrate degradability, enzyme activity, and microbial communities. The results showed that the removal rates of erythromycin for all tested concentrations were higher than 90% after fermentation. Erythromycin addition inhibited biogas production. The more erythromycin added, the lower the CH4 content obtained. The high concentration of erythromycin (20 and 40 mg/L) resulted in more remarkable variations of pH values than the control group and 1 mg/L erythromycin added during the fermentation process. Erythromycin inhibited the hydrolysis process in the early stage of anaerobic fermentation. The contents of chemical oxygen demand (COD), NH4+-N, and volatile fatty acids (VFA) of erythromycin added groups were lower than those of the control group. Erythromycin inhibited the degradation of lignocellulose in the late stage of fermentation. Cellulase activity increased first and then decreased during the fermentation and addition of erythromycin delayed the peak of cellulase activity. The inhibitory effect of erythromycin on the activity of coenzyme F420 increased with elevated erythromycin concentrations. The relative abundance of archaea in erythromycin added groups was lower than the control group. The decrease in archaea resulted in the delay of the daily biogas peak. Additionally, the degradation rate of erythromycin was significantly correlated with the cumulative biogas yield, COD, pH, and ORP. This study supports the reutilization of antibiotic-contaminated biowaste and provides references for further research.

Muscle fatigue related to human posture using a brush cutter for landscape gardening: a preliminary study.

BACKGROUND: Brush cutters are widely used in Chinese landscape gardening and agricultural laboring which leads the operators being exposed to many risks. Low back pain (LBP) is particularly common and can lead to substantial personal, community and financial burdens. The aim of the presented study was to measure the activity and function of each torso muscle of the operator when using the bush cutter, so as to evaluate the muscle injury of the operator during using several common brush cutters for different landscape tasks. MATERIAL AND METHODS: The human postures of 6 workers using 2 types of brush cutters in the 3 working conditions were recorded and measured by using a surface electromyography (sEMG) system and a camera. The test results were compared by t-test and sign test. The effect of human posture on the sEMG signal of trunk muscles in different working condition were analyzed by ANOVA. RESULTS: In the 3 working conditions, except for the left trapezius muscle, the muscle load of operating the backpack brush cutter is higher than that of operating side-mounted brush cutter. When operating the side-mounted backpack brush cutter, the force on both sides of the trapezius muscle is uneven, the load of the left trapezius muscle is >25%, and the maximum value is >30%. CONCLUSIONS: The results highlighted significant differences in the effects of different working postures on the muscle activities of workers' trunk. Safe operation standards should therefore be taken to protect the exposed workers and to improve the industrial design of irrigation cutters to prevent the occurrence of occupational diseases. Med Pr. 2022;73(3):201-7.

Effects of Fermented Milk Containing Bifidobacterium animalis Subsp. lactis MN-Gup (MN-Gup) and MN-Gup-Based Synbiotics on Obesity Induced by High Fat Diet in Rats.

Given the probiotic effects previously found in Bifidobacterium animalis subsp. lactis MN-Gup (MN-Gup) and its great application potential in dairy products, this study aimed to investigate the effects of fermented milk containing MN-Gup or MN-Gup-based synbiotics on high fat diet (HFD)-induced obesity in rats. Galacto-oligosaccharides (GOS) and xylo-oligosaccharides (XOS) were selected as the tested prebiotics in MN-Gup-based synbiotics due to their promotion of MN-Gup growth in vitro. After nine weeks of HFD feeding, the obese rats were intervened with fermented milk containing MN-Gup (MN-Gup FM) or its synbiotics (MN-Gup + GOS FM, MN-Gup + XOS FM) for eight weeks. The results showed that the interventions could alleviate HFD-induced body weight gain, epididymal fat deposition, adipocyte hypertrophy, dyslipidemia and inflammation, but GOS and XOS did not exhibit significant synergies with MN-Gup on those alleviations. Furthermore, the interventions could regulate the HFD-affected gut microbiota and microbial metabolites, as shown by the increases in short chain fatty acids (SCFAs) and alterations in obesity-related bile acids (BAs), which may play important roles in the mechanism underlying the alleviation of obesity. This study revealed the probiotic effects of MN-Gup on alleviating obesity and provided the basis for MN-Gup applications in the future.

Dexamethasone enhances glucose uptake by SGLT1 and GLUT1 and boosts ATP generation through the PPP-TCA cycle in bovine neutrophils.

BACKGROUND: Clinical dexamethasone (DEX) treatment or stress in bovines results in extensive physiological changes with prominent hyperglycemia and neutrophils dysfunction. OBJECTIVES: To elucidate the effects of DEX treatment in vivo on cellular energy status and the underlying mechanism in circulating neutrophils. METHODS: We selected eight-month-old male bovines and injected DEX for 3 consecutive days (1 time/d). The levels of glucose, total protein (TP), total cholesterol (TC), and the proinflammatory cytokines interleukin (IL)-1ß, IL-6 and tumor necrosis factor (TNF)-α in blood were examined, and we then detected glycogen and adenosine triphosphate (ATP) content, phosphofructosekinase-1 (PFK1) and glucose-6-phosphate dehydrogenase (G6PDH) activity, glucose transporter (GLUT)1, GLUT4, sodium/glucose cotransporter (SGLT)1 and citrate synthase (CS) protein expression and autophagy levels in circulating neutrophils. RESULTS: DEX injection markedly increased blood glucose, TP and TC levels, the Ca2+/P5+ ratio and the neutrophil/lymphocyte ratio and significantly decreased blood IL-1ß, IL-6 and TNF-α levels. Particularly in neutrophils, DEX injection inhibited p65-NFκB activation and elevated glycogen and ATP contents and SGLT1, GLUT1 and GR expression while inhibiting PFK1 activity, enhancing G6PDH activity and CS expression and lowering cell autophagy levels. CONCLUSIONS: DEX induced neutrophils glucose uptake by enhancing SGLT1 and GLUT1 expression and the transformation of energy metabolism from glycolysis to pentose phosphate pathway (PPP)-tricarboxylic acid (TCA) cycle. This finding gives us a new perspective on deeper understanding of clinical anti-inflammatory effects of DEX on bovine.

Bifidobacterium animalis subsp. lactis MN-Gup protects mice against gut microbiota-related obesity and endotoxemia induced by a high fat diet.

Obesity has become a public health concern due to its global prevalence and high risk of complications such as endotoxemia. Given the important role of gut microbiota in obesity, probiotics targeting gut microbiota have been developed and applied to alleviate obesity. However, most studies focused on the effects of probiotics on pre-existing obesity, and the preventive effects of probiotics against obesity were rarely studied. This study aimed to investigate the preventive effects of Bifidobacterium animalis subsp. lactis MN-Gup (MN-Gup) and fermented milk containing MN-Gup against high fat diet (HFD)-induced obesity and endotoxemia in C57BL/6J mice. The results showed that MN-Gup, especially the high dose of MN-Gup (1 × 1010CFU/kg b.w.), could significantly protect mice against HFD-induced body weight gain, increased fat percentage, dyslipidemia, and increased lipopolysaccharides (LPS). Fermented milk containing MN-Gup had better preventive effects on fat percentage and dyslipidemia than fermented milk without MN-Gup, but its overall performance was less effective than MN-Gup. Furthermore, MN-Gup and fermented milk containing MN-Gup could alter HFD-affected gut microbiota and regulate obesity- or endotoxemia-correlated bacteria, which may contribute to the prevention of obesity and endotoxemia. This study revealed that MN-Gup could reduce obesity and endotoxemia under HFD, thereby providing a potential application of MN-Gup in preventing obesity.

Effect of Zn Addition on the Cd-Containing Anaerobic Fermentation Process: Biodegradation and Microbial Communities.

Anaerobic fermentation is considered as a cost-effective way of biomass waste disposal. However, the compound heavy metals contained in the biomass may induce complex effects on anaerobic fermentation, which limit the utilization of metal-contaminated biowaste. In this study, the impacts of Cd and Zn addition on biogas properties, process stability, substrate biodegradation, enzyme activity, and microbial properties were studied. The results showed that the addition of Cd together with Zn (Cd+Zn) increased the maximum daily and cumulative biogas yields, and brought forward the gas production peak compared with the Cd-added group. Taking the whole fermentation process into account, the promotion effects of adding Zn into the Cd-containing fermentation system on biogas yields were mainly attributable to better process stability, higher average NH4+-N concentration in the later stage of fermentation, reduced COD (p < 0.05), and increased biodegradability of lignocelluloses (p < 0.01), especially cellulose (p < 0.05) and lignin (p < 0.01). Meanwhile, the addition of Zn promoted the coenzyme M activity (p < 0.05), and increased the absolute abundance of Methanothermobacter. The bacteria communities during the fermentation process were responsible for the degradation of lignocelluloses. The results demonstrated that the addition of appropriate Zn into the Cd-containing fermentation system enhanced the efficiency of anaerobic fermentation and utilization of biowaste.

Process Analysis of Anaerobic Fermentation Exposure to Metal Mixtures.

Anaerobic fermentation is a cost-effective biowaste disposal approach. During fermentation, microorganisms require a trace amount of metals for optimal growth and performance. This study investigated the effects of metal mixtures on biogas properties, process stability, substrate degradation, enzyme activity, and microbial communities during anaerobic fermentation. The addition of iron (Fe), nickel (Ni), and zinc (Zn) into a copper (Cu)-stressed fermentation system resulted in higher cumulative biogas yields, ammonia nitrogen (NH4+-N) concentrations and coenzyme F420 activities. Ni and Zn addition enhanced process stability and acetate utilization. The addition of these metals also improved and brought forward the peak daily biogas yields as well as increased CH4 content to 88.94 and 86.58%, respectively. Adding Zn into the Cu-stressed system improved the abundance of Defluviitoga, Fibrobacter and Methanothermobacter, the degradation of cellulose, and the transformation of CO2 to CH4. The bacterial and archaeal communities were responsible for the degradation of lignocelluloses and CH4 production during the fermentation process. This study supports the reutilization of heavy metal-contaminated biowaste and provides references for further research on heavy metals impacted anaerobic fermentation.

Palladium-Catalyzed Dialkylation of C-C Triple Bonds: Access to Multi-Functionalized Indenes.

A palladium-catalyzed dialkylation of 1,3-dien-5-ynes was developed using alkenyl double bonds as the initiator and terminator for the synthesis of functionalized indene derivatives. The reactions were performed under mild reaction conditions, affording the corresponding multi-substituted indene derivatives in high efficiency via unprecedented 5- endo cyclization and alkylation processes. It was found that the substituent location at the alkenyl double bond was essential for the chemoselective synthesis of the indene and naphthalene derivatives, respectively.

An ontology-driven, case-based clinical decision support model for removable partial denture design.

We present the initial work toward developing a clinical decision support model for specific design of removable partial dentures (RPDs) in dentistry. We developed an ontological paradigm to represent knowledge of a patient's oral conditions and denture component parts. During the case-based reasoning process, a cosine similarity algorithm was applied to calculate similarity values between input patients and standard ontology cases. A group of designs from the most similar cases were output as the final results. To evaluate this model, the output designs of RPDs for 104 randomly selected patients were compared with those selected by professionals. An area under the curve of the receiver operating characteristic (AUC-ROC) was created by plotting true-positive rates against the false-positive rate at various threshold settings. The precision at position 5 of the retrieved cases was 0.67 and at the top of the curve it was 0.96, both of which are very high. The mean average of precision (MAP) was 0.61 and the normalized discounted cumulative gain (NDCG) was 0.74 both of which confirmed the efficient performance of our model. All the metrics demonstrated the efficiency of our model. This methodology merits further research development to match clinical applications for designing RPDs. This paper is organized as follows. After the introduction and description of the basis for the paper, the evaluation and results are presented in Section 2. Section 3 provides a discussion of the methodology and results. Section 4 describes the details of the ontology, similarity algorithm, and application.