Analytical Methods
·
2017
·Vol. 9
·期 35
·5174-5180
·
IF 1.9
Yu-Xiu Yang,Su-Ying Li,Qian Zhang,Hua Chen,Zhi-Ning Xia,Feng-Qing Yang
As a serine protease, thrombin (THR) plays an important role in the coagulation cascade. In this study, a frontal affinity chromatography (FAC) method was developed for the characterization of interactions between immobilized thrombin and phenolic acids, which have potential thrombin inhibitory activity. First, a simple method to immobilize thrombin was developed, where an immobilized artificial membrane (IAM) chromatographic column was used as the carrier material for entrapping THR. Then the frontal analysis of five compounds was performed both on the THR-IAM column and argatroban-THR-IAM column. Finally, the dissociation constants were determined and the values are 2.46, 12.42, 44.93, 76.91 and 100.69 μmol L−1 for gallic acid, protocatechuic acid, ferulic acid, chlorogenic acid and sinapic acid, respectively. In addition, molecular docking was further applied to study the interaction between the compounds and thrombin, and the rank order of docking energy values is similar to that of the dissociation constants of the compounds determined by FAC. The results of the present study demonstrate that the interaction between the compounds and thrombin can be well characterized by FAC experiments along with molecular docking.
研究领域:
生物化学
·
药理学
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DOI
高影响因子
BIOMEDICAL CHROMATOGRAPHY
·
2024
·Vol. 38
·期 9
·e5957
·
IF 1.8
Yu Yi,Xuewang Gong,Mengyuan Cui,Yuting Liang,Jianfeng Mei,Guoqing Ying,Yinfei Wu
Microbial metabolites are an important source of tyrosinase (TYR) inhibitors because of their rich chemical diversity. However, because of the complex metabolic environment of microbial products, it is difficult to rapidly locate and identify natural TYR inhibitors. Affinity-based ligand screening is an important method for capturing active ingredients in complex samples, but ligand immobilization is an important factor affecting the screening process. In this paper, TYR was used as ligand, and the SpyTag/SpyCatcher coupling system was used to rapidly construct affinity chromatography vectors for screening TYR inhibitors and separating active components from complex samples. We successfully expressed SpyTag–TYR fusion protein and SpyCatcher protein, and incubated SpyCatcher protein with epoxy-activated agarose. The SpyTag–TYR protein was spontaneously coupled with SpyCatcher to obtain an affinity chromatography filler for immobilization of TYR, and the performance of the packaging material was characterized. Finally, compound 1 with enzyme inhibitory activity was successfully obtained from the fermentation product of marine microorganism C. Through HPLC, MS, 1 H NMR and 13 C NMR analyses, its structure was deduced as azelaic acid, and its activity was analyzed. The results showed that this is a feasible method for screening TYR inhibitors in complex systems.
研究领域:
生物化学
·
药物化学
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DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2024
·
IF 10.7
Guoning Chen,Shuxian Zhang,Xiaofei Wang,Xiaoxuan Fan,Gidion Wilson,Yuping Sa,Xueqin Ma
The discovery of enzyme inhibitors from natural products is a crucial aspect in the development of therapeutic drugs. However, the complexity of natural products presents a challenge in developing simple and efficient methods for inhibitor screening. Herein, we have developed an integrated analytical model for screening xanthine oxidase (XOD) inhibitors that combines simplicity, accuracy, and efficiency. This model utilizes a colorimetric sensor and affinity chromatography technology with immobilized XOD . The colorimetric sensor procedure can quickly identify whether there are active components in complex samples. Subsequently, the active components in the samples identified by the colorimetric sensor procedure were further captured, separated, and identified through affinity chromatography . The integrated analytical model can significantly enhance the efficiency and accuracy of inhibitor screening. The proposed method was applied to screen for an activity inhibitor of XOD in five natural medicines. As a result, a potential active ingredient for XOD , polydatin , was successfully identified from Polygoni Cuspidati Rhizoma et Radix. This work is anticipated to offer new insights for the screening of enzyme inhibitors from natural medicines.
研究领域:
药物化学
·
生物分析化学
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高影响因子
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
·
2025
·
IF 3.8
Ma Fen,Wang Weibiao,Wang Mei,Zhang Weiman,Zhang Shuxian,Wilson Gidion,Sa Yuping,Zhang Yue,Chen Guoning,Ma Xueqin
Given the inherent complexity of natural medicines, finding a straightforward and efficient method for identifying active ingredients remains a significant challenge, yet it is of paramount importance. Influenza virus neuraminidase (NA), a primary target for anti-influenza drug development, plays a crucial role in the infection process, making it essential to develop rapid and facile methods for screening NA inhibitors. Herein, we developed a novel and efficient analytical technique for the identification of NA inhibitors from complex herbal medicines by integrating dual sensing with affinity chromatography. This approach simplifies the experimental process and highlights the benefits of being quicker, more sensitive, and cost-effective. Regarding the biosensing section, the innovative concept of a 4-methylumbelliferyl-N-acetylneuraminic acid-NA-based fluorescence paper sensor strategy enables the rapid detection of NA inhibitors in complex herbal samples. In affinity chromatography, bioactive compounds were precisely captured, separated, and identified. The efficacy and reliability of the developed method were confirmed using both negative and positive controls. Then, the method was applied to screen for NA inhibitors in 20 different herbal medicines. The results revealed that Bupleurum chinense DC. exhibited the most pronounced inhibitory effect on NA. Subsequent analysis utilizing affinity chromatography identified three bioactive compounds, namely saikosaponin a, saikosaponin d, and baicalin, as the active agents responsible for this inhibitory effect, with IC 50 values of 177.3 μM, 262.9 μM, and 241.4 μM, respectively. Molecular docking studies further indicated that these three bioactive compounds exhibit a strong binding affinity with NA. This research provides novel insights into the screening of enzyme inhibitors within herbal medicines. Graphical abstract
研究领域:
分析化学
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药理学
·
生物医学工程
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PubMed
高影响因子
JOURNAL OF CHROMATOGRAPHY A
·
2014
·
IF 4.258
Shao-Ting Wang,Wei Huang,Yi-Fan Deng,Qiang Gao,Bi-Feng Yuan,Yu-Qi Feng
The metal oxide affinity chromatography (MOAC) materials have been extensively used for extraction of phosphate compounds in the past decade. Actually, some of these materials also possess adsorption affinity towards cis -diol-containing compounds, which was seldom explored in separation field so far. Here we present the proof-of-concept study to evaluate the feasibility of expanding MOAC for specific capture of cis -diol biomolecules. Benefitting from the high commercialisation of the metal oxide materials, such MOAC strategy possesses several advantages, like synthesis-free, low cost and high expandability. Firstly, the recognition of adenosine against 2′-deoxyadenosine was performed using zirconium oxide and cerium oxide, two typical commercial MOAC materials. The results showed that efficient adsorption and elution could be achieved easily by pH switching from basic to acidic. The isotherm curves demonstrated the adsorption process fitted well with Freundlich isotherm model and was spontaneous at room temperature (Δ G 0 < 0) with an exothermic nature (Δ H 0 < 0). Afterwards, the highly efficient and selective enrichment of various model cis -diol biomolecules, including ribonucleosides, glycopeptides and glycoproteins, was achieved using this MOAC strategy. Finally, the endogenous ribonucleosides and modified ribonucleosides were successfully purified from human urine sample, which demonstrated the potential application of MOAC materials in the enrichment of target compounds from complex biological samples. Besides the excellent performance of extraction for cis -diol-containing compounds, equally important is that these materials are commercially available with low cost, which makes the MOAC a promising strategy for the study of cis -diol biomolecules in metabolomics and proteomics.
研究领域:
分析化学
·
生物化学
·
代谢组学
·
蛋白质组学
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DOI
高影响因子
FOOD CHEMISTRY
·
2018
·
IF 4.946
Hui Cao,Xiaojuan Liu,Nataša Poklar Ulrih,Pradeep K. Sengupta,Jianbo Xiao
Herein, the protein binding rates of structurally different flavonoids to human serum albumin (HSA) were elucidated by applying the high performance affinity chromatography (HPAC). The flavonoids with hydroxyl groups on ring A showed a higher protein binding rate compared with those that there was no hydroxyl on ring A. However, the hydroxylation of ring B lowered the protein binding rate. It was also found that an additional methoxy group in flavone ring A would decrease the protein binding rate. Nevertheless, the methoxy group in flavanone ring A (position 6) and isoflavone ring B (position 4′) increased the protein binding rate. Methoxy group at other positions of flavonoids slightly enhanced or no significantly affected the binding rates on human serum albumin. Hydrogenation of C2 C3 double bond of flavonoids decreased the protein binding rate and had the same effect as glycosylation which decrease the protein binding rate by 5%–25%.
研究领域:
药理学
·
生物化学
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PubMed
高影响因子
MACROMOLECULAR MATERIALS AND ENGINEERING
·
2018
·Vol. 303
·期 12
·1800411
·
IF 2.69
Huimin Gu,Yibin Liu,Dezhong Yin,LuKui Cai,Baoliang Zhang,Qiuyu Zhang
In this study, glycidyl methacrylate-based polymerized high internal phase emulsion monolithic column (polyHIPE) with submicron skeletons and well-defined macropores is prepared and functionalized by immobilizing heparin to obtain a novel affinity chromatographic separation medium for effective purification of Enterovirus 71 (EV71). The pore structure of the obtained monoliths differs significantly from conventional polyHIPE monoliths. The prepared polyHIPE presents well-defined 3D network skeletons of 0.2–0.5 µm, interconnected macropores of 0.5–2 µm, large specific surface area (47–61 m 2 g −1 ), and excellent permeability ( k = 1.0 × 10 −13 m 2 ). Further, successful immobilization of heparin is proved by FTIR and XPS and the monoliths display superiority in the purification of EV71 from culture supernatant. The final recovery and binding capacity of EV71 are calculated to be 60% and 152.8 ng per column, respectively. SDS-PAGE gel electrophoresis and TEM analysis indicate a high efficiency in the removal of impurities from culture supernatant of EV71. This, together with easy fabrication, makes polyHIPE monolith a promising alternative to commercially available monolithic supports for virus purification.
研究领域:
化学工程
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生物技术
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病毒学
·
材料科学
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DOI
高影响因子
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
·
2024
·
IF 2.8
Hongbei Liu,Qiumin Xu,Michael Adu-Frimpong,Yuchu Chen,Ran Li,Fei Xu,Xia Cao,Shanshan Tong
In order to enrich the selection of biological ligands, realize the miniaturization analysis, and broaden the application of monolith materials for active ingredients screening and separating, we sough to construct a lipid raft @capillary monolith microcolumn affinity chromatography model. Single factor experiments and various characterization methods, including scanning electron microscopy (SEM) and thermogravimetric analysis, were employed to investigate the polymerization of the monolith column under different material ratios to determine optimal preparation conditions. Subsequently, the lipid raft from U251 cells was integrated with the monolith materials based on epoxy-based covalent crosslinking principle and characterized through SEM and immunofluorescence methods. Afterwards, the retention of positive drug gefitinib, negative drug gemcitabine and four licorice standards solution on the prepared lipid raft monolith microcolumn was then detected via electrochemical detection. The results exhibited that there was no specific adsorption for any active compounds on the blank monolith materials. Significantly, the lipid raft monolith microcolumn packed with TrkA-target proteins could be successfully validated for positive drug gefitinib with a high affinity sorption efficiency of 51.2%. This work expands the range of the utilization of affinity chromatography carriers and the selection of biological ligands, providing a new idea for the screening of active ingredients.
研究领域:
分析化学
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药理学
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生物技术
·
分子生物学
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PubMed
高影响因子
JOURNAL OF PHARMACEUTICAL AND BIOMEDICAL ANALYSIS
·
2025
·
IF 3.1
Yaqi Wang,Ping Shu,Mei Lv,Yonghao Xing,Haoran Zhang,Xiaojing Yan,Bo Wang,Tingting Huang,Minfeng Fang
Farfarae Flos and its honey-processed product are used to treat respiratory diseases; yet their active components and therapeutic targets remained unclear. Herein, we developed an integrated strategy combining receptor chromatography and mice models to screen and evaluate active components in both raw and honey-processed Farfarae Flos. We focused on three key receptors,β 2 -AR, CysLT 1 R, and M 3 R, known to be involved in antitussive, anti-inflammatory, and expectorant activities. These receptors were immobilized onto silica gels to construct affinity columns. Using these columns, we identified the bioactive compounds in both forms of Farfarae Flos. The results revealed an identical set of target-specific compounds across raw and honey-processed samples: chlorogenic acid (CGA) and caffeic acid (CA) targeted β 2 -AR; CGA and rutin bound to CysLT 1 R; and CGA along with isochlorogenic acids A, B, and C interacted with M 3 R. Notably, the interactions of rutin with CysLT 1 R and isochlorogenic acids A, B, and C with M 3 R are reported for the first time. These findings indicate that both raw and honey-fired Farfarae Flos share the same material basis for their antitussive, anti-inflammatory, and expectorant effects, with no significant alteration in pharmacological efficacy due to honey processing. Furthermore, the binding affinities of the bioactive compounds for the receptors correlated well with the molecular docking-predicted binding energies of the corresponding compound-receptor complexes. To our knowledge, this study represents the first target-centric comparative analysis of raw and processed Farfarae Flos. It establishes a generalizable framework for investigating processing-induced changes in herbal pharmacology and provides empirical evidence connecting metabolomic profiles to clinical efficacy.
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高影响因子
JOURNAL OF SEPARATION SCIENCE
·
2025
·Vol. 48
·期 7
·e70219
·
IF 2.8
Xiao-Bei Huang,Yong-Ming Wei,Lin Yu,Zhen-Liang Xu,Hu Yang
Immobilized metal affinity chromatography is a green method applied to the separation and purification of proteins and biomolecules in downstream processing. With its unique internal pores, the membrane carriers have more active sites than typical magnetic nanoparticles and beads. It also greatly overcomes the mass-transfer resistance brought by traditional column chromatography. In this study, polysulfone was selected as the pristine membrane. Tannic acid and 3-aminopropyltriethoxysilane coatings were employed to construct the secondary reaction platform, which was then covalently coupled with polyethyleneimine and grafted with iminodiacetic acid groups to chelate Cu 2+ . Bovine serum albumin was utilized as the adsorption protein to adjust and optimize the modification parameters. The results demonstrated that the adsorption amount per unit membrane area reached 0.19 mg/cm 2 under the solution condition of pH 8.5. The adsorption thermodynamics followed the Langmuir model, and the corresponding model parameters were obtained.
研究领域:
生物技术
·
生物化学
·
材料科学
·
分析化学
·
分离科学
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JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
·
2020
·
IF 3.004
Pengru Liu,Xiongdiao Lan,Muhammad Yaseen,Kungang Chai,Liqin Zhou,Jianhua Sun,Ping Lan,Zhangfa Tong,Dankui Liao,Lixia Sun
Purification of small bioactive peptides from complex biological samples is a difficult task due to the interference of concentrated large biomolecules. In this study, a magnetic immobilized metal affinity chromatography matrix modified by poly (ethylene glycol) methyl ether (IMACM@mPEG) was prepared and applied for the rapid purification of angiotensin I-converting enzyme (ACE) inhibitory peptides from casein hydrolysate. The proposed IMACM@mPEG considerably reduced the non-specific adsorption of large proteins and exhibited improved purification efficiency towards ACE inhibitory peptides. A novel peptide with moderate ACE inhibitory activity (IC 50 value of 274 ± 5 μM) was identified as LLYQEPVLGPVR. Lineweaver-Burk plot confirmed the non-competitive inhibition pattern of LLYQEPVLGPVR. The purified peptide was digested after simulated gastrointestinal digestion and produced shorter peptides which contributed to enhanced ACE inhibitory activity. These results indicated that the IMACM@mPEG is an effective method for the prepurification of ACE inhibitory peptide and the purified peptide LLYQEPVLGPVR may have potential as nutraceutical ingredient in functional foods for hypertension treatments.
研究领域:
生物化学
·
药剂学
·
食品科学
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DOI
PubMed
高影响因子
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