BIOPROCESS AND BIOSYSTEMS ENGINEERING
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2023
·Vol. 46
·期 9
·1293-1302
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IF 3.8
Duan Zhiwen,Wang Yaowu,Ouyang Bin,Wang Pu
Ionic liquids (ILs) which synthesized from bio-renewable materials have recently attracted much attention for their applications in biocatalysis. Ethyl ( R )-3-hydroxybutyrate (( R )-EHB) as a versatile chiral intermediate is of great interest in pharmaceutical synthesis. This study focuses on evaluating the performances of choline chloride (ChCl)-based and tetramethylammonium (TMA)-based neoteric ILs in the efficient synthesis of ( R )-EHB via the bioreduction of ethyl acetoacetate (EAA) at high substrate loading by recombinant Escherichia coli cells. It was found that choline chloride/glutathione (ChCl/GSH, molar ratio 1:1) and tetramethylammonium/cysteine ([TMA][Cys], molar ratio 1:1) as eco-friendly ILs not only enhanced the solubility of water-insoluble EAA in the aqueous buffer system, but also appropriately improved the membrane permeability of recombinant E. coli cells, thus boosting catalytic reduction efficiency of EAA to ( R )-EHB. In the developed ChCl/GSH- or [TMA][Cys]-buffer systems, the space-time yields of ( R )-EHB achieved 754.9 g/L/d and 726.3 g/L/d, respectively, which are much higher than neat aqueous buffer system (537.2 g/L/d space-time yield). Meanwhile, positive results have also been demonstrated in the bioreduction of other prochiral ketones in the established IL-buffer systems. This work exhibits an efficient bioprocess for ( R )-EHB synthesis under 325 g/L (2.5 M) substrate loading, and provides promising ChCl/GSH- and [TMA][Cys]-buffer systems employed in the biocatalysis for hydrophobic substrate.
研究领域:
生物化学
·
工业微生物学
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DOI
PubMed
高影响因子
ORGANIC PROCESS RESEARCH & DEVELOPMENT
·
2022
·
IF 3.858
Ling He,Wenjie Ye,Youyu Xie,Qinghai Liu,Hualei Wang,Dongzhi Wei
Bioreductions catalyzed by ketoreductases play an important role in the synthesis of chiral alcohols. However, the synthesis of (R)-2-chloro-1-(3,4-difluorophenyl)ethanol (24b), an important chiral intermediate for the synthesis of the anticoagulant ticagrelor, poses significant challenges in terms of high substrate concentration requirements that limit its production. In this study, a novel NADH-dependent, short-chain dehydrogenase, PpKR8, from Paraburkholderia phymatum STM815, exhibited excellent enantioselectivity and high activity for the production of (R)-CFPL (24b) through the reduction of 2-chloro-1-(3,4-difluorophenyl)ethanone (24a). The coexpression of PpKR8 and glucose dehydrogenase from Bacillus subtilis in Escherichia coli allowed up to 300 g/L (1.57 M) CFPO (24a) to be completely converted into 24b with 99.9% enantiomeric excess and a high space–time yield (728 g/(L day)). Substrate specificity assays demonstrated a broad substrate spectrum for PpKR8, which included 35 α/β-ketoesters, aromatic ketones, and heterocyclic ketones. Moreover, three additional optically pure chiral alcohols that are used as important drug intermediates were synthesized at high substrate concentrations (150–330 g/L), demonstrating the excellent industrial potential of PpKR8-mediated bioreductions.
研究领域:
生物化学
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酶工程
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制药工业
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DOI
高影响因子
CHIRALITY
·
2018
·Vol. 30
·期 12
·1277-1286
·
IF 1.833
Lu-Ning Sun,Yi-Wen Shen,Yu-Wen Ying,Duo Li,Teng-Fei Li,Xue-Hui Zhang,Ping Zhao,Li Ding,Yong-Qing Wang
Rabeprazole is an effective proton pump inhibitor to treat acid-related diseases. To achieve the simultaneous determination of rabeprazole enantiomers in human plasma, a chiral LC-MS/MS method was developed and validated. Acetonitrile including 0.1% ammonium were used as protein precipitating agent. Analytes were separated within 8 minutes on a Chiralpak IC column (4.6 mm × 150 mm, 5 μm). The mobile phase was 10 mM ammonium acetate including 0.2% acetic acid-acetonitrile (35:65, v/v). An API 4000 mass spectrometer was used as detector for the analysis, and the multiple reactions monitoring transitions of m/z 360.1 → 242.2 and 346.1 → 198.1 were opted for quantifying rabeprazole enantiomers and internal standard. Matrix effects were not apparent for each enantiomer and internal standard (esomeprazole), the calibration curves were linear over the concentration of 0.500 to 400 ng·mL −1 , the intra-run precisions were below 5.4%, the inter-run precisions were below 9.9%, and the accuracy was between −9.2% and 9.3%. There was no chiral inversion observed during sample storage, preparation procedure, and analysis, demonstrating that analytes were stable in this study. This method was applied to the stereoselective pharmacokinetic study of (R)-(+)- and (S)-(−)-rabeprazole after oral administration of 10-mg rabeprazole sodium enteric-coated tablet in healthy Chinese subjects.
研究领域:
药物分析
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临床药理学
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生物化学
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产品关联
DOI
PubMed
高影响因子
Catalysts
·
2021
·Vol. 11
·期 9
·1126
·
IF 4.146
Qingman Li,Qihua Jiang,Pengcheng Gu,Lianju Ma,Yiwu Wang
Ni2+-NTA-boosted magnetic porous silica nanoparticles (Ni@MSN) to serve as ideal support for bi-functional enzyme were fabricated for the first time. The versatility of this support was validated by one-step purification and immobilization of bi-functional enzyme MLG consisting of 3-Quinuclidinone reductase and glucose dehydrogenase, which can simultaneously catalyze both carbonyl reduction and cofactor regeneration, to fabricate an artificial bi-functional nanobiocatalyst (namely, MLG-Ni@MSN). The enzyme loading of 71.7 mg/g support and 92.7% immobilization efficiency were obtained. Moreover, the immobilized MLG showed wider pH and temperature tolerance and greater storage stability than free MLG under the same conditions. The nanosystem was employed as biocatalyst to accomplish the 3-quinuclidinone (70 g/L) to (R)-3-quinuclidinol biotransformation in 100% conversion yield with >99% selectivity within 6 h and simultaneous cofactor regeneration. Furthermore, the immobilized MLG retained up to 80.3% (carbonyl reduction) and 78.0% (cofactor regeneration) of the initial activity after being recycled eight times. In addition, the MLG-Ni@MSN system exhibited almost no enzyme leaching during biotransformation and recycling. Therefore, we have reason to believe that the Ni@MSN support gave great promise for constructing a new biocatalytic nanosystem with multifunctional enzymes to achieve some other complex bioconversions.Keywords:porous silica nanoparticles;immobilized enzyme;nanosystem;cofactor regeneration;(R)-3-quinuclidinol;biocatalysts’ recycling
研究领域:
生物工程
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酶学
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纳米技术
·
合成生物学
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产品关联
DOI
高影响因子
CHEMICAL ENGINEERING JOURNAL
·
2023
·
IF 15.1
Xinman Hu,Fang-Yuan Chen,Jie Ding,Zihe Zhai,Huang Yang,Liyin Shen,Yang Zhu,Dong-Sheng Guo,Changyou Gao
The clinically restored blood supply post myocardial infarction (MI) will break the physiological and metabolic balance of the ischemic myocardial region, leading to a new injury known as myocardial ischemia/reperfusion (I/R) injury. Abundant oxygen brought by the fresh blood would trigger the damaged myocardium to produce excessive reactive oxygen species (ROS) and thereby inflammation along with the recruitment and infiltration of activated neutrophils and macrophages. Inspired by the active targeting process of inflammatory cells, a novel “neutrophil-like” nanoassembly (At@NQA) was prepared by modifying neutrophil membrane fragments onto At@QA NPs containing 1:1 (molar ratio) calixarene molecule (QA) and anti-inflammatory and anti-fibrotic atorvastatin calcium (At). Tail vein injection of the At@NQA could efficiently target the damaged myocardium, inhibit oxidative damage and inflammation, reduce myocardial fibrosis, increase myocardial troponin T (cTnT) and connexin 43 (Cx43) expression, and thus improve cardiac functions effectively.
研究领域:
纳米技术
·
心脏病学
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药学
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生物医学工程
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产品关联
DOI
高影响因子
Polymer Chemistry
·
2016
·Vol. 7
·期 22
·3756-3765
·
IF 5.687
Mingguang Yu,Jianbo Tan,Jianwen Yang,Zhaohua Zeng
In a common RAFT dispersion polymerization induced self-assembly (PISA) formulation, R-type macro-RAFT agents with a solvophilic chain attached at the leaving group side are normally used to mediate the polymerization, meanwhile Z-type macro-RAFT agents with a solvophilic chain attached at the Z-group side are seldom employed. In this paper, we have tried to use a Z-type macro-RAFT agent (mPEG113-BTPA) to mediate RAFT dispersion polymerization of styrene, and found that the process exhibited a poorer PISA tendency and weaker molecular weight control. In comparison, we have conducted the same process mediated with an R-type macro-RAFT agent (mPEG113-DDMAT), and found that the formulation exhibited a typical PISA process, and nanoparticles were produced in situ. Further experiments suggest that the location of RAFT groups plays a key role for the PISA process in RAFT dispersion polymerization. The RAFT groups will be embedded in the produced particles when an R-type macro-RAFT agent is employed, but locate on the particle surface in the case of the Z-type macro-RAFT agent. We have tried to employ the RAFT groups on the surface of the mPEG113-BTPA stabilized particles to modify the particle surface, and successfully removed the solvophilic block via breaking the RAFT group, and generated the poly(dimethylacrylamide) block by surface-initiated RAFT polymerization. Based on the above mechanism perspective, we have designed a preliminary experiment, and found that well-defined monodisperse nanospheres could be prepared by heterogeneous RAFT dispersion polymerization mediated with a Z-type macro-RAFT agent-based block copolymer which was synthesized via pre-polymerization of styrene in 1,4-dioxane mediated with mPEG113-BTPA.
研究领域:
高分子化学
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胶体与界面科学
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纳米材料
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可控自由基聚合
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自组装
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药物递送系统
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DOI
高影响因子
JOURNAL OF BIOTECHNOLOGY
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2015
·
IF 2.871
Yu-Cai He,Zhi-Cheng Tao,Yun Ding,Dan-Ping Zhang,Yin-Qin Wu,Yun Lu,Feng Liu,Yu-Feng Xue,Cheng Wang,Jian-He Xu
To avoid adding NAD + and effectively transform ethyl 4-chloro-3-oxobutanoate, the mixture of l -glutamine (200 mM) and d -xylose (250 mM) was added into in n -butyl acetate–water (10:90, v/v) biphasic system instead of NAD + for increasing the biocatalytic efficiency. To further improve the synthesis of optically pure ethyl ( R )-4-chloro-3-hydroxybutanoate (>99% ee ), β-cyclodextrin was also added into this reaction media, and ethyl ( R )-4-chloro-3-hydroxybutanoate (>99% ee ) could be effectively synthesized from 800 mM ethyl 4-chloro-3-oxobutanoate in the yield of 100% by whole-cells of recombinant E. coli CCZU-A13. Finally, the possible mechanism for improving the reductase activity by supplementation of l -glutamine, d -xylose and β-CD was proposed. In conclusion, this strategy has high potential for the effective biosynthesis of ethyl ( R )-4-chloro-3-hydroxybutanoate (>99% ee ).
研究领域:
生物化学
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酶工程
·
合成生物学
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DOI
高影响因子
Journal of Natural Medicines
·
2025
·
IF 2.5
Zhou Yue,Li Yiran,Yang Zhenzhong,Zhao Lu,Wang Yule
Blood–brain barrier (BBB) dysfunction is a well-established pathological phenotype of ischemic stroke, and targeting BBB integrity has emerged as a promising therapeutic strategy. Danshen-Chuanxiong formula (DS-CX), an effective herbal combination against ischemic stroke, has demonstrated regulatory effects on the BBB at various stages of ischemic stroke. However, its specific BBB-protective components and underlying molecular mechanisms remain unclear. Recent advances in multicellular self-assembled BBB spheroids have shown distinct advantages in disease modeling and drug discovery, offering a novel approach to address these questions. To simulate ischemic stroke-induced BBB dysfunction, we developed an oxygen–glucose deprivation/reoxygenation (OGD/R)-induced BBB disruption model using multicellular spheroids. To identify the effective substances of DS-CX responsible for BBB protection, we conducted a multi-parametric evaluation to assess BBB permeability, tight junctions, cell viability, reactive oxygen species (ROS) levels, inflammatory markers, and apoptotic phenotypes using high-content imaging. Further immunofluorescence and transcription analyses were performed to elucidate the BBB-protective mechanisms of DS-CX and its active components. Similar to the overall effects of DS-CX on BBB protection, preliminary screening fortunately found that both protocatechuic acid, ferulic acid, and senkyunolide I significantly reduced OGD/R-induced leakage, and upregulated the protein and mRNA levels of ZO-1 and Claudin-5 in BBB spheroids. Further multi-phenotypic assessments manifested that DS-CX and its active compounds effectively improved cell survival, reduced ROS production, inhibited inflammation, and decreased apoptosis, compared to the damaged BBB spheroids without drug intervention. Molecular experiments showed that DS-CX and its active constituents not only rescued the abnormal protein levels of pivotal targets related to oxidative stress (HO-1), inflammation (MMP-9, TLR-4), and apoptosis (Caspase-3, Bax, Bcl-2) in OGD/R-treated BBB spheroids, but also normalized the dysregulated mRNA levels of vWF, HO-1, MMP-9, TLR-4, TNF-α, IL-6, IL-1β, and IL-18 caused by OGD/R stimulation. Collectively, the present work successfully identified protocatechuic acid, ferulic acid, and senkyunolide I as key BBB-protective components of DS-CX against ischemic stroke. These compounds likely exert their therapeutic effects through multi-target regulation of oxidative stress, inflammation, and apoptosis. Our findings provide a novel spheroid-based multi-parametric screening approach for discovering BBB-targeted therapies in ischemic stroke. Graphical abstract
研究领域:
神经生物学
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药理学
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中医学
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分子生物学
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转化医学
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药物筛选
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产品关联
DOI
PubMed
高影响因子
BIOMEDICINE & PHARMACOTHERAPY
·
2021
·
IF 6.53
Yi-yu Liang,Kun-wei Li,Feng-jv Niu,Ying Li,Hao-cheng Wei,Yun-lei Dai,Yun-yu Wang,Chang-zheng Zhou,Xin-huan Wan
Objective To investigate the antiviral effect of Salvia plebeia R. Br. polysaccharides (SPP) against RSV and underlying mechanisms. Methods SPP was extracted via alcohol-precipitation method and extract was separated into various fractions using ultrafiltration method. The polysaccharide content was determined using UV-Vis. Antiviral effect of SPP and fractions was measured using MTT method and Reed-Muench method. Sixty Balb/c mice were randomly divided into 6 groups, and received either Ribavirin or SPP. Their body weight and food intake were recorded every day throughout the experiment period. The lung index inhibition ratio and pulmonary virus titer were determined followed by the histological analysis of lungs. Furthermore, time-of-addition and effective stage analysis were carried out to determine the mechanism of action. The TLR-3 and TLR-4 levels in the lungs were determined using qRT-PCR. The levels of IFN-γ, IL-2 and TNF-α in serum were determined using ELISA. Results The SPP content is 4.396%. SPP has shown a good anti-RSV effect both in vitro (TI = 123.041) and in vivo models. The antiviral activity of fractions with molecular weight ≥ 10,000 is found to possess more potent antiviral activity than other fractions. SPP inhibits the RSV proliferation and reduces the lung lesions induced by RSV. The mechanism of action involves the inhibition of TLR-3 and TLR-4 in lungs, up-regulation of IFN-γ and IL-2, and down-regulation of TNF-α in serum. It is also shown to improve the body’s immune function. Conclusion SPP has a potential to treat diseases caused by RSV.
研究领域:
药理学
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免疫学
·
病毒学
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产品关联
DOI
PubMed
高影响因子
CHEMICAL ENGINEERING JOURNAL
·
2019
·
IF 8.355
Kiran Gupta,Baoling Yuan,Chen Chen,Naresh Varnakavi,Ming-Lai Fu
Novel composite of immobilized layered K 2x Mn x Sn 3−x S 6 (x = 0.5–0.95) (KMS-1) on the surface of reduced graphene oxide (r-GO), KMS-1/r-GO (KGO), has been fabricated by employing hydrothermal technique and used for the removal of radioactive Cs + and Sr 2+ ions from aqueous solutions. The KGO was characterized thoroughly with different characteristic techniques and found that the laminar KMS-1 nanoplate were immobilized on the surface of r-GO nanosheets, forming a highly porous stacked structure. Batch studies have been done for obtaining optimal experimental parameters and investigating the effect of adsorption time, initial concentration of pollutant, pH of solution, temperature, and competing anions on the removal properties. At room temperature, the maximum adsorption capacity of KGO was 338.18 mg·g −1 and 220.12 mg·g −1 for both Cs + and Sr 2+ , respectively. Langmuir isotherm and pseudo-second-order kinetic models were fitted well with experimental data of removed Cs + and Sr 2+ . Adsorption showed good selectivity of composite towards Cs + and Sr 2+ in the presence of co-existing ions (Ca 2+ and Mg 2+ ). The proposed adsorption mechanism involved ion exchange through the presence of K + ions in KMS-1 and formation of surface ionic interactions between r-GO and ions through COO X or S X (X denoted Cs + and Sr 2+ ). This study highlighted the facile applicability of novel material KGO and could be used as a new adsorbent for the removal of radioactive ions.
研究领域:
环境科学
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材料科学
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产品关联
DOI
高影响因子
Microbial Biotechnology
·
2021
·Vol. 15
·期 5
·1486-1498
·
IF 5.813
Baoqi Zhang,Han Du,Yanqiu Zheng,Jiale Sun,Yu Shen,Jinping Lin,Dongzhi Wei
Summary Bioproduction of optical pure ( R )-citronellal from ( E /Z)-citral at high substrate loading remains challenging. Low catalytic efficiency of ( R )-stereoselective ene reductases towards crude citral mixture is one of the major bottlenecks. Herein, a structure-based engineering strategy was adopted to enhance the catalytic efficiency and stereoselectivity of an ene reductase (OYE2p) from Saccharomyces cerevisiae YJM1341 towards ( E / Z )-citral. On basis of homologous modelling, molecular docking analysis and alanine scanning at the binding pocket of OYE2p, a mutant Y84A was obtained with simultaneous increase in catalytic efficiency and stereoselectivity. Furthermore, site-saturation mutagenesis of Y84 yielded seven mutants with improved activity and stereoselectivity in the ( E / Z )-citral reduction. Among them, the variant Y84V exhibited an 18.3% and 71.3% rise in catalytic efficiency ( k cat / K m ) for ( Z )-citral and ( E )-citral respectively. Meanwhile, the stereoselectivity of Y84V was improved from 89.2% to 98.0% in the reduction in ( E / Z )-citral. The docking analysis and molecular dynamics simulation of OYE2p and its variants revealed that the substitution Y84V enabled ( E )-citral and ( Z )-citral to bind with a smaller distance to the key hydrogen donors at a modified ( R )-selective binding mode. The variant Y84V was then co-expressed with glucose dehydrogenase from Bacillus megaterium in E . coli D4, in which competing prim -alcohol dehydrogenase genes were deleted to prevent the undesired reduction in the aldehyde moiety of citral and citronellal. Employing this biocatalyst, 106 g l −1 ( E / Z )-citral was completely converted into ( R )-citronellal with 95.4% ee value and a high space-time yield of 121.6 g l −1 day −1 . The work highlights the synthetic potential of Y84V, which enabled the highest productivity of ( R )-citronellal from ( E / Z )-citral in high enantiopurity so far.
研究领域:
生物催化
·
酶工程
·
分子生物学
·
生物化学
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2020
·
IF 4.192
Ying Zhou,Rufang Deng,Xinlan Xu,Ziyin Yang
Linalool is abundant in tea leaves and contributes greatly to tea aroma. The two isomers of linalool, (R)-linalool and (S)-linalool, exist in tea leaves. Our study found that (R)-linalool was the minor isomer in nine of Camellia sinensis var. sinensis cultivars. The (R)-linalool synthase of tea plant CsRLIS was identified subsequently. It is a chloroplast-located protein and specifically catalyzes the formation of (R)-linalool in vitro and in vivo. CsRLIS was observed to be a stress-responsive gene and caused the accumulation of internal (R)-linalool during oolong tea manufacture, mechanical wounding, and insect attack. Further study demonstrated that the catalytic efficiency of CsRLIS was much lower than that of (S)-linalool synthase CsSLIS, which might explain the lower (R)-linalool proportion in C. sinensis var. sinensis cultivars. The relative expression levels of CsRLIS and CsSLIS may also affect the (R)-linalool proportions among C. sinensis var. sinensis cultivars. This information will help us understand differential distributions of chiral aroma compounds in tea.
研究领域:
植物学
·
分子生物学
·
芳香化合物代谢
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产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）