Enrichment of trace bioactive constituents and metabolites from complex biological samples is challenging.This study presented a one-pot synthesis of magnetic polydopamine nanoparticles(Fe3O4@-SiO2@PDA NPs)with multip...Enrichment of trace bioactive constituents and metabolites from complex biological samples is challenging.This study presented a one-pot synthesis of magnetic polydopamine nanoparticles(Fe3O4@-SiO2@PDA NPs)with multiple recognition sites for the magnetic dispersive solid-phase extraction(MDSPE)of ginsenosides from rat plasma treated with white ginseng.The extracted ginsenosides were characterized by combining an ultra-high-performance liquid chromatography coupled to a highresolution mass spectrometry with supplemental UNIFI libraries.Response surface methodology was statistically used to optimize the extraction procedure of the ginsenosides.The reusability of Fe3O4@-SiO2@PDA NPs was also examined and the results showed that the recovery rate exceeded 80%after recycling 6 times.Furthermore,the proposed method showed greater enrichment efficiency and could rapidly determine and characterize 23 ginsenoside prototypes and metabolites from plasma.In comparison,conventional methanol method can only detect 8 ginsenosides from the same plasma samples.The proposed approach can provide methodological reference for the trace determination and characterization of different bioactive ingredients and metabolites of traditional Chinese medicines and food.展开更多
Magnetic nanoparticles(MNPs) are widely used for the immobilization of enzyme owing to the unique properties such as good biocompatibility and rapid separation. Herein, we used Fe_3O_4 magnetic nanoparticles(Fe_3O_4 M...Magnetic nanoparticles(MNPs) are widely used for the immobilization of enzyme owing to the unique properties such as good biocompatibility and rapid separation. Herein, we used Fe_3O_4 magnetic nanoparticles(Fe_3O_4 MNPs) as the carrier core with(3-aminopropyl)triethoxysilane(APTES)modification by our approach, in which a-glucosidase was stereoscopically immobilized on the surface of Fe_3O_4 MNPs via covalent binding. The result of immobilization was characterized by scanning electron microscope(SEM) and fourier transform-infrared spectroscopy(FT-IR). Then we optimized some key parameters of the immobilization reaction, including the ratio of MNPs to enzyme, GA concentration,crosslinking time and immobilization time. Moreover, under the optimal conditions, pH tolerance,thermo stability and reusability of the immobilized enzyme were investigated and compared with the free one. In order to evaluate the change of the affinity of the enzyme to its specific substrate after immobilization, the Michaelis-Menten constant(K_m) was also studied. Finally, the immobilized α-glucosidase combining with high performance liquid chromatography-tandem mass spectrometry technique(HPLC-MS/MS) was applied to screen and identify eight inhibitors from Polygonum cuspidatum extract. These results indicated that the established method had the broad prospects for biotechnological applications.展开更多
Aromatic carboxylic acids(ACAs) may be as transformed key metabolites via gut microbiome for playing better pharmacological effects. However, it's rare to achieve high-specificity, high-sensitivity, and highthroug...Aromatic carboxylic acids(ACAs) may be as transformed key metabolites via gut microbiome for playing better pharmacological effects. However, it's rare to achieve high-specificity, high-sensitivity, and highthroughput detection simultaneously, especially, for tracing trace ACAs in gut microbiome. In this work,firstly, a novel dual-template and double-shelled molecularly imprinted 96-well microplates(DDMIPs)was designed and amplified signal for p-hydroxybenzoic acid(PBA) and 3,4,5-trimethoxycinnamic acid(TMA). Additionally, the DDMIPs and a stable isotope labeling derivatization(SILD) method combined with the ultra-high performance liquid chromatography triple quadrupole tandem mass spectrometry(UHPLC-TQ MS) was firstly stepwise integrated, achieving high-effective, high-sensitive, and highthroughput study of gut microbiome metabolism. The whole strategy showed lower limits of detections(LODs) up to 1000 folds than the traditional method, and revealed a more real metabolism-time profile of PBA and TMA by 3-step signal amplification. The platform also laid the foundation for fast, simple,high-selective, high-effective, and high-throughput metabolism and pharmacological research.展开更多
基金This work was supported by grants from the National Natural Science Foundation of China Key Program(NO.81530094)General Program(NO.81573574,81873193)the Science and Technology Development Project of Jilin Province(20190201283JC).
文摘Enrichment of trace bioactive constituents and metabolites from complex biological samples is challenging.This study presented a one-pot synthesis of magnetic polydopamine nanoparticles(Fe3O4@-SiO2@PDA NPs)with multiple recognition sites for the magnetic dispersive solid-phase extraction(MDSPE)of ginsenosides from rat plasma treated with white ginseng.The extracted ginsenosides were characterized by combining an ultra-high-performance liquid chromatography coupled to a highresolution mass spectrometry with supplemental UNIFI libraries.Response surface methodology was statistically used to optimize the extraction procedure of the ginsenosides.The reusability of Fe3O4@-SiO2@PDA NPs was also examined and the results showed that the recovery rate exceeded 80%after recycling 6 times.Furthermore,the proposed method showed greater enrichment efficiency and could rapidly determine and characterize 23 ginsenoside prototypes and metabolites from plasma.In comparison,conventional methanol method can only detect 8 ginsenosides from the same plasma samples.The proposed approach can provide methodological reference for the trace determination and characterization of different bioactive ingredients and metabolites of traditional Chinese medicines and food.
基金funded by the National Natural Science Foundation of China(Nos.81473537,81773690,21673219)
文摘Magnetic nanoparticles(MNPs) are widely used for the immobilization of enzyme owing to the unique properties such as good biocompatibility and rapid separation. Herein, we used Fe_3O_4 magnetic nanoparticles(Fe_3O_4 MNPs) as the carrier core with(3-aminopropyl)triethoxysilane(APTES)modification by our approach, in which a-glucosidase was stereoscopically immobilized on the surface of Fe_3O_4 MNPs via covalent binding. The result of immobilization was characterized by scanning electron microscope(SEM) and fourier transform-infrared spectroscopy(FT-IR). Then we optimized some key parameters of the immobilization reaction, including the ratio of MNPs to enzyme, GA concentration,crosslinking time and immobilization time. Moreover, under the optimal conditions, pH tolerance,thermo stability and reusability of the immobilized enzyme were investigated and compared with the free one. In order to evaluate the change of the affinity of the enzyme to its specific substrate after immobilization, the Michaelis-Menten constant(K_m) was also studied. Finally, the immobilized α-glucosidase combining with high performance liquid chromatography-tandem mass spectrometry technique(HPLC-MS/MS) was applied to screen and identify eight inhibitors from Polygonum cuspidatum extract. These results indicated that the established method had the broad prospects for biotechnological applications.
基金the National Natural Science Foundation of China (Nos. 82073973, 81872969)the Jilin Provincial Industrial Innovation Special Fund Project (No. 20200703015ZP)the Science and the Youth Innovation Promotion Association of CAS (No. 2019227)。
文摘Aromatic carboxylic acids(ACAs) may be as transformed key metabolites via gut microbiome for playing better pharmacological effects. However, it's rare to achieve high-specificity, high-sensitivity, and highthroughput detection simultaneously, especially, for tracing trace ACAs in gut microbiome. In this work,firstly, a novel dual-template and double-shelled molecularly imprinted 96-well microplates(DDMIPs)was designed and amplified signal for p-hydroxybenzoic acid(PBA) and 3,4,5-trimethoxycinnamic acid(TMA). Additionally, the DDMIPs and a stable isotope labeling derivatization(SILD) method combined with the ultra-high performance liquid chromatography triple quadrupole tandem mass spectrometry(UHPLC-TQ MS) was firstly stepwise integrated, achieving high-effective, high-sensitive, and highthroughput study of gut microbiome metabolism. The whole strategy showed lower limits of detections(LODs) up to 1000 folds than the traditional method, and revealed a more real metabolism-time profile of PBA and TMA by 3-step signal amplification. The platform also laid the foundation for fast, simple,high-selective, high-effective, and high-throughput metabolism and pharmacological research.
