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In this study, a capillary electrophoresis‐based online immobilized enzyme microreactor was developed for evaluating the inhibitory activity of green tea catechins and tea polyphenol extracts on trypsin. The immobilized trypsin activity and other kinetic parameters were evaluated by measuring the peak area of the hydrolyzate of chromogenic substrate S‐2765. The results indicated that the activity of the immobilized trypsin remained approximately 90.0% of the initial immobilized enzyme activity after 30 runs. The value of Michaelis–Menten constant (Km) was (0.47 ± 0.08) mM, and the half‐maximal inhibitory concentration (IC50) and inhibition constant (Ki) of benzamidine were measured as 3.34 and 3.00 mM, respectively. Then, the inhibitory activity of four main catechins (epicatechin, epigallocatechin, epicatechin gallate, and epigallocatechin gallate) and three tea polyphenol extracts (green tea, white tea, and black tea) on trypsin were investigated. The results showed that four catechins and three tea polyphenol extracts had potential trypsin inhibitory activity. In addition, molecular docking results illustrated that epigallocatechin gallate, epicatechin gallate, epicatechin, and epigallocatechin were all located not only in the catalytic cavity, but also in the substrate‐binding pocket of trypsin. These results indicated that the developed method is an effective tool for evaluating inhibitory activity of catechins on trypsin. [ABSTRACT FROM AUTHOR]

Capillary electrochromatography, which combined the high selectivity of high‐performance liquid chromatography and the high separation efficiency of capillary electrophoresis, is an attractive separation tool. In this review, the developments on monolithic and open tubular capillary electrochromatography during 2017 to August 2019 are summarized. Considering the development of novel stationary phases is the most active research field in capillary electrochromatography, monolithic capillary electrochromatography is classified according to the polymer‐based and hybrid monolithic columns, while open‐tubular capillary electrochromatography is categorized by cyclodextrin, silica, polymer, nanomaterials, microporous materials, and biomaterials‐based open tubular columns. [ABSTRACT FROM AUTHOR]

Background: The dry root and rhizome of Salvia miltiorrhiza Bunge, or Danshen, is a well-known traditional Chinese medicine with anticoagulant activity. Taking into account that thrombin (THR) and factor Xa (FXa) play crucial roles in the coagulation cascade, it is reasonable and meaningful to screening THR and/or FXa inhibitors from Danshen. Methods: Four extracts [butanol (BA), ethyl acetate (EA) and remained extract (RE) from 75% ethanol extract, and water extract (WE)] of Danshen were prepared, and their THR/FXa inhibitory activities were assessed in vitro. Then, the active EA extract was further separated by silica-gel column chromatography (SC), and its fractions (SC1–SC5) were analyzed by LC–MS. The principal component analysis (PCA) and orthogonal partial least squares discriminate analysis (OPLS-DA) were employed for predicting the specific marker compounds. The chemical structures of targeted compounds were identified by LC–MS/MS and their interactions with THR/FXa were analyzed by the molecular docking analysis. Results: Danshen EA extract showed strong activity against THR and FXa, and its fractions (SC1–SC5) exhibited obvious difference in inhibitory activity against these two enzymes. Furthermore, four marker compounds with potential THR/FXa inhibitory activity were screened by PCA and OPLS-DA, and were identified as cryptotanshinone, tanshinone I, dihydrotanshinone I and tanshinone IIA. The molecular docking study showed that all these four tanshinones can interact with some key amino acid residues of the THR/FXa active cavities, such as HIS57 and SER195, which were considered to be promising candidates targeting THR and/or FXa with low binding energy (< − 7 kcal mol−1). Conclusions: LC–MS combined with multivariate statistical analysis can effectively screen potential THR/FXa inhibitory components in Danshen. [ABSTRACT FROM AUTHOR]

Background: Salvia miltiorrhiza (Danshen, DS) and Panax notoginseng (Sanqi, SQ) are famous traditional Chinese herbs, and their herbal pair (DS–SQ) has been popular used as anti-thrombotic medicines. However, there is still a lack of sufficient scientific evidence to illustrate the optimum combination ratio of these two herbs as well as its action mechanisms. The purpose of this study is to investigate the anti-thrombotic effects of DS–SQ on zebrafish and explore its possible action mechanism. Methods: Firstly, the chemical components in DS–SQ extract were analyzed by LC–ESI–MS/MS. Then, a phenylhydrazine (PHZ)-induced zebrafish thrombosis model was developed for evaluating the anti-thrombotic effects of DS–SQ extracts with different combination ratios and their nine pure compounds. Followed, Real-time quantitative PCR (RT-qPCR) assays were performed to investigate the potential antithrombotic mechanisms of DS–SQ. Results: Thirty-three components were tentatively identified by LC–MS analysis. DS–SQ at the ratio of 10:1 presented the best anti-thrombotic effect, and rosmarinic acid, lithospermic acid and salvianolic acid B of DS showed good anti-thrombotic activity on zebrafish thrombosis model. The RT-qPCR assays indicated that DS–SQ (10:1) could cure the PHZ-induced thrombosis by downregulating the expression of PKCα, PKCβ, fga, fgb, fgg and vWF in zebrafish. Conclusions: DS–SQ with the combination ratio of 10:1 showed optimum anti-thrombotic effect on PHZ-induced zebrafish thrombosis model, which provided a reference for reasonable clinical applications of DS–SQ herbal pair. [ABSTRACT FROM AUTHOR]

• The applications of immobilized biomaterials in herbal analysis are reviewed. • Different immobilization strategies for biomaterials are discussed. • The carrier materials applied in biomaterials immobilization are summarized. Immobilization of biomaterials developed rapidly due to the great promise in improving their stability, activity and even selectivity. In this review, the immobilization strategies of biomaterials, including physical adsorption, encapsulation, covalent attachment, cross-linking and affinity linkage, were briefly introduced. Then, the major emphasis was focused on the reported various types of immobilized biomaterials, including proteins, enzymes, cell membrane and artificial membrane, living cells, carbohydrates and bacteria, used in the herbal analysis for bioactive compound screening, drug-target interaction evaluation and chiral separation. In addition, a series of carrier materials applied in biomaterials immobilization, such as magnetic nanoparticles, metal-organic frameworks, silica capillary column, cellulose filter paper, cell membrane chromatography, immobilized artificial membrane chromatography and hollow fiber, were also discussed. Perspectives on further applications of immobilized biomaterials in herbal analysis were finally presented. [ABSTRACT FROM AUTHOR]

With the development of nanotechnology, various nanomaterials with enzyme-like activity (nanozymes) have been reported. Due to their superior properties, nanozymes have shown important application potential in the fields of bioanalysis, disease detection, and environmental remediation. However, only a few nanomaterials with multi-enzyme mimicry activity have been reported. In this study, a novel multienzyme mimic was synthesized through a simple and rapid preparation protocol by coordinating copper ions with N3, N6 (amino), N7, and N9 on adenine phosphate. The prepared adenine phosphate-Cu complex exhibits significant peroxidase, laccase, and oxidase mimicking activities. The Michaelis-Menten constant (K m) and the maximal velocity (V max) values of the peroxidase, laccase, and oxidase mimicking activities of AP-Cu nanozyme are 0.052 mM, 0.14 mM, and 2.49 mM; and 0.552 μM min−1, 6.70 μM min−1, and 2.24 μM min−1, respectively. Then, based on its laccase mimicking activity, the nanozyme was applied in the degradation of phenolic compounds. The calculated kinetic constant for the degradation of 2,4-dichlorophenol is 0.468 min−1 and the degradation efficiency of 2,4-dichlorophenol (0.1 mM) reaches 96.14% at 7 min. Finally, based on the multienzyme mimicking activity of adenine phosphate-Cu nanozyme, simple colorimetric sensing methods with high sensitivity and good selectivity were developed for the detection of hydrogen peroxide, epinephrine, and glutathione in the ranges of 20.0–200.0 μM (R2 = 0.9951), 5.0–100.0 μM (R2 = 0.9970), and 5.0–200.0 μM (R2 = 0.9924) with the limits of quantitation of 20.0 μM, 5.0 μM, and 5.0 μM, respectively. In short, the synthesis of nanozymes with multi-enzyme mimicry activity through coordination between copper ions and small molecule mimicry enzymes provides new ideas for the design and research of multi-enzyme mimics. • Adenine phosphate-Cu nanozyme is synthesized via simple coordination reaction. • It exhibits significant peroxidase, laccase, and oxidase mimicking activities. • Adenine phosphate-Cu nanozyme can efficiently degrade phenolic compounds. • It has excellent performance in detecting H 2 O 2 , epinephrine, and glutathione. [ABSTRACT FROM AUTHOR]

In this paper, Cu2+ functionalized Fe 3 O 4 @polydopamine core-shell (Fe 3 O 4 @PDA@Cu2+) magnetic microspheres were prepared by the chelation between Cu2+ and catechol of polydopamine surface. The synthetic magnetic adsorbent was characterized by Fourier transform infrared spectroscopy, energy dispersive X-ray spectroscopy, vibrating sample magnetometry, thermogravimetric analysis, scanning electron microscope and transmission electron microscopy. Four purines include guanine, adenine, hypoxanthine and xanthine were selected as model compounds to evaluate the applicability of this adsorbent. Several parameters that effected the extraction efficiency, such as extraction time, adsorbent amount, solution pH, ionic strength, eluent type, concentration of eluent and eluent time, were investigated. Under the optimized conditions, good linearity was obtained with correlation coefficients between 0.9983 and 0.9999 for the four analytes, and their LOD and LOQ were 0.42–2.15 ng/mL and 1.41–6.50 ng/mL, respectively. Meanwhile, the RSDs of intra-day and inter-day precision were in the range of 1.43%–5.55% and 4.56%–7.01%, respectively. The spiked recoveries of four purines in real sample were 70.01%–102.42%, indicating this proposed method might have potential applications for the analysis of purines in real samples. In addition, the developed method was used to monitor the concentrations of adenine in rat plasma at different time points after intragastric administration. • A Fe 3 O 4 @PDA@Cu2+ magnetic microsphere was prepared and characterized. • Fe 3 O 4 @PDA@Cu2+ was successfully applied in the determination of purines in plasma. • Potential for the extraction of purines from complex matrices with easy separation [ABSTRACT FROM AUTHOR]

Highlights • Metal ions functionalized Fe 3 O 4 @PDA particles for the extraction of nucleotides. • To extract nucleotides from complex matrix by Fe 3 O 4 @PDA@Ti4+ particles. • Potential applications in phosphorylated small molecular compounds extraction. Abstract In this study, a titanium ion (Ti4+) functionalized polydopamine coated ferroferric oxide (Fe 3 O 4 @PDA@Ti4+) core-shell magnetic particle was prepared for the selective extraction of nucleotides. Firstly, different metal ions including Ti4+, Zr4+, Fe3+, Al3+, Cu2+, Zn2+, Ni2+ and Mg2+ were respectively immobilized onto Fe 3 O 4 @PDA particles and their extraction efficiency for five nucleotides [cytidine-5′-monophosphate (CMP), uridine-5′-monophosphate (UMP), guanosine-5′-monophosphate (GMP), thymidine-5′-monophosphate (TMP) and adenosine-5′-monophosphate (AMP)] were compared. Among these prepared materials, Fe 3 O 4 @PDA@Ti4+, which exhibited the highest extraction efficiency for nucleotides, was further characterized by Fourier transform infrared spectroscopy, scanning electron microscopy, transmission electron microscopy and energy dispersive X-ray spectroscopy. After being optimized of the extraction parameters including adsorbent amounts, extraction time, extraction temperature, type and concentration of the eluent, the prepared Fe 3 O 4 @PDA@Ti4+ magnetic particles were successfully applied for the selective extraction and determination of CMP, UMP, GMP, TMP and AMP in Cordyceps and Lentinus edodes. Good linearity (varying from 0.063 to 19.000 μg/mL, R2 > 0.999) and low limit of detection (LODs) (ranging between 0.0047 and 0.0141 μg/mL) for target analytes were achieved. These results demonstrated that the synthesized material in this study had potential for selective extraction of phosphorylated small molecular compounds in complicated matrix. [ABSTRACT FROM AUTHOR]

Abstract In this study, a novel amino-terminated supramolecular cucurbit [6] uril (CB [6]) pseudorotaxane motif coated magnetic nanoparticles (MNPs) was successfully fabricated through cucurbit [6] uril promoted azide-alkyne cycloaddition. The physicochemical properties of the obtained functional MNPs were investigated by scanning electron microscope, transmission electron microscope, energy dispersive X-ray spectroscopy, Fourier transform infrared spectroscopy, X-ray diffractometer, vibrating sample magnetometer, thermal gravimetric-derivative thermogravimetric analysis, and Zeta potential analyzer. Due to the abundant carboxyl groups in the structure of CB [6] and amino residue on the surface of the pseudorotaxane motif MNPs, the prepared materials were employed to extract salvianolic acids (SAs) for the first time. A series of significant parameters in the extraction and elution process, including sample pH, acetonitrile percentage, ionic strength and extraction temperature, as well as eluent type and desorption time, were examined systematically. Based on the results, the electrostatic interaction, van der Waals interaction, hydrophilic interaction and hydrogen bonding formation between the analytes and sorbent were speculated. The equilibrium state of static adsorption study indicated the outstanding adsorption capacity of pseudorotaxane motif MNPs toward six SAs to be 15–80 mg/g, providing much better adsorption affinity for SAs compared with MNPs only modified with amino, and their adsorption behaviors were all fitted with Freundlich adsorption isotherm model. In addition, the developed pseudorotaxane motif MNPs offered good recyclability and stability. Taking advantage of these merits, four SAs from Salvia miltiorrhiza Bunge were successfully extracted using the prepared pseudorotaxane motif MNPs with relative recovery in the range from 95.1% to 106.5% and RSDs below 9.7%. These results demonstrated that the synthesized pseudorotaxane motif MNPs showed potential as a candidate in the extraction of SAs. Graphical abstract fx1 Highlights • Novel MNPs material was fabricated by CB [6] promoted azide-alkyne click chemistry. • First attempt to extract SAs by the synthetic CB [6] pseudorotaxane motif MNPs. • High adsorption capacity, good recyclability and stability were achieved for SAs. [ABSTRACT FROM AUTHOR]

Abstract A capillary electrophoresis (CE)-based beta-glucosidase (beta-Glu) immobilized enzyme microreactor (IMER) was constructed for enzyme kinetics study and inhibitor screening with the aid of polydopamine coating. The enzyme kinetic and inhibition studies of beta-Glu were comprehensively evaluated using p -nitrophenyl beta- d -glucopyranoside as a model substrate and castanospermine as a model inhibitor. The Michaelis-Menten constant value of the immobilized beta-Glu in the developed IMER was calculated to be 2.79 mmol/L. The half-maximal inhibitory concentration and inhibition constant of castanospermine were 13.22 μg/mL and 1.54 μg/mL, respectively. In addition, after 50 consecutive runs, the IMER activity was remained at 89.5% of the initial immobilized beta-Glu activity, which showed that the constructed IMER has good stability and repeatability. Finally, the developed method was successfully applied to screen beta-Glu inhibitors from twelve flavonoids. Four flavonoids include genistein, baicalein, epicatechin gallate and epigallocatechin gallate had significant inhibitory effect on beta-Glu, and their binding mode with enzyme was further verified via the molecular docking analysis. In summary, the developed CE based beta-Glu-IMER is a reliable method for screening beta-Glu inhibitors from natural products. Highlights • A stable and reliable online CE-based beta-Glu-IMER was firstly constructed. • The prepared IMER was successfully applied to screen beta-Glu inhibitors. • Four flavonoids were discovered as new inhibitors targeting beta-Glu. • The enzyme-inhibitor binding mode was further verified by molecular docking. [ABSTRACT FROM AUTHOR]

Background: Salvia miltiorrhiza (Danshen [DS]) and Carthamus tinctorius (Honghua [HH]) are commonly used traditional Chinese medicines for activating blood and removing stasis, and they were usually used as DS-HH (Danshen-Honghua [DH]) herbal pair in clinical applications. Characterizing the antioxidant active ingredients in DH herbal pair may be helpful for understanding their curative effect on cardiovascular diseases. Objective: The objective of the study is to screen the 1, 1-diphenyl-2-trinitrophenylhydrazine (DPPH) scavenging active compounds in DH herbal pair by spectrum-effect relationship analysis. Materials and Methods: First, the water extracts of DH herbal pair with different ratios (DS: HH = 1:0, 0:1, 1:1, 2:1, 3:1, 5:1, 1:5, 1:3, and 1:2) were prepared. Then, the clearance effects of DH herbal pair and single drugs on DPPH were compared, meanwhile, high performance liquid chromatography was applied for chemical analysis of DH extracts. Finally, DPPH scavengers in DH herbal pair were predicted and identified by spectrum-effect relationship analysis and liquid chromatography-mass spectrometry (LC-MS) analysis. Results: Compared with single drugs, the extracts of herbal pairs had higher clearance rate to DPPH. Eighteen potential active compounds (peaks) in the extract of DH herbal pair were predicted, and 13 of them were tentatively identified by LC-MS analysis. Furthermore, antioxidant activities of eight pure compounds from the DH herbal pair were validated by DPPH radical scavenging assay with Vitamin C as positive control drug. Among them, six compounds including danshensu, protocatechuic acid, coffee acid, chlorogenic acid, rutin, and salvianolic acid A were found to have high antioxidant activity. Conclusion: DH herbal pair showed strong clearance effect on DPPH, and danshensu, protocatechuic acid, coffee acid, chlorogenic acid, rutin and salvianolic acid A are the active components. Abbreviations Used: DS: Danshen, HH: Honghua, DH: Danshen-Honghua, TCMs: Traditional Chinese medicines, LC-MS: Liquid chromatography-mass spectrometry, DPPH: 1, 1-diphenyl-2-trinitrophenylhydrazine, VC: Vitamin C, CCA: Canonical correlation analysis. [ABSTRACT FROM AUTHOR]

Context: Selaginella tamariscina (P. Beauv.) Spring (Selaginellaceae) (ST) has been widely used in China as a medicine for improving blood circulation. However, its processed product, S. tamariscina carbonisatus (STC), possesses opposite haemostatic activity. Objective: To comprehensively evaluate the activity of ST and STC on physiological coagulation system of rats, and seek potential active substances accounting for the activity transformation of ST during processing. Materials and methods: The 75% methanol extracts of the whole grass (fine powder) of ST and STC were prepared, respectively. Male Sprague-Dawley rats were randomly divided into five groups: control group, model group, model + ST group, model + STC group and positive control group (model + Yunnanbaiyao). The duration of intragastric administration was 72 h at 12 h intervals. Haemorheology parameters were measured using an LB-2 A cone-plate viscometer and the existed classic methods, respectively. SC40 semi-automatic coagulation analyzer was employed to determine coagulation indices. Meanwhile, HPLC and LC-MS were applied for chemical analyses of ST and STC extracts. Results: STC shortened tail-bleeding time, increased whole blood viscosity (WBV) and plasma viscosity (PV), decreased erythrocyte sedimentation rate blood (ESR), reduced activated partial thromboplastin time (APTT) and increased the fibrinogen (FIB) content in the plasma of bleeding model rats. Although ST could shorten APTT and TT, the FIB content was significantly decreased by ST. Dihydrocaffeic acid with increased content in STC vs. ST showed haemostatic activity for promoting the platelet aggregation induced by collagen and trap-6, and reducing APTT and PT significantly with a concentration of 171.7 μM in vitro. Amentoflavone with reduced content in STC vs. ST inhibited ADP and AA-induced platelet aggregation significantly with a concentration of 40.7 μM. Discussion and conclusions: As the processed product of ST, STC showed strong haemostatic activity on bleeding rat through regulating the parameters involved in haemorheology and plasma coagulation system. Two active compounds, dihydrocaffeic acid and amentoflavone, might be partially responsible for the haemostatic and anticoagulant activity of STC and ST, respectively. [ABSTRACT FROM AUTHOR]

Highlights • A mathematical model to characterize confounding electrostatic effects on IAM surface for diverse analytes. • The intrinsic phospholipid-water sorption coefficient (K IAM,intr) was explored. • Sigmoidal correlations between Caco-2 permeability and K IAM,intr were investigated. Abstract The passive membrane permeability of drugs may be potentially predicted by phospholipid-water sorption coefficient (K PLIPW) through immobilized artificial membrane (IAM) chromatography. However, for predominantly ionized compounds, unexpected confounding electrostatic interaction would influence the accurate determination of K PLIPW , accompanied by the mobile phase with varied pH and ionic strength. In order to measure the intrinsic phospholipid-water sorption coefficient (K IAM,intr) for diverse analytes, a mathematic model by characterizing the confounding electrostatic effects was developed in this study. Based on the developed electrostatic model, additional electrostatic repulsion/attraction interactions with the charged IAM surface were characterized for organic cations/anions. Considering that most of the IAM chromatography data was determined under physiological conditions, the K PLIPW values of all compounds were calculated with the ionic strength of 0.1 M and the pH value of 7. Generally, K PLIPW was hardly affected by the pH and ionic strength of mobile phase for those electroneutral compounds. K IAM,intr was 0.28 log units smaller than apparent phospholipid-water sorption coefficient (K IAM,app) for cations. In contrast, K IAM,intr was larger than K IAM,app by 0.28 log units for anions. Herein, K IAM,intr of diverse analytes can be easily calculated by the developed electrostatic model. For application, K IAM,intr was further compared with intrinsic Caco-2 permeability, and good sigmoidal correlations (R2 = 0.76) were established between them for organic cations and neutral compounds but not for organic anions. [ABSTRACT FROM AUTHOR]

[Display omitted] • The silver nanocomposite is in situ synthesized on paper substrate. • Detection of Fe3+ is based on the aggregation of silver nanocomposite. • The sensitivity for Fe3+ detection can be tuned by the pre-concentration steps. • On site detection of Fe3+ can be realized with the aid of a smartphone. In this study, a green and novel method for in situ synthesis of silver nanocomposites on paper substrate was developed and used for the pre-concentration and determination of Fe3+. Silver nanocomposites were in situ synthesized using tannic acid as reducing and capping agent under a mildly alkaline condition. The mechanism for the Fe3+ detection is based on its coordination with the tannic acid present on the surface of the silver nanocomposites and the formation of aggregates. The colorimetric response between Fe3+ and tannic acid modified silver nanocomposites deposited on the surface of the paper substrate was collected by a smartphone followed by processing with Adobe Photoshop CS 5.1 software. Under the optimal conditions, the color response showed a good linear relationship with Fe3+ concentration in the range of 25.0–500.0 μM. The limit of detection for Fe3+ was calculated to be 10.0 μM. Finally, the developed smartphone assisted paper-based chemical sensor was used to detect Fe3+ in two lake water samples, and the recoveries are in the range of 83.5 %−106.7 %. The sensitivity for Fe3+ detection can be tuned through the pre-concentration steps based on the porous nature of the filter paper. The platform highlights the advantage of simplicity, ease of synthesis, fast response time, and low reagent/analyte volume requirement, which is suitable for point-of-use application in iron ions contamination, especially in the remote or resource-limited regions. [ABSTRACT FROM AUTHOR]

Abstract: For lead compound discovery from natural products, hollow fiber cell fishing with chromatographic analysis is a newly developed method. In this study, an adsorbed hollow fiber‐based biological fingerprinting method was firstly developed to discover potential platelet aggregation inhibitors from Danshen–Honghua decoction. Platelets were seeded on the fiber and their survival rate was tested. Results indicated that more than 92% platelets survived during the whole operation process. Ranitidine and tirofiban were used as positive and negative control respectively to verify the reliability of the presented approach. The main variables such as amount of extract and stirring time that affect the adsorbed hollow fiber‐based biological fingerprinting process were optimized, and the repeatability of this method was also investigated. Finally, 12 potential active compounds in Danshen–Honghua decoction were successfully detected using the established approach and structures for nine of them were tentatively identified by liquid chromatography with mass spectrometry analysis. In addition, the in vitro platelet aggregation inhibition test was carried out for five of the nine hit compounds, and three active components, namely, lithospermic acid, salvianolic acid A, and salvianolic acid B were confirmed. These results proved that the proposed method could be an effective approach for screening platelet inhibitors from plant extracts. [ABSTRACT FROM AUTHOR]

Tyrosinase (TYR) is a rate-limiting enzyme in the synthesis of melanin, while direct TYR inhibitors are a class of important clinical antimelanoma drugs. This study established a spectrum-effect relationship analysis method and high-performance liquid chromatography-mass spectrometry (LC-MS) analysis method to screen and identify the active ingredients that inhibited TYR in Salvia miltiorrhiza–Carthamus tinctorius (Danshen–Honghua, DH) herbal pair. Seventeen potential active compounds (peaks) in the extract of DH herbal pair were predicted, and thirteen of them were tentatively identified by LC-MS analysis. Furthermore, TYR inhibitory activities of five pure compounds obtained from the DH herbal pair were validated in the test in which kojic acid served as a positive control drug. Among them, three compounds including protocatechuic aldehyde, hydroxysafflor yellow A, and tanshinone IIA were verified to have high TYR inhibitory activity (IC50 value of 455, 498, and 1214 μM, resp.) and bind to the same amino acid residues in TYR catalytic pocket according to the results of the molecular docking test. However, the other two compounds lithospermic acid and salvianolic acid A had a weak effect on TYR, as they do not combine with the active amino acid residues or act on the active center of TYR. Therefore, the developed methods (spectrum-effect relationship analysis and molecular docking) could be used to effectively screen TYR inhibitors in complex mixtures such as natural products. [ABSTRACT FROM AUTHOR]

Thrombin plays a vital role in blood coagulation, which is a key process involved in thrombosis by promoting platelet aggregation and converting fibrinogen to form the fibrin clot. In the receptor concept, drugs produce their therapeutic effects via interactions with the targets. Therefore, investigation of interaction between thrombin and small molecules is important to find out the potential thrombin inhibitor. In this study, affinity capillary electrophoresis (ACE) and in silico molecular docking methods were developed to study the interaction between thrombin and ten phenolic compounds (p-hydroxybenzoic acid, protocatechuic acid, vanillic acid, gallic acid, catechin, epicatechin, dihydroquercetin, naringenin, apigenin, and baicalein). The ACE results showed that gallic acids and six flavonoid compounds had relative strong interactions with thrombin. In addition, the docking results indicated that all of optimal conformations of the six flavonoid compounds were positioned into the thrombin activity centre and had interaction with the HIS57 or SER195 which was the key residue to bind thrombin inhibitors such as argatroban. Herein, these six flavonoid compounds might have the potential of thrombin inhibition activity. In addition, the developed method in this study can be further applied to study the interactions of other molecules with thrombin. [ABSTRACT FROM AUTHOR]

In present study, magnetic molecularly imprinted polymers (MMIPs) were successfully prepared for specific recognition and selective enrichment of phloridzin from the leaves of Malus doumeri (Bois) A. Chev and rats' plasma. The magnetic Fe 3 O 4 were prepared by the solvothermal reaction method and followed by the modification of TEOS and functionalization with APTES. Using functionalized Fe 3 O 4 particles as the magnetic cores, phloridzin as template, ethylene glycol dimethacrylate (EGDMA) as cross-linker and 2,2-azobisisobutyonnitrile (AIBN) as initiator, the MMIPs were prepared through APTES to associate the template on the surface of the magnetic substrate. The structural features and morphological characterizations of MMIPs were performed by FT-IR, SEM, TEM, XRD, TGA and VSM. The adsorption experiments revealed that the MMIPs presented high selective recognition property to phloridzin. The selectivity experiment indicated that the adsorption capacity and selectivity of polymers to phloridzin was higher than that of baicalin and 2,3,5,4′-ttrahydroxy stilbene-2-O-β-D-glucoside. Furthermore, the MMIPs were employed as adsorbents for extraction and enrichment of phloridzin from the leaves of M. doumeri and rats' plasma. The recoveries of phloridzin in the leaves of M. doumeri ranged from 81.45% to 90.27%. The maximum concentration (C max ) of phloridzin in rats' plasma was detected as 12.19 ± 0.84 μg/mL at about 15 min after oral administration of phloridzin (200 mg/kg). These results demonstrate that the prepared MMIPs are suitable for the selective adsorption of phloridzin from complex samples such as natural medical plants and biological samples. [ABSTRACT FROM AUTHOR]

Context:Corydalis yanhusuoW.T. Wang (Papaveraceae) (Rhizoma Corydalis) showed inhibitory effects on rabbit platelet aggregation induced by ADP, thrombin (THR) or arachidonic acid (AA). Objective:This study separates and identifies the possible target-related platelet proteins and suggests possible signal cascades of RC antiplatelet aggregation. Materials and methods:Based on comparative proteomics, the differentially expressed platelet proteins treated before and after with 50 mg/mL RC 90% ethanol extract (for 15 min at 37 °C) were analyzed and identified by two dimensional gel electrophoresis (2-DE) and MALDI-TOF-MS/MS. To further verify the possible signalling pathways of RC antiplatelet aggregation function, the concentration of calcium (Ca2+) was measured by Fura-2/AM fluorescence (Ex 340/380 nm, Em 500 nm) (RC final concentrations of 0.0156–0.1563 mg/mL), the levels of P-selectin and cyclic guanosine monophosphate (cGMP) were quantified by ELISA (OD. 450 nm) (RC final concentrations of 0.0156–1.5625 mg/mL), and the 5-hydroxytryptamine (5-HT) level was measured usingortho-phthalaldehyde (OPT) fluorescence (Ex 340 nm, Em 470 nm) (RC final concentrations of 0.3125–1.5625 mg/mL). Results:The expression of 52 proteins were altered in rabbit platelets after the treatment and the MALDI-TOF-MS analysis indicated that those proteins include 12 cytoskeleton proteins, 7 cell signalling proteins, 3 molecular chaperone proteins, 6 proteins related to platelet function, 16 enzymes and 7 other related proteins. Furthermore, RC extract could decrease the levels of 5-HT [inhibition rate of 96.80% (p < 0.05,vs. THR-activated group) treated with 0.7813 mg/mL of RC], Ca2+ [172.73 ± 5.07 to 113.56 ± 5.46 nM (p < 0.001,vs. THR-activated group) treated with 0.0313 mg/mL of RC] and P-selectin [13.48 ± 0.96 ng/3 × 108to 11.64 ± 0.17 ng/3 × 108(p < 0.05,vs. THR-activated group) treated with 0.0156 mg/mL of RC], and increase in cGMP level [38.93 ± 0.57 to 50.26 ± 4.05 ng/3 × 108(p <0.05,vs. THR-activated group) treated with 1.5165 mg/mL of RC] in ADP (10 μmol/L), THR (0.25 u/mL) or AA-(0.205 mmol/L) activated rabbit platelets. Discussion and conclusion:The present study indicated that P2Y12 receptor might be one of the direct target proteins of RC in platelets. The signal cascades network of RC after binding with P2Y12 receptor is mediating Gαi proteins to activate downstream signalling pathways (AC and/or PI3K signalling pathways) for the inhibition of platelet aggregation. [ABSTRACT FROM AUTHOR]

Thrombin (THR) plays a significant role in thromboembolic diseases, direct THR inhibitors are a class of important clinical anticoagulant drugs. This study established a THR in-solution based biospecific extraction combined with ultrafiltration and high performance liquid chromatography coupled with diode array detector and mass spectrometry analysis (TUA) method to screen and identify ligands for THR in Rhizoma Chuanxiong. After evaluating the reliability of the present TUA method using positive (argatroban) and negative (adenosine, tirofiban, ticagrelor) control drugs, this method was successfully applied to detect eight potential active compounds in Rhizoma Chuanxiong. Two new THR-targeted compounds isochlorogenic acid C and senkyunolide I with high THR inhibitory activity (IC50 206.48 and 197.23 μM, respectively) were identified by liquid chromatography/mass spectrometry and enzyme inhibitory activity test finally. They were reported with direct THR inhibition activity for the first time and their ligand-THR interactions were explored by in silico molecular docking research. In addition, based on the TUA screening result, four compounds gained similar structure with the two hit compounds were also investigated as promising candidates targeting THR with high binding energy (>5.0 kcal/mol). These results may prove that the proposed method could effectively screen THR inhibitors in complex mixtures. [ABSTRACT FROM AUTHOR]

A bisubstrate colorimetric system was developed for rapid and multi-color assay of copper ion (Cu2+). Cu2+-triethylamine (Cu2+-TEA) complex with the peroxidase-like activity can catalyze the 3,3′,5,5′-tetramethylbenzidine (TMB) to produce oxidized TMB (TMB+, color in blue) and dopamine (DA) to oxidized DA (oxDA, color in orange) in the presence of hydrogen peroxide, which can obtain a three-color colorimetric result due to the couple of complementary colors of blue and orange. The determination of Cu2+ is achieved based on the absorbance value at 460 nm of the composite peak of TMB+ and oxDA. After systematically optimizing the reaction conditions, the developed bisubstrate colorimetric method exhibits high sensitivity for the detection of Cu2+ (limit of quantitation (LOQ) is 0.0625 μM) with a wide linear range of 0.0625–8.0 μM (R2 = 0.991). Meanwhile, the reaction solution shows an orange, brown, and green color change with the increase in Cu2+ concentration, which can realize the semi-quantitatively detection by the naked eyes. Furthermore, TMB was also used as a single substrate for Cu2+ detection with a linear range of 0.5–9.0 μM (R2 = 0.994), and the LOQ is determined to be 0.5 μM. The LOQ of bisubstrate system is 8 times lower than that of single substrate system, indicating a higher sensitivity for the detection of Cu2+ using the bisubstrate colorimetric system. In addition, the developed bisubstrate colorimetric method exhibits excellent selectivity for Cu2+ detection, which was applied in the determination of Cu2+ in lake water and soil samples, and satisfactory spiked recoveries were obtained. [Display omitted] • Cu2+-triethylamine (TEA) complex exhibits excellent peroxidase-like activity. • A bisubstrate (TMB + dopamine) colorimetric sensor is developed based on Cu2+-TEA complex. • The developed senor displays three-color change for the detection of Cu2+. • The bisubstrate sensor shows a higher sensitivity than that of single substrate. [ABSTRACT FROM AUTHOR]

Nano-sized molecularly imprinted polymers for tiliroside were successfully prepared by a precipitation polymerization method. Acrylamide, ethylene glycol dimethacrylate, azobisisobutyronitrile, and acetonitrile/dimethyl sulfoxide were used as functional monomer, cross-linker, initiator, and porogen, respectively. The structural features and morphological characterization of tiliroside-imprinted polymers were characterized by Fourier transform infrared spectroscopy and scanning electron microscopy, respectively. The adsorption experiments indicated that the tiliroside-imprinted polymers exhibited high selective recognition property to tiliroside. Scatchard analysis indicated that the homogeneous-binding sites were formed in the polymers. The selectivity test revealed that the adsorption capacity and selectivity of polymers to tiliroside was significantly higher than that of rutin, astragalin, and kaempferol. Finally, the tiliroside-imprinted polymers were employed as adsorbents in solid-phase extraction for the extraction of tiliroside from the ethyl acetate extract of the flowers of Edgeworthia gardneri (wall.) Meisn. The results demonstrated that the extraction recoveries of tiliroside ranged from 69.3 to 73.5% by using tiliroside-imprinted polymers coupled with solid-phase extraction method. These results indicated that the tiliroside-based molecularly imprinted solid-phase extraction method was proven to be an effective technique for the separation and enrichment of tiliroside from natural medicines. [ABSTRACT FROM AUTHOR]

Context:The rising problem of atherosclerosis and ischemic heart disease emphasizes the need to look for new antithrombotic components with effective modes of action.Corydalis yanhusuo(Y.H. Chou & Chun C. Hsu) W.T. Wang ex Z.Y. Su & C.Y. Wu (Papaveraceae) (Rhizoma Corydalis) has been used in the traditional medicines for the treatment of cardiovascular disease. Objective:The antiplatelet aggregation compounds in Rhizoma Corydalis were screened to validate its traditional medicinal use. Material and methods:Total alkaloid extract (TAE) of Rhizoma Corydalis was obtained by refluxing 100 g Rhizoma Corydalis powder with 600 mL 70% ethanol, and purified by acidification (20% HCl) and alkalization (5 M NaOH) process. Potential antiplatelet aggregation compounds in TAE were screened by a method involving platelet bio-specific extraction and HPLC-DAD/LC–MS analysis. Furtherin vitroantiplatelet aggregation activity confirmation of TAE and seven main alkaloids were achieved by turbidimetry method within 3 h after blood collection from rabbit carotid artery, and all the test drugs were at the concentration range of 25–350 μg/mL. Finally, HPLC-DAD was employed for the quantitative determination of seven main components in TAE. Results:Five alkaloids, identified as glaucine, dehydrocorydaline, canadine, tetrahydrocoptisine and corydaline, can be specifically extracted with platelets. The results indicated that all these five alkaloids can inhibit thrombin-induced platelet aggregation in a low dose (IC50of glaucine, dehydrocorydaline, canadine, tetrahydrocoptisine and corydaline were 49.057, 34.914, 33.547, 84.261 and 54.164 μg/mL, respectively) as compared to TAE (IC50 = 175.426 μg/mL) and aspirin (IC50 = 300.340 μg/mL), while the unbound compounds (palmatine and tetrahydropalmatine) had a very weak antiplatelet effect (IC50 > 200 μg/mL). Discussion and conclusion:This study is the first reported work for antiplatelet components screening in Rhizoma Corydalis. Seven compounds were detected and identified by HPLC-DAD/LC–MS, of which five platelet-targeted compounds were discovered. [ABSTRACT FROM AUTHOR]

In this study, the cobalt carbonate (CoCO 3) was in situ prepared using cobalt (Ⅱ) and carbonate at room temperature, and the intrinsic peroxidase-like activity of CoCO 3 was demonstrated for the first time through the oxidation of chromogenic substrate 3, 3', 5, 5' - tetramethylbenzidine (TMB) in the presence of hydrogen peroxide (H 2 O 2). The effects of pH, reaction temperature, and incubation time on the catalytic activity of CoCO 3 were studied. Under the optimal reaction conditions, the Michaelis constant (K m) of CoCO 3 to H 2 O 2 is 0.443 mM, which is lower than that of natural horseradish peroxidase and the previously reported peroxidase mimics. Furthermore, the mechanism exploration experiments confirm that the catalytic activity of CoCO 3 is mainly attributed to the hydroxyl radical (•OH). Finally, based on the outstanding peroxidase-like activity of CoCO 3 , a colorimetric sensor for the detection of H 2 O 2 , glucose, and ascorbic acid (AA) was developed, which shows satisfactory selectivity and recovery. In addition, the total antioxidant capacity of three types of tea, dried lemon, three kinds of fresh fruit juice, and vitamin C tablet were determined. In short, this study reports a novel in situ generated CoCO 3 , which can be prepared rapidly at room temperature, eliminating the harsh, cumbersome, and time-consuming synthesis and purification process. The developed sensor based on the newly found excellent peroxidase-like activity of in situ prepared CoCO 3 for the determination of H 2 O 2 , glucose, and AA is fast, simple, low cost, and high sensitivity. [Display omitted] • The excellent peroxidase-like activity of CoCO 3 is firstly investigated. • The CoCO 3 is in situ prepared of rapidity and simplicity. • The hydroxyl radical (•OH) is the major active specie in the catalytic reaction. • The nanozyme has preferable kinetics with a low K m value towards substrate H 2 O 2. • A colorimetric detection platform for H 2 O 2 , glucose, and ascorbic acid is developed. [ABSTRACT FROM AUTHOR]

[Display omitted] • Products of urease catalyzes urea induce color change of Prussian blue nanoparticles. • A paper-based sensor depending on the Prussian blue pH sensitivity is developed. • Smartphone captures color signals of the sensor to achieve rapid detection of urea. Depending on the pH sensitivity of Prussian blue nanoparticles (PB NPs) and the specificity of urease-catalyzed urea, a paper-based colorimetric sensor was developed using urease, PB NPs, and cellulose filter paper. With the assistance of a smartphone, the rapid detection of urea can be realized. Under the optimal conditions, the linear range of the paper-based sensor for detection of urea is 0.5–5.0 mM, and the limit of detection is 0.27 mM. The RSD values of within-batch and between-batch repeatability are 2.5 % and 0.9 %, respectively. The method was successfully applied to the detection of urea in spiked milk samples and artificial urine sample, and the recoveries are in the range of 91.8–107.9%. The developed paper-based colorimetric sensor is a simple, economic, and rapid method that can be used for the determination of urea in real samples, particularly in resource-poor regions. [ABSTRACT FROM AUTHOR]

The formation of Pb-O bond between Pb SP 2+ sp and L-Cysteine can induce significant aggregation of Au NPs (blue color), conversely, Cu can catalyze the oxidation of Cysteine and prevent the aggregation of Au NPs (pink color). DOI: 10.1002/sscp.202200019 The cover picture shows that the lead ion (Pb SP 2+ sp ) and copper ion (Cu SP 2+ sp ) in lake water are detected by UV-Vis analysis using gold nanoparticles (Au NPs) as the colorimetric probe. [Extracted from the article]

A simple and time-saving colorimetric method was developed to quantify sulfonamides (SAAs) in milk via inhibition of the human carbonic anhydrase II (hCAII)-like activity of ZIF-8 that can hydrolyze p -nitrophenyl acetate (p NPA) to p -nitrophenol (p NP), following the color change from yellow to colorless. Effects of different reaction conditions, including pH, temperature, amount of ZIF-8, and incubation time, were investigated. The value of Michaelis-Menten constant (K m) is measured to be 0.15 mM, which exhibits high affinity to p NPA. The IC 50 (0.17, 0.24, and 0.60 mM) and inhibition constant (K i) (0.09, 0.13, and 0.33 mM) of sulfamethazine (SD), sulfadimethoxine (SDM), and sulfathiazole (ST) on ZIF-8 were measured, respectively. Moreover, the activity of ZIF-8 remains more than 90.0% of its initial activity after 30 days' storage. The colorimetric method for SD, SDM, and ST determination was established at the linear ranges of 6.3–750.0 μM (1.75–208.75 mg/kg), 6.3–750.0 μM (1.96–232.75 mg/kg), and 5.0–1250.0 μM (1.28–319.15 mg/kg) with limit of detection of 4.3, 3.2, and 3.9 μΜ (1.2, 0.99, and 0.96 mg/kg), respectively. In addition, the spiked recoveries of SAAs in milk sample are in the range of 81.6%–106.7% with RSD less than 6.5%. In short, the developed colorimetric method can achieve rapid analysis of SAAs in milk with simple operations. [Display omitted] • A simple colorimetric method is developed for sulfonamides detection. • The method is based on the human carbonic anhydrase II-like activity of ZIF-8. • The method can achieve rapid analysis of sulfonamides with simple operation. • It provides a possible strategy for the detection of sulfonamides in milk. [ABSTRACT FROM AUTHOR]

The Co 3 (PO 4) 2.8 H 2 O nanozyme was in situ synthesized through direct reaction between cobalt (Ⅱ) and phosphate, and its peroxidase-like activity was studied in the presence of hydrogen peroxide (H 2 O 2) using a chromogenic substrate 3, 3′, 5, 5′ - tetramethylbenzidine (TMB). The effects of pH value, temperature, and incubation time on the conversion of TMB to oxidized TMB (oxTMB) catalyzed by Co 3 (PO 4) 2.8 H 2 O in the presence of H 2 O 2 were optimized. The peroxidase-like activity of Co 3 (PO 4) 2.8 H 2 O conforms to the Michaelis-Menten kinetics, and its Michaelis constant (K m) is 0.073 mM, showing a higher H 2 O 2 affinity than natural horseradish peroxidase (HRP) and some other peroxidase nanozymes. The mechanism exploration experiment confirmed that the hydroxyl radical (•OH) is the main active species in the catalytic reaction of Co 3 (PO 4) 2.8 H 2 O. Finally, based on the inherent peroxidase-like activity of Co 3 (PO 4) 2.8 H 2 O, a simple colorimetric detection method for H 2 O 2 was established, with a linear range of 15.0 − 100.0 μM and a detection limit of 4.385 μM. The method was applied to the determination of H 2 O 2 in milk samples with the spiked recovery of 90.2%− 106.2%. This study reports a novel nanozyme that can be easily produced in situ, which is exempt from the harsh, cumbersome, and time-consuming synthesis and purification procedures. In addition, the H 2 O 2 colorimetric detection method based on the nanozyme has the advantages of low cost, convenient, high sensitivity, and good selectivity. [Display omitted] • A novel nanozyme Co 3 (PO 4) 2.8 H 2 O of excellent catalytic activity is discovered. • This peroxidase-mimic nanozyme is in situ synthesized rapidly. • The hydroxyl radical is the major active specie in the catalytic reaction. • A sensitive colorimetric method for hydrogen peroxide determination is established. [ABSTRACT FROM AUTHOR]

Salvia miltiorrhiza (Danshen, DS), the dried root and rhizome of Salvia miltiorrhiza Bunge and Ligusticum chuanxiong (Chuanxiong, CX), the dried rhizomes of Ligusticum striatum DC are effective in invigorating blood circulation and eliminating stasis which is highly related with cardiovascular disease (CVD). The identification of activity-based chemical markers is very important, but the complex mechanism of "multi-component, multi-target, and multi-effect" within traditional Chinese medicine (TCM) poses a great challenge to this work. In this study, we combined network pharmacological prediction with experimental validation of the DS and CX to explore an effective method for discovering quality control (QC) of antithrombotic herbs by clarifying the intermediate layer "module/cluster" between the whole complex system and a single component. Based on structural similarity analysis of compound and the thrombosis network published before, we firstly modularized two layers called chemical cluster (CC) network and functional module (FM) network respectively and linked them into one bilayer modularized compound target (BMCT) network. "Two-step" calculation was applied on identifying the significant compounds as the potential QC markers from CC. The in vitro inhibitory activity of selected QC marker compounds on thrombin was evaluated to partially verify their pharmacological activities. HPLC was used to determine contents. According to the network-based analysis, nine compounds with great importance in the BMCT network were identified as QC markers of DS-CX, including tanshinone I, tanshinone IIA, cryptotanshinone, salvianolic acid B, ferulic acid, salvianolic acid A, rosmarinic acid, chlorogenic acid, and coniferyl ferulate. Enzyme inhibitory test partially verified the activity of tanshinone I and tanshinone IIA. Chemical profiling indicated that the nine marker compounds are the main components in the herbal pair. This study identified activity-based QC markers of DS-CX herbal pair and provided a new methodology that can be used in the QC of other herbs, herbal pairs, or formulas. [Display omitted] [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance In Tibet, the flower of Edgeworthia gardneri (Wall.) Meisn., locally named “Lvluohua, 绿萝花”, has been traditionally used to treat diabetes mellitus for many years. Aim of this study To evaluate the activity of dual agonists for PPARγ/β from the flower of E.gardneri in vitro . Materials and methods HeLa cells were transiently co-transfected with the re-constructed plasmids of pBIND-PPARγ-LBD or pBIND-PPARβ-LBD and rL4.35. The activities of crude extracts, secondary fractions and compounds from the flower of E.gardneri were evaluated with the transfected cells. Rosiglitazone (at 0.5 μg/mL) and L-165041 (at 0.5 μg/mL) were used as the positive controls for PPARγ and PPARβ respectively. Results The results demonstrated that n-hexane, ethyl acetate and n-butanol extracts from the flower of E.gardneri were able to significantly activate PPARγ and PPARβ respectively, and the activity of ethyl acetate extract was much better. We further observed that, among the 11 secondary fractions of ethyl acetate extract, the fr. 9 could activate PPARγ and PPARβ significantly. Moreover, umbelliferone (from fr.9) and pentadecanoic acid could activate PPARγ and PPARβ at the same time. Conclusions The extracts from the flower of E.gardneri could significantly activate PPARγ and PPARβ. Besides, umbelliferone and pentadecanoic acid isolated from the flower of E.gardneri were the new agonists for PPARγ and PPARβ. [ABSTRACT FROM AUTHOR]

[Display omitted] • A sensitive visual colorimetric sensor is developed for the detection of trichlorfon. • The sensor is based on inhibition of enzyme-induced metallization of gold nanorods. • The method is able to achieve rapid analysis of trichlorfon with simple operation. • It provides a possible strategy for the detection of pesticides i.e. trichlorfon. In this study, a highly sensitive visual colorimetric sensor was designed for the detection of trichlorfon based on inhibiting ascorbate oxidase (AAO)-induced metallization of gold nanorods (Au NRs). Ascorbic acid (AA) can reduce silver ion (Ag+) to metal silver (Ag) that will be deposited on the surface of Au NRs, which results in the blue shift of longitudinal localized surface plasmon resonance (LSPR) peak of Au NRs, accompanying by perceptible color changes from red to cyan to red to yellow. In the presence of trichlorfon, the activity of AAO will be inhibited, resulting in less AA is hydrolyzed to dehydroascorbic acid (DHA), and therefore more Ag+ is reduced to Ag by AA. Under the optimized conditions, detection of trichlorfon has a wide linear range of 27.8–11111.1 μg/L with a limit of detection as low as 132.6 ng/L. Moreover, the sensor has a good sample spiked recovery (84.7–96.8%) for the determination of trichlorfon in lake water samples. The proposed method can achieve rapid analysis (about 10 min) of trichlorfon with simple operation when there are no other types of organophosphorus pesticides in the real samples. [ABSTRACT FROM AUTHOR]

A convenient and green method was developed using a personal glucose meter (PGM) for direct determination of hydrogen peroxide and hypochlorite in household disinfectants based on the acetylthiocholine iodide (ATCI)-mediated reaction. In this method, acetylcholinesterase catalyzes the hydrolysis of ATCI to generate thiocholine iodide, which can trigger the reduction of K 3 [Fe(CN) 6 ] to K 4 [Fe(CN) 6 ] and generate a PGM detectable signal. When the hydrogen peroxide or hypochlorite is pre-incubated with the ATCI, the yield of thiocholine will be decreased because the iodine molecules can initiate the oxidation of thiocholine to disulphides, leading to a lower PGM readout. Thus, the hydrogen peroxide or hypochlorite can be directly quantified by PGM as simple as detecting glucose in blood. After being systematically investigated, the optimum conditions of the PGM method are as follows: 5.0 min of enzymatic reaction time, 6.0 mM final concentration of ATCI solution, and 10.0 min of reaction time between ATCI/H 2 O 2 and thiocholine. Moreover, the recoveries of hydrogen peroxide in real sample spiked with three different concentrations (final concentrations of 5.0, 10.0, and 20.0 mM) are in the range of 96.3–108.4%. The recoveries of hypochlorite in real sample spiked with three different concentrations (final concentrations of 1.0, 5.0, and 10.0 mM) are in the range of 99.4–117.0%. These results indicate that the developed method can be employed for the detection of hydrogen peroxide and hypochlorite in household disinfectants. [Display omitted] • A simple and green method for direct detection of hydrogen peroxide and hypochlorite. • Generating personal glucose meters detectable signals based on acetylthiocholine iodide-mediated reaction. • The user-friendly detection process is as simple as detection of glucose in blood. [ABSTRACT FROM AUTHOR]

• The covalently bonded liposome capillary column is prepared by a new three-step method. • The immobilized liposome capillary column (ILCC) has good repeatability and stability. • Retention factors of seventeen drugs are determined using ILCC and compared with other systems. • Partitions of drugs in ILCC are explained by linear free energy relationship analysis with seven solute descriptors. • This study provide reference information for evaluating the membrane permeability of drugs [Display omitted] Two liposomes, including 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) + 1,2-dimyristoyl- sn -glycero-3-phosphoethanolamine (PE) + cholesterol (Chol) (DSPC/PE/Chol liposomes) and soybean lecithin (SPC) + PE + Chol (SPC/PE/Chol liposomes), were prepared and fixed on the inner wall of capillary column by using the adhesion of polydopamine (PDA) membrane and the cross-linking property of glutaraldehyde (GA). The immobilized liposome capillary column (ILCC) has good repeatability and stability based on the electrophoretic mobility of analyte. A CE method based on the immobilized liposome capillary column chromatography (ILCCC) was successfully developed to study the retention behavior of drugs on ILCC, and the logarithm of retention factor (log k) of neutral and ionic drugs were determined. The results show that the log k measured by the ILCCC based on two liposomes have a good linear fitting (R2 = 0.86). Moreover, the linear relationship between ILCCC system and other related research systems (octanol-water system and immobilized artificial membrane (IAM)) was analyzed, and the results indicate that SPC/PE/Chol ILCCC, DSPC/PE/Chol ILCCC and IAM systems have good fitting results, R2 values are 0.86 and 0.78, respectively. In addition, the normalization coefficients of ILCCC and IAM systems obtained by the linear free energy relationship (LFER) analysis are close and the d value is small. In short, the ILCCC is a simple and feasible method for studying drug membrane permeability. [ABSTRACT FROM AUTHOR]

[Display omitted] • Adenine phosphate (AP) is first found exhibiting intrinsic peroxidase-like activity. • The hydroxyl radical (•OH) is the active specie of catalytic reaction of AP. • A sensitive colorimetric method for hydrogen peroxide and ascorbic acid detection is established. • The findings facilitate the development of new small-molecule enzyme mimics. The peroxidase-like activity of adenine phosphate (AP) was investigated for the first time using chromogenic substrate 3, 3′, 5, 5′-tetramethylbenzidine (TMB) in the presence of H 2 O 2. Effects of different reaction conditions, including pH, temperature, AP concentration and incubation time on the catalytic property of AP, were investigated. The peroxidase-like activity of AP complies with Michaelis-Menten kinetics, and the calculated kinetic parameter (K m , 0.038 mM) of AP catalysis exhibits higher affinity to H 2 O 2 than natural horseradish peroxidase and some of the other small-molecule peroxidase mimics. The radical scavenging results confirm that hydroxyl radical (•OH) is the active specie of catalytic process. Based on the excellent catalytic performance of AP, a facile colorimetric method for the H 2 O 2 and ascorbic acid detection with good sensitivity and selectivity was established at the linear ranges of 2.0–50.0 μM and 20.0–70.0 μM, respectively. In addition, the developed method was applied to detect H 2 O 2 and ascorbic acid in practical samples. In short, this work will not only broaden the understanding of biological function of AP, but also facilitate the development of new category of small-molecule enzyme mimics. [ABSTRACT FROM AUTHOR]

In this study; a spectrum–effect relationship analysis combined with a high-performance liquid chromatography–mass spectrometry (LC–MS) analysis was established to screen and identify active components that can inhibit thrombin and factor Xa (THR and FXa) in Salviae Miltiorrhizae Radix et Rhizoma–Chuanxiong Rhizoma (Danshen–Chuanxiong) herbal pair. Ten potential active compounds were predicted through a canonical correlation analysis (CCA), and eight of them were tentatively identified through an LC–MS analysis. Furthermore; the enzyme inhibitory activity of six available compounds; chlorogenic acid; Z-ligustilide; caffeic acid; ferulic acid; tanshinone I and tanshinone IIA; were tested to verify the feasibility of the method. Among them; chlorogenic acid was validated to possess a good THR inhibitory activity with IC50 of 185.08 µM. Tanshinone I and tanshinone IIA are potential FXa inhibitors with IC50 of 112.59 µM and 138.19 µM; respectively. Meanwhile; molecular docking results show that tanshinone I and tanshinone IIA; which both have binding energies of less than −7.0 kcal·mol−1; can interact with FXa by forming H-bonds with residues of SER214; GLY219 and GLN192. In short; the THR and FXa inhibitors in the Danshen–Chuanxiong herbal pair have been successfully characterized through a spectrum–effect relationship analysis and an LC–MS analysis. [ABSTRACT FROM AUTHOR]

To date, peroxisome proliferator-activated receptors (PPARs) are becoming the new therapeutic targets for the treatment of metabolic diseases, such as Type 2 diabetes, obesity, and cardiovascular disease. In this study, a cell-based high-throughput PPARs (PPARα/β/γ) model was developed for the screening of PPARs agonists. The screening conditions were evaluated through analyzing the expression value of luciferase. Finally, 24 h of drug acting time, 5 times of the dilution factor of luciferase zymolyte, and about 2 × 104cells/ well on HeLa cells in 96-well plates were used, respectively. Furthermore, the quality of high-throughput screening (HTS) in stability and reliability was evaluated by the Z′-factor. Additionally, different extracts of Rhizoma Coptis and berberine were tested by the developed method. The results suggested that both the EtOAc extract and berberine were able to activate PPARα/β/γ, and Rhizoma Coptis contains potential natural agonists of PPARs besides berberine. In conclusion, the developed HTS assay is a simple, rapid, stable, and specific method for the screening of PPARs natural agonists. [ABSTRACT FROM PUBLISHER]

As a key enzyme regulating postprandial blood glucose, α-Glucosidase is considered to be an effective target for the treatment of diabetes mellitus. In this study, a simple, rapid, and effective method for enzyme inhibitors screening assay was established based on α-glucosidase catalyzes reactions in a personal glucose meter (PGM). α-glucosidase catalyzes the hydrolysis of maltose to produce glucose, which triggers the reduction of ferricyanide (K3[Fe(CN)6]) to ferrocyanide (K4[Fe(CN)6]) and generates the PGM detectable signals. When the α-glucosidase inhibitor (such as acarbose) is added, the yield of glucose and the readout of PGM decreased accordingly. This method can achieve the direct determination of α-glucosidase activity by the PGM as simple as the blood glucose tests. Under the optimal experimental conditions, the developed method was applied to evaluate the inhibitory activity of thirty-four small-molecule compounds and eighteen medicinal plants extracts on α-glucosidase. The results exhibit that lithospermic acid (52.5 ± 3.0%) and protocatechualdehyde (36.8 ± 2.8%) have higher inhibitory activity than that of positive control acarbose (31.5 ± 2.5%) at the same final concentration of 5.0 mM. Besides, the lemon extract has a good inhibitory effect on α-glucosidase with a percentage of inhibition of 43.3 ± 3.5%. Finally, the binding sites and modes of four active small-molecule compounds to α-glucosidase were investigated by molecular docking analysis. These results indicate that the PGM method is feasible to screening inhibitors from natural products with simple and rapid operations. [ABSTRACT FROM AUTHOR]

[Display omitted] • Enzyme-mediated sodium alginate gelation on paper-based sensor for glucose detection. • The sodium alginate viscosity is converted to distance-readout on paper-based sensor. • Combination of paper-based sensor and distance-readout method for glucose detection. In this study, a paper-based sensor, combined with a visual distance-readout method, was developed to determine glucose in fruit samples based on the glucose oxidase-mediated sodium alginate gelation. The type of filter paper, the concentration of sodium alginate and the enzymatic reaction conditions were systematically investigated. Under optimal conditions, the increase in diffusion diameter showed a good linear relationship with glucose concentration between 1.4–7.0 mM, and the limit of quantification was 1.4 mM. Finally, the applicability of the proposed strategy was successfully verified by measuring glucose concentrations in fruit samples. The results generated by the developed paper-based sensor were in good agreement with the results obtained from a glucose assay kit. The recoveries were 91.8%–99.1%. In short, the present study developed a simple, low-cost and efficient method for assessing fruit quality and for guiding fruit intake for diabetic patients, especially in remote or resource-limited regions. [ABSTRACT FROM AUTHOR]

In this study, a polydopamine (PDA)-modified hollow fiber-immobilized xanthine oxidase (XOD) was prepared for screening potential XOD inhibitors from flavonoids. Several parameters for the preparation of PDA-modified hollow fiber-immobilized XOD, including the dopamine concentration, modification time, XOD concentration and immobilization time, were optimized. The results show that the optimal conditions for immobilized XOD activity were a dopamine concentration of 2.0 mg/mL in 10.0 mM Tris-HCl buffer (pH 8.5), a modification time of 3.0 h, an XOD concentration of 1000 μg/mL in 10.0 mM phosphate buffer (pH 7.5) and an immobilization time of 3.0 h. Subsequently, the enzymatic reaction conditions such as the pH value and temperature were investigated, and the enzyme kinetics and inhibition parameters were determined. The results indicate that the optimal pH value (7.5) and temperature (37 °C) of the PDA-modified hollow fiber-immobilized XOD were consistent with the free enzyme. Moreover, the PDA-modified hollow fiber-immobilized XOD could still maintain above 50% of its initial immobilized enzyme activity after seven consecutive cycles. The Michaelis–Menten constant (Km) and the half-maximal inhibitory concentration (IC50) of allopurinol on the immobilized XOD were determined as 0.25 mM and 23.2 μM, respectively. Furthermore, the PDA-modified hollow fiber-immobilized XOD was successfully applied to evaluate the inhibitory activity of eight flavonoids. Quercetin, apigenin, puerarin and epigallocatechin showed a good inhibition effect, and their percentages of inhibition were (79.86 ± 3.50)%, (80.98 ± 0.64)%, (61.15 ± 6.26)% and (54.92 ± 0.41)%, respectively. Finally, molecular docking analysis further verified that these four active compounds could bind to the amino acid residues in the XOD active site. In summary, the PDA-modified hollow fiber-immobilized XOD is an efficient method for the primary screening of XOD inhibitors from natural products. [ABSTRACT FROM AUTHOR]

• A Fe 3 O 4 @COF@NH 2 nanoparticle is fabricated via Schiff-base and thiol-ene click reactions. • The material has high selectivity and adsorption capacity for flavonoid glycosides. • The material has good recyclability and storage stability. In this study, a surface functionalized Fe 3 O 4 composite with core–shell structure was fabricated based on Schiff-base and thiol-ene click reactions. The Fe 3 O 4 @COF@NH 2 magnetic nanoparticles were characterized by scanning electron microscopy, transmission electron microscopy, energy dispersive X-ray spectrometry, Fourier transform infrared spectroscopy, X-ray diffraction, Brunauer-Emmett-Teller and vibrating sample magnetometer. The material showed high specific surface area, high pore volume and satisfactory magnetism. Furthermore, the parameters for the extraction of four flavonoid glycosides, including the amount of adsorbent, adsorption time, pH value, adsorption temperature, ion strength, elution solvent and elution time, were investigated. While the adsorption capacity and adsorption mechanism were explored through static adsorption experiment, high adsorption capacity for rutin (31.6 mg/g), vitexin (36.7 mg/g), hyperoside (47.0 mg/g) and icariin (54.1 mg/g) were obtained based on the strongly π-π stacking, hydrogen-bonding and hydrophobic interactions. Furthermore, Freundlich model with higher R2 value than Langmuir model, was more appropriate for fitting the isotherm adsorption. Finally, the material was applied for the separation, pre-concentration and analysis of four flavonoid glycosides in the fruit of Crataegus pinnatifida extract and rat plasma. Under the optimal conditions, the recoveries of spiked samples were between 89.2% and 106.5% with RSDs ranged from 0.2% to 6.2%. The results revealed a fast binding kinetics, high adsorption capacity, satisfactory selectivity, very good reusability and storage stability of the Fe 3 O 4 @COF@NH 2. [ABSTRACT FROM AUTHOR]

• A phytic acid modified α-Glu/Cu 3 (PO 4) 2 ·3H 2 O hybrid nanoflower for enzyme immobilization. • The developed method improves the activity and stability of α-Glucosidase. • This reported strategy provides a reference for efficient enzyme immobilization. A simple, convenient and efficient enzyme immobilization method through phytic acid (PA) modified α-Glucosidase (α-Glu)/Cu 3 (PO 4) 2 ·3H 2 O hybrid nanoflower was developed. The structural properties of the materials were studied by several characterization techniques. Subsequently, the enzymatic reaction conditions such as the pH value and temperature were optimized, and the enzyme kinetics and inhibition parameters were determined. The PA modified α-Glu/Cu 3 (PO 4) 2 ·3H 2 O hybrid nanoflower had better enzymatic activity under a wide pH range and high temperature than the free one. After seven successive cycles, the PA modified α-Glu/Cu 3 (PO 4) 2 ·3H 2 O hybrid nanoflower could still maintain approximately 63.0 % of its initial immobilized enzyme activity. The Michaelis-Menten constant (K m) and the half-maximal inhibitory concentration (IC 50) of acarbose were determined as 0.77 mM and 15.01 μM, respectively. In addition, the material was applied to evaluate the inhibitory activity of ten phenolic compounds on α-Glu, and epicatechin gallate, gallocatechin gallate, epigallocatechin gallate and rosmarinic acid showed good inhibitory activity with % of inhibition of (53.42 ± 2.39)%, (37.28 ± 1.32)%, (37.08 ± 0.63)% and (35.53 ± 0.23)%, respectively. These results indicate that the PA modified hybrid nanoflower is an efficient method of α-Glu immobilization. [ABSTRACT FROM AUTHOR]

• The tyrosinase (TYR) is simply immobilized by an adsorb hollow fiber. • An enzyme inhibitor screening method is developed based on the immobilized TYR. • Three TYR inhibitors are identified from Pueraria lobata. In this study, a method based on adsorbed hollow fiber immobilized tyrosinase (TYR) was developed to screening potential TYR inhibitors from Pueraria lobate extract. Kojic acid and ranitidine were used as positive and negative control to verify the reliability of the proposed method, respectively. Several significant parameters of the screening process, including the amount of P. lobata extract, adsorption time and incubation time, were optimized. After investigating the repeatability of the developed method, seven potential active compounds in P. lobata extract were successfully detected and their chemical structures were tentatively identified by liquid chromatography – mass spectrometry analysis. Furthermore, the inhibitory activity of four identified compounds on TYR was tested in vitro, and three of them, namely, puerarin, puerarin-6″- O -xyloside and puerarin apioside were verified to have good TYR inhibitory activity with IC 50 value of 478.5, 513.8, and 877.3 μM, respectively. In addition, the molecular docking results indicated that these compounds could bind to the amino acid residues in TYR catalytic pocket. These results proved that the proposed method is a feasible approach for screening of TYR inhibitors from plant extract. [ABSTRACT FROM AUTHOR]

In this work, efficient, sensitive bifunctional-monomer chitosan magnetic molecularly imprinted polymers (BCMMIPs) were fabricated and successfully applied to concentrate the metabolites of Epimedium flavonoids in rat testis and bone that were later analyzed using UPLC-Q-TOF-MS. Using chitosan and methacrylic acid as co-functional monomers, BCMMIPs exhibited a large adsorption capacity (7.60 mg/g), fast kinetics (60 min), and good selectivity. Chitosan is bio-compatible and non-toxic, and methacrylic acid provides multiple hydrogen bond donors. The BCMMIPs were injected into rat testis to specifically enrich the total flavonoid metabolites in vivo and were used to extract metabolites from bone in vitro. The results showed that the BCMMIPs coupled with UPLC-Q-TOF-MS successfully identified 28 compounds from testis and 18 compounds from bone, including 19 new compounds. This study provided a reliable protocol for the concentration of metabolites from complex biological samples, and several new metabolites of Epimedium flavonoids were found in vivo and in vitro. Image 1 • Bifunction-monomer chitosan magnetic molecularly imprinted polymers were created. • The BCMMIPs were applied to enrich the epimedium glycosides metabolites. • 19 new metabolites of Epimedium flavonoids were found. • Provides a protocol for the concentration of metabolites from biological samples. [ABSTRACT FROM AUTHOR]

In this report, a non-toxic Dual Template Molecularly Imprinted Polymers (DMIPs) was synthesized with quercetin and schisandrin b as template molecules, using deep-eutectic solvents as functional monomers for the first time. The DMIPs were used to efficiently and simultaneously enrich quercetin and schisandrin b from the mixed crude extracts of penthorum and schisandra. The results indicated that the DMIPs exhibited rapid adsorption kinetics (80 min for adsorption equilibrium) and high selectivity. The largest adsorbing capacities to quercetin and schisandrin b were 23.58 mg/g and 41.64 mg/g, respectively. After presaturation with quercetin and schisandrin b, the nontoxic saturated DMIPs were fed to the mice. Blood samples of the mice were taken and both quercetin and schisandrin b were successfully detected. The pharmacokinetics of quercetin and schisandrin b were similar to reports in the literature where mice were directly fed with botanicals. Our study provides a reliable protocol such that DMIPs can be used to separate and enrich several target molecules simultaneously from complex biological systems. Our findings suggested that the DMIPs have potential application as a drug delivery system of compound herbal formulas. Image 1 • Dual Template Molecularly Imprinted Polymers (DMIPs) was synthesized with DES as functional monomers. • DMIP was used to enrich quercetin and schisandrin b from the mixed crude extracts. • Enrich several target molecules simultaneously from complex biological systems. [ABSTRACT FROM AUTHOR]

Thrombotic events act as a critical factor that interferes with Cardiovascular Diseases (CVDs), and antithrombotic herbal medicine is a long-standing controversial issue. Although a dispute is involved in their clinical application, all parties unanimously agree that herbal products have been widely used in folk medicine, and their interactions with conventional drugs are of high concern. This study aims to investigate how antithrombotic herbal medicines interact with Western cardiovascular drugs on the molecular level by taking an example of the most frequently used herbal pair, Danshen−Chuanxiong (DS−CX), and to discover more scientific evidence on their potential herb−drug interactions. Network pharmacology (NP), as an analytical approach of a complex system, is used to visualize and compare target profiles of DS−CX and Western cardiovascular drugs, which can be applied to predict common herb-drug targets and to construct a solid context for discussing herb−drug interactions. These interactions are further validated by in vitro assays, while in vivo zebrafish model employed for evaluating an overall pharmacological efficacy of herbal pairs in specific combination ratios. The study finds that DS could react directly to the Western cardiovascular drug targets relevant to antithrombotic pathways (i.e. , thrombin, coagulation factor Xa and cyclooxygenase-1), whereas CX could not react directly and can synergistically affect antithrombotic effects with DS in specific combination ratios. Moreover, it is indicated that DS−CX may generate wide biological functions by a complicated mechanism of "neuro−immune−metabolism/endocrine" (NIM), which can further cause multiple direct and indirect interactions with Western cardiovascular drugs. From the clinical perspective, herb−drug interactions should be given high attention, especially when multiple herbs are used simultaneously. [ABSTRACT FROM AUTHOR]

Background: The dry root and rhizome of Ligusticum chuanxiong Hort., or Chuanxiong, has been used as a blood-activating and stasis-removing traditional Chinese medicine for 1000 years. Our previous studies have shown the inhibitory activity on platelet and thrombin (THR) of Chuanxiong. THR and factor Xa (FXa) play significant roles in the coagulation cascade and their inhibitors are of valuable in the treatment of thromboembolic diseases. The aim of the present study is to screen THR and FXa inhibitors from Chuanxiong. Methods: Four extracts [ethyl acetate (EA), butanol (BA) and remained extract (RE) from 75% ethanol extract, and water extract (WE)] of Chuanxiong were prepared, and their THR/FXa inhibitory activities were assessed in vitro. Following silica-gel column chromatography (SC), the active EA extract and BA extract was further partitioned, respectively. Their active fractions (EA-SC1 to EA-SC5; BA-SC1 to BA-SC5) were obtained and analyzed by LC–MS. After modeling by the principal component analysis (PCA) and orthogonal partial least squares discriminate analysis (OPLS-DA), the specific marker compounds were predicted and identified. Their enzyme inhibitory was assessed in vitro and interactions with THR/FXa were investigated by molecular docking analysis. Results: Chuanxiong EA extract showed strong activity against THR and BA extract was more effective in inhibiting FXa activity, and their fractions exhibited obvious difference in enzyme inhibitory activity. Furthermore, marker compounds a–h were predicted by PCA and OPLS-DA, and their chemical structures were identified. Among them, senkyunolide A, Z-ligustilide, ferulic acid and senkyunolide I (IC50 was determined as 0.77 mM) with potential THR inhibitory activity, as well as isochlorogenic acid A with FXa inhibitory activity were screened out. It was found that the four components could interact with the active site of THR, and the binding energy was lower than − 5 kcal/mol. Isochlorogenic acid A were bound to the active site of FXa, and the binding energy was − 9.39 kcal/mol. The IC50 was determined as 0.56 mM. Conclusions: THR/FXa inhibitory components in different extracts of Chuanxiong were successfully characterized by the method of enzyme inhibition activity assays with ultra performance liquid chromatography-quadrupole time of flight mass spectrometry-based multivariate statistical analysis. [ABSTRACT FROM AUTHOR]

• A simple, economical and fast on-line immobilized trypsin microreactor is developed. • The develop method is applied for evaluating inhibitory activity of phenolic acids. • The structure-activity relationship of phenolic acids on trypsin is discussed. Phenolic acids, which are important aromatic secondary metabolites, are widely distributed in plant foods. In this study, a simple, economical and fast on-line immobilized trypsin microreactor was developed for evaluating the inhibitory activity of phenolic acids by capillary electrophoresis. The Michaelis-Menten constant (K m) of immobilized trypsin was determined as 0.99 mM, and the half-maximal inhibitory concentration (IC 50) and inhibition constant (K i) of benzamidine were measured as 3.39 and 1.68 mM, respectively. Then, the developed strategy was applied to investigate the inhibitory activity of six phenolic acids on trypsin. The results showed that gallic acid, caffeic acid and ferulic acid had high inhibitory activity at concentration of 150 μM. Molecular docking results illustrated that gallic acid, caffeic acid and ferulic acid can interact indirectly with the catalytic and substrate-binding sites of trypsin. The developed strategy is an effective tool for evaluating inhibitory activity of phenolic acids on trypsin. [ABSTRACT FROM AUTHOR]

• TYR & β-Glu are simultaneously entrapped in PDA modified magnetic alginate beads. • TYR-mediated polymerization & internal setting strategy are integrated. • Leakage of enzymes is significantly reduced by introducing PDA to alginate beads. • The developed method can be used for dual-enzymes inhibition assays. In this study, a simple and efficient method to obtain entrapment of mixtures of double enzymes is developed. As a proof of principle, double enzymes (tyrosinase (TYR) and β-glucosidase (β-Glu)) were co-immobilized in magnetic alginate-polydopamine (PDA) beads using in situ TYR-mediated dopamine polymerization and internal setting strategy-mediated magnetic alginate-PDA gelation. The leakage of enzymes from the magnetic alginate beads was significantly reduced by exploiting the double network cross-linking of alginate and PDA, which was induced by the d -(+)-Gluconic acid δ-lactone (GDL) and TYR, respectively. The physicochemical properties of the prepared magnetic alginate beads were characterized by Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD) and scanning electron microscopy (SEM) analysis. After that, the enzymatic reaction conditions and the performance of the entrapped TYR and β-Glu, such as enzyme kinetics and inhibition kinetics, were investigated. The Michaelis-Menten constants (K m) of the entrapped TYR and β-Glu were determined as 2.72 and 3.45 mM, respectively. The half-maximal inhibitory concentrations (IC 50) of kojic acid and castanospermine for the entrapped TYR and β-Glu were determined as 13.04 and 56.23 μM, respectively. Finally, the entrapped double enzymes magnetic alginate beads were successfully applied to evaluate the inhibitory potency of six kinds of tea polyphenols extracts. Black tea and white tea showed high inhibition activity against TYR were (36.14 ± 1.43)% and (36.76 ± 2.35)%, respectively, while the black tea and dark tea showed high inhibition activity against β-Glu were (37.89 ± 6.70)% and (21.28 ± 4.68)%, respectively. [ABSTRACT FROM AUTHOR]

A magnetite@graphene oxide nanocomposite was first coated with polyethylenimine and then modified with phytic acid and titanium(IV) ions. The high loading with Ti(IV) and the good hydrophilicity of PEI and PA result in a material that can be applied to the efficient extraction of highly polar nucleobases, nucleosides and nucleotides. The physicochemical properties of the composite were investigated by scanning electron microscopy, transmission electron microscopy, energy dispersive X-ray spectroscopy, Fourier transform infrared spectroscopy, water contact angle measurements, thermogravimetric analysis, and vibrating sample magnetometry. A series of parameters that affect extraction and elution under the conditions of immobilized metal affinity chromatography (IMAC) and hydrophilic interaction liquid chromatography (HILIC) were examined. The analytes were eluted from the nanocomposites using 10 mM trisodium phosphate as the elution solution in the IMAC mode, and 50% methanol-water as elution solution in the HILIC mode. Figures of merit include (a) an intra-day precision of 0.1-1.0% in the IMAC mode; (b) an intra-day precision of 0.4%-0.8% in the HILIC mode; (c) detection limits between 1.8-2.8 ng mL−1 in the IMAC mode; and (d) detection limits of 4.0-10.5 ng mL−1 in the HILIC mode. The method was applied to the extraction of the nucleotides cytidine-5′-monophosphate (CMP), uridine-5′-monophosphate (UMP), guanosine-5′-monophosphate (GMP), and adenosine-5′-monophosphate (AMP), and the nucleobases and nucleosides hypoxanthine, adenosine, cytosine, inosine and cytidine from Cordyceps sinensis, Lentinus edodes and plasma samples.Schematic presentation of the workflow for the extraction of nucleobases, nucleosides and nucleotides using phytic acid-Ti(IV) functionalized magnetite@graphene oxide nanocomposites under two distinct modes. [ABSTRACT FROM AUTHOR]