ECOTOXICOLOGY AND ENVIRONMENTAL SAFETY
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2024
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IF 6.2
Bu-Da Su,Xiao-Meng Li,Zhi-Wei Huang,Yue Wang,Jia Shao,Yan-Yan Xu,Le-Xin Shu,Yu-Bo Li
Ochratoxin A (OTA) is a common fungal toxin frequently detected in food and human plasma samples. Currently, the physiologically based toxicokinetic (PBTK) model plays an active role in dose translation and can improve and enhance the risk assessment of toxins. In this study, the PBTK model of OTA in rats and humans was established based on knowledge of OTA-specific absorption, distribution, metabolism, and excretion (ADME) in order to better explain the disposition of OTA in humans and the discrepancies with other species. The models were calibrated and optimized using the available kinetic and toxicokinetic (TK) data, and independent test datasets were used for model evaluation. Subsequently, sensitivity analyses and population simulations were performed to characterize the extent to which variations in physiological and specific chemical parameters affected the model output. Finally, the constructed models were used for dose extrapolation of OTA, including the rat-to-human dose adjustment factor (DAF) and the human exposure conversion factor (ECF). The results showed that the unbound fraction (F up ) of OTA in plasma of rat and human was 0.02–0.04% and 0.13–4.21%, respectively. In vitro experiments, the maximum enzyme velocity (V max ) and Michaelis–Menten constant (K m ) of OTA in rat and human liver microsomes were 3.86 and 78.17 μg/g min −1 , 0.46 and 4.108 μg/mL, respectively. The predicted results of the model were in good agreement with the observed data, and the models in rats and humans were verified. The PBTK model derived a DAF of 0.1081 between rats and humans, whereas the ECF was 2.03. The established PBTK model can be used to estimate short- or long-term OTA exposure levels in rats and humans, with the capacity for dose translation of OTA to provide the underlying data for risk assessment of OTA.
研究领域:
毒理学
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药代动力学
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食品安全
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生物医学工程
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产品关联
DOI
PubMed
高影响因子
PROTEIN EXPRESSION AND PURIFICATION
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2022
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IF 2.025
Xiaowen Wang,Sibo Chen,Lin Fan,Hao Liu,Si Chen,Xianfeng Ye,Zhoukun Li,Zhongli Cui,Yan Huang
A new glycoside hydrolase family 2 (GH2) β-galactosidase encoding gene galM was cloned from Microvirga sp. strain MC18 and overexpressed in Escherichia coli . The recombinant β-galactosidase GalM showed optimal activity at pH 7.0 and 50 °C, with a stability at pH 6.0–9.0 and 20–40 °C, which are conditions suitable for the diary environment. The K m and V max values for o-nitrophenyl-β- d -galactopyranoside (oNPG) were 1.30 mmol/L and 15.974 μmol/(min·mg), respectively. GalM showed low product inhibition by galactose with a K i of 73.18 mM and high tolerance for glucose that 86.5% activity retained in the presence of 500 mM glucose. It was also stable and active in 20% of methanol, ethanol and isopropanol. In addition, the enzyme activity of GalM was activated significantly over 0–2 mol/L NaCl (1.6–4.3 fold). These favorable properties make GalM a potential candidate for the industrial application.
研究领域:
生物化学
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酶学
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生物技术
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DOI
PubMed
高影响因子
TALANTA
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2019
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IF 4.916
Wen Zheng,Dongjun Shen,Yadi Pan,Danyang Yi,Yijuan Long,Huzhi Zheng
The peroxidase-like activity of ficin is relatively low, which limits its application. It was found that thiol groups of ficin could inhibit its peroxidase-like activity. So, two procedures, i.e., direct blocking with N -ethylmaleimide (NEM), or using tris (2-carboxyethyl) phosphine hydrochloride (TCEP) to interrupt disulfide bonds then blocking thiol groups with NEM, were applied to block thiol groups of ficin, ficin-NEM (ficin-N) and ficin-TCEP-NEM (ficin-TN) were produced, respectively. The blocking of thiol groups accelerated the peroxidase activity dramatically. The peroxidase catalytic activity of ficin-N and ficin-TN toward the peroxidase substrate 3,3′,5,5′-tetramethylbenzidine (TMB) oxidation by H 2 O 2 was about 2.5-fold and 5-fold increase compared with ficin, respectively, which accompanied a color change from colorless to blue and followed classic Michaelis-Menten model. The kinetic parameters indicated that higher affinity of ficin-N ( K m = 0.31) and ficin-TN ( K m = 0.39) to H 2 O 2 compared with ficin ( K m = 0.58), and ficin-TN had the highest K cat which increased by 6.5 times and 4.5 times for TMB and H 2 O 2 , respectively. According to these findings, a colorimetric method with high sensitivity for the detection of biothiols was developed due to sulfhydryl compounds inhibited the peroxidase activity of ficin. Comparing with ficin and ficin-N, ficin-TN had the widest detection range (0.01–16 μM) and the lowest detection limit (3 nM). The practical applications of ficin-TN for biothiol determination in human serum samples have been demonstrated with satisfactory results. Ficin-N and ficin-TN are promising to apply to the bioanalysis.
研究领域:
生物化学
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分析化学
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生物技术
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DOI
PubMed
高影响因子
JOURNAL OF CHROMATOGRAPHY A
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2023
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IF 4.1
Wen Li,Xinyue Cui,Zilin Chen
An online capillary electrophoresis method based-lipase immobilized enzyme microreactor was developed for lipase kinetic study and inhibitor screening from compounds from natural products. Zeolitic Imidazolate Framework-8 (ZIF-8) has the advantages of large pore size, mild synthesis conditions and good biocompatibility. Lipase was immobilized on the inner wall of capillary with the help of the metal-organic framework ZIF-8. The results of electron microscopy showed that lipase could be aggregated and fixed on the inner wall of capillary by ZIF-8. After the experimental conditions including electrophoretic separation and enzymatic reaction were optimized, the baseline separation of substrate p-nitrophenyl acetate (pNPA) and product p-nitrophenol (pNP) was achieved within 3 min. The immobilized enzyme microreactor showed good repeatability and stability, and the determined Michaelis-Menten constant ( K m ) of lipase was 2.75 mM, which was lower than the kinetic constant determined in off-line reaction, indicating that the immobilized enzyme had a high affinity with the substrate. In addition, the IC 50 value of the positive control compound orlistat on lipase inhibition was 7.26 nM, which was consistent with the literature. Then the inhibitory activity of 10 compounds from natural products on lipase was evaluated by the ZIF-8-IMER. Among them, 7 compounds including baicalein, luteolin, epicatechin gallic acid, and chlorogenic acid, had a certain inhibitory effect on lipase. The molecular docking technology proved the interaction between the enzyme and the screened inhibitor, which provides a new method for the screening of lipase inhibitors.
研究领域:
生物化学
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分析化学
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药学
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酶学
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产品关联
DOI
PubMed
高影响因子
ANALYTICA CHIMICA ACTA
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2023
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IF 6.2
Yaoyu Tan,Mengyao Qi,Huan Jiang,Baihui Wang,Xia Zhang
The level of uric acid is crucial to human health. Octahedral oxygen vacancy MnCo 2 O 4 /Ag (V O -MnCo 2 O 4 /Ag) nanozyme was successfully prepared by simple hydrothermal, calcination and self-reduction methods. V O -MnCo 2 O 4 /Ag nanozyme is rich in Mn 2+ /Mn 3+ and C O 2+ /C O 3+ redox electron pairs, large specific surface area and oxygen vacancies. V O -MnCo 2 O 4 /Ag nanozyme showed high uricase-like activity and peroxidase-like activity. At the same time, the SERS signal of the detected molecule could be significantly enhanced after the catalytic reaction of the V O -MnCo 2 O 4 /Ag nanozyme. The K m values of V O -MnCo 2 O 4 /Ag nanozyme for H 2 O 2 and TMB were 0.04 mM and 0.027 mM respectively. Based on the uric acid oxidase-like and peroxidase-like activities of V O -MnCo 2 O 4 /Ag, we developed a label-free, sensitive, and reliable SERS uric acid detection system. The detection linear range of uric acid is 0.01 μM–1000 μM and the detection of limit is 7.8 × 10 −9 M. The results show that the sensing system has good accuracy, sensitivity, selectivity , and stability. It can be applied to the determination of samples under different conditions. This study provides profound insights into the design of enzyme-like activity regulation and SERS properties regulation of nanozymes, provides guidance for the study of reaction kinetics and catalytic mechanism of nanozymes, and has broad application prospects in the field of nanozymes and SERS sensing analysis.
研究领域:
纳米技术
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分析化学
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生物化学
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材料科学
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医学诊断
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产品关联
DOI
PubMed
高影响因子
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