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Abstract Plants usually produce defence metabolites in non-active forms to minimize the risk of harm to themselves and spatiotemporally activate these defence metabolites upon pathogen attack. This so-called two-component system plays a decisive role in the chemical defence of various plants. Here, we discovered that Panax notoginseng, a valuable medicinal plant, has evolved a two-component chemical defence system composed of a chloroplast-localized β-glucosidase, denominated PnGH1, and its substrates 20(S)-protopanaxadiol ginsenosides. The β-glucosidase and its substrates are spatially separated in cells under physiological conditions, and ginsenoside hydrolysis is therefore activated only upon chloroplast disruption, which is caused by the induced exoenzymes of pathogenic fungi upon exposure to plant leaves. This activation of PnGH1-mediated hydrolysis results in the production of a series of less-polar ginsenosides by selective hydrolysis of an outer glucose at the C-3 site, with a broader spectrum and more potent antifungal activity in vitro and in vivo than the precursor molecules. Furthermore, such β-glucosidase-mediated hydrolysis upon fungal infection was also found in the congeneric species P. quinquefolium and P. ginseng. Our findings reveal a two-component chemical defence system in Panax species and offer insights for developing botanical pesticides for disease management in Panax species.

Chiral molecules have similar physicochemical properties, which are different in terms of physiological activities and toxicities, rendering their differentiation and recognition highly significant. Nanozymes, which are nanomaterials with inherent enzyme-like activities, have garnered significant interest owing to their high cost-effectiveness, enhanced stability, and straightforward synthesis. However, constructing nanozymes with high activity and enantioselectivity remains a significant challenge. This review briefly introduces the synthesis methods of chiral nanozymes and systematically summarizes the latest research progress in enantioselective recognition of chiral molecules based on electrochemical methods and ultraviolet–visible absorption spectroscopy. Moreover, the challenges and development trends in developing enantioselective nanozymes are discussed. It is expected that this review will provide new ideas for the design of multifunctional chiral nanozymes and broaden the application field of nanozymes.

Due to the copper (Cu) active sites, its complexes with oxidase-like activity have superior catalytic properties, which can catalyze a series of specific substrates like 3,3′,5,5′-tetramethylbenzidine (TMB), producing colorimetric reactions for the detection of different reducing small-molecule compounds. Attribute to the competitive coordination effects between water molecules and central Cu ions, most of the Cu complexes can hardly be used in the pure aqueous reaction system. In this study, a Cu-based material (Cu-imidazole-2-carboxaldehyde, Cu-ICA) was prepared using copper ions and ICA through a one-step process in the water solution. After the morphology of the material being characterized, the mimetic enzyme behavior of the Cu-ICA was demonstrated through the TMB oxidation. Compared to the other reported oxidase-like mimics, Cu-ICA has better aqueous stability and oxidase-like activity, and shows a higher vmax. Furthermore, basing on the oxidase-like activity of Cu-ICA, a colorimetric method was developed for the ascorbic acid and glutathione detections with linear ranges of 0.5–5 μM and 0.5–4 μM, and limit of detection of 0.1304 μM and 0.097 μM, respectively. Owing to its excellent aqueous stability and oxidase-like activity, Cu-ICA has bright application prospects in the analysis of reducing small-molecule compounds.

The high toxicity of arsenic (As) can cause irreversible harm to the environment and human health. In this study, the chlorin e6 (Ce6), which emits fluorescence in the infrared region, was introduced as the luminescence center, and the addition of copper ion (Cu2+) and As(V) provoked a regular change in fluorescence at 652 nm, whereas that of As(III) was 665 nm, which was used to optionally detect Cu2+, arsenic (As(III), and As(V)). The limit of detection (LOD) values were 0.212 μM, 0.089 ppm, and 1.375 ppb for Cu2+, As(III), and As(V), respectively. The developed method can be used to determine Cu2+ and arsenic in water and soil with good sensitivity and selectivity. The 1:1 stoichiometry of Ce6 with Cu2+ was obtained from the Job plot that was developed from UV–visible spectra. The binding constants for Cu2+ and As(V) were established to be 1.248 × 105 M−1 and 2.35 × 1012 M−2, respectively, using B–H (Benesi–Hildebrand) plots. Fluorescence lifetimes, B–H plots, FT–IR, and 1H-NMR were used to postulate the mechanism of Cu2+ fluorescence quenching and As(V) fluorescence restoration and the interactions of the two ions with the Ce6 molecule.

A core-shell structure UiO-66-(OH)2@UiO-66-NH2 (MOF@MOF) nanoparticle was synthesized through a simple hydrothermal method and employed as an adsorbent and laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) matrix for the quantitative analysis of rhubarb anthraquinones (RAs). The properties of the materials were characterized by field emission scanning electron microscopy, transmission electron microscopy, energy-dispersive X-ray spectroscopy, X-ray powder diffraction, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, thermogravimetric analysis, and Brunauer-Emmett-Teller. The results indicate that MOF@MOF is regular octahedral structure with a size distribution of about 100 nm, having large BET specific surface area (920 m2/g). Using the MOF@MOF as a matrix shows lower background interference, higher sensitivity, and better storage stability than that of traditional matrices. The MOF@MOF matrix exhibits excellent salt tolerance even under a NaCl concentration of 150 mM. Then, the enrichment conditions were optimized, and the adsorption time of 10 min, adsorption temperature of 40 °C and adsorbent amount of 100 μg were selected. In addition, the possible mechanism of MOF@MOF as an adsorbent and matrix was discussed. Finally, the MOF@MOF nanoparticle was employed as a matrix for the sensitive MALDI-TOF-MS analysis of RAs in spiked rabbit plasma, and the recoveries are in the range of 88.3–101.5% with RSD ≤9.9%. In short, the novel MOF@MOF matrix has demonstrated its potential in the analysis of small-molecule compounds in biological samples.

Colorimetric sensors have attracted considerable attention in many sensing applications because of their specificity, high sensitivity, cost-effectiveness, ease of use, rapid analysis, simplicity of operation, and clear visibility to the naked eye [...]

In this work, dopamine (DA) was polymerized on the surface of CuO nanoparticles (CuO NPs) to form a molecularly imprinted polymer (MIP@PDA/CuO NPs) for the colorimetric detection of astragaloside-IV (AS-IV). The synthesis process of MIP is simple and easy to operate, without adding other monomers or initiators. CuO NPs has high peroxidase (POD)-like activity that can catalyze the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB) to generate oxidized TMB (OxTMB) in the presence of H2O2, having a maximum ultraviolet-visible (UV-Vis) absorption peak at 652 nm. The AS-IV can specifically bind to the surface imprinted cavities and prevent the entry of TMB and H2O2, which will lead to the inhibition of the catalytic reaction. Therefore, a new approach based on the POD-like activity of MIP@PDA/CuO NPs for AS-IV detection was developed with a linear range from 0.000341 to 1.024 mg/mL. The LOD and LOQ are 0.000991 and 0.000341 mg/mL, respectively. The developed method can accurately determine AS-IV in Huangqi Granules and different batches of Ganweikang Tablets, which are similar to the results measured by HPLC-ELSD and meet the requirements of Chinese Pharmacopoeia (2020 edition) for the amount of AS-IV in Huangqi Granules. The combination of MIP with CuO NPs not only endows the detection of AS-IV with high selectivity and reliability, but also expands the application of nanozymes in the detection of small-molecule compounds that have weak UV absorption, and do not have reducibility or oxidation properties.

Phenolic compounds are one of the main organic pollutants in the environment that can seriously affect ecosystems, even at very low concentrations. Due to the resistance of phenolic compounds to microorganisms, conventional biological treatment methods face challenges in effectively addressing this pollution problem. In this study, a novel laccase mimic (Tris-Cu nanozyme) is prepared using a simple and rapid synthesis strategy based on the coordination of copper ions and amino groups in Tris(hydroxymethyl)aminomethane (Tris). It is found that the Tris-Cu nanozyme exhibits good catalytic activity against a variety of phenolic compounds, the Km, Vmax and Kcat are determined to be 0.18 mM, 15.62 μM·min−1 and 1.57 × 107 min−1 using 2,4-dichlorophenol (2,4-DP) as the substrate, respectively. Then, based on the laccase-like activity of the Tris-Cu nanozyme, a novel colorimetric method for 2,4-DP (the limit of detection (LOD) = 2.4 μM, S/N = 3) detection in the range of 10–400 μM was established, and its accuracy was verified by analyzing tap and lake water samples. In addition, the Tris-Cu nanozyme shows excellent removal abilities for six phenolic compounds in experiments. The removal percentages for 2,4-DP, 2-chlorophenol (2-CP), phenol, resorcinol, 2,6-dimethoxyphenol (2,6-DOP), and bisphenol A (BPA) are 100%, 100%, 100%, 100%, 87%, and 81% at 1 h, respectively. In the simulated effluent, the Tris-Cu nanozyme maintains its efficient catalytic activity towards 2,4-DP, with a degradation percentage of 76.36% at 7 min and a reaction rate constant (k0) of 0.2304 min−1. Therefore, this metal–organic complex shows promise for applications in the monitoring and degrading of environmental pollutants.

A fast, simple, sensitive, and selective colorimetric method for the detection of Cu2+ was developed using Prussian blue/Turnbull’s blue nanoparticles (PBNPs/TBNPs) as the probe. The colorimetric sensor is based on the following principle. Cu2+ can induce the aggregation of L-cysteine (L-cys) modified-PBNPs/TBNPs (L-cys-PBNPs/TBNPs), resulting in an obvious red shift of its maximum absorption peak. Thus, the concentration of Cu2+ can be determined based on the peak shift in the UV–Vis spectra. The optimal pH, concentration of L-cys, reaction temperature between L-cys-PBNPs/TBNPs and Cu2+, the formation time of L-cys-PBNPs/TBNPs, and the reaction time between L-cys-PBNPs/TBNPs and Cu2+ of the method were determined to be pH 4.5, 2.0 mM, 20 °C, 5.0 min, and 2.0 min, respectively. A good linearity for the colorimetric determination of Cu2+ at the range of 0.25–2.5 μM (R2 = 0.986) was obtained, with a limit of detection (LOD) of 0.12 μM. Moreover, the negligible response of other metal ions demonstrates good selectivity and specificity of the sensor. In addition, the method was employed in the detection of Cu2+ in lake water samples, and the spiked recoveries are in the range of 96.7–106.6% with a relative standard deviation less than 7.4%. Therefore, the colorimetric method is applicable for Cu2+ detection in real water samples of high sensitivity and selectivity.

A biomimetic mineralization method was used in the facile and rapid preparation of nanoflowers for immobilizing alcohol dehydrogenase (ADH). The method mainly uses ADH as an organic component and zinc phosphate as an inorganic component to prepare flower-like ADH/Zn3(PO4)2 organic-inorganic hybrid nanoflowers (HNFs) with the high specific surface area through a self-assembly process. The synthesis conditions of the ADH HNFs were optimized and its morphology was characterized. Under the optimum enzymatic reaction conditions, the Michaelis-Menten constant (Km) of ADH HNFs (β-NAD+ as substrate) was measured to be 3.54 mM, and the half-maximal inhibitory concentration (IC50) of the positive control ranitidine (0.2–0.8 mM) was determined to be 0.49 mM. Subsequently, the inhibitory activity of natural medicine Penthorum chinense Pursh and nine small-molecule compounds on ADH was evaluated using ADH HNFs. The inhibition percentage of the aqueous extract of P. chinense is 57.9%. The vanillic acid, protocatechuic acid, gallic acid, and naringenin have obvious inhibitory effects on ADH, and their percentages of inhibition are 55.1%, 68.3%, 61.9%, and 75.5%, respectively. Moreover, molecular docking analysis was applied to explore the binding modes and sites of the four most active small-molecule compounds to ADH. The results of this study can broaden the application of immobilized enzymes through biomimetic mineralization, and provide a reference for the discovery of ADH inhibitors from natural products.

A core–shell-derived structural magnetic zeolite imidazolate framework-67 (Fe3O4-COOH@ZIF-67) nanocomposite was fabricated through a single-step coating of zeolite imidazolate framework-67 on glutaric anhydride-functionalized Fe3O4 nanosphere for the magnetic solid-phase extraction (MSPE) of theophylline (TP). The Fe3O4-COOH@ZIF-67 nanocomposite was characterized through scanning electron microscopy, transmission electron microscopy, energy dispersive X-ray spectrometry, Fourier transform infrared spectroscopy, Zeta potential analysis, X-ray diffraction, Brunauer–Emmett–Teller, and vibrating sample magnetometer. The material has a high specific surface area and good magnetism, which maintains the regular dodecahedron structure of ZIF-67 without being destroyed by the addition of Fe3O4-COOH nanospheres. The Fe3O4-COOH@ZIF-67 can rapidly adsorb TP mainly through the strong coordination interaction between undercoordinated Co2+ on ZIF-67 and –NH from imidazole of TP. The adsorption and desorption conditions, such as the amount of adsorbent, adsorption time, pH value, and elution solvent, were optimized. The kinetics of TP adsorption on Fe3O4-COOH@ZIF-67 was found to follow pseudo-second-order kinetics. The Langmuir model fits the adsorption data well and the maximum adsorption capacity is 1764 mg/g. Finally, the developed MSPE-HPLC method was applied in the enrichment and analysis of TP in four tea samples and rabbit plasma. TP was not detected in oolong tea and rabbit plasma, and its contents in jasmine tea, black tea, and green tea are 5.80, 4.31, and 1.53 μg/g, respectively. The recoveries of spiked samples are between 74.41% and 86.07% with RSD in the range of 0.81–3.83%. The adsorption performance of Fe3O4-COOH@ZIF-67 nanocomposite was nearly unchanged after being stored at room temperature for at least 80 days and two consecutive adsorption–desorption cycles. The results demonstrate that Fe3O4-COOH@ZIF-67 nanocomposite is a promising magnetic adsorbent for the preconcentration of TP in complex samples.

Small-molecule enzyme mimics as biocatalysts have been extensively applied in diverse colorimetric sensors fabrication. However, excavating potential organic enzyme mimics with high catalytic activity still remains challenging. In this study, the peroxidase mimicking activity of nicotinic acid (VB3) was demonstrated for the first time through chromogenic substrate 3, 3′, 5, 5′-tetramethylbenzidine (TMB) at the existence of hydrogen peroxide (H2O2). The catalytic activity of VB3 kept more than 80% of its optimum activity in a broad pH range of 3.0–9.0. In addition, the kinetic parameter (Michaelis constant, Km = 0.037 mM) of VB3 catalysis to H2O2 is smaller than natural horseradish peroxidase (HRP) and previously reported peroxidase mimics. The catalytic mechanism of VB3 is mainly attributed to the active species of hydroxyl radical (·OH) and partially attributed to the superoxide free radicals (·O2−). A convenient and sensitive colorimetric method based on VB3-H2O2-TMB chromogenic system for H2O2 and glutathione detection was fabricated with the linear ranges of 5.0–100.0 μM and 5.0–50.0 μM, respectively. In short, this work will not only bring new enlightenment on the physiological functions and practical applications in the analytical field of VB3, but also provide a new type of structural reference for small-molecule enzyme mimics.

In this study, a copper hexacyanoferrate nanoparticle with excellent oxidase-mimetic behaviour has been synthesized through a simple precipitation method. The synthesized copper hexacyanoferrate nanoparticle has intrinsic oxidase-like activity, which can catalyze the chromogenic reaction of 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulphonate) through an O2•− reactive oxygen-species-participated process. On the other hand, K3[Fe(CN)6] can be reduced by ascorbic acid (AA) to produce K4[Fe(CN)6], thereby inhibiting the formation of the copper hexacyanoferrate nanoparticles. Furthermore, ascorbate oxidase (AAO) can catalyze the oxidation of AA to produce dehydroascorbic acid, which cannot reduce K3[Fe(CN)6]. Thus, a system for an AAO-regulated in situ formation of copper hexacyanoferrate nanoparticles was constructed by coupling a prepared copper hexacyanoferrate nanozyme with AA for the detection of AAO activity. This colorimetric sensing assay shows high sensitivity and selectivity for the detection of AAO activity (the limit of detection is 0.52 U/L) with a linear range of 1.1–35.7 U/L. Finally, the developed method was applied to detect the activity of AAO in normal human serum with a satisfactory sample spiked recovery (87.4–108.8%). In short, this study provides a good strategy for the construction of nanozyme-based multi-enzyme cascade-signal amplification assay.

Abstract 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 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 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 (

In this study, the personal glucose meter (PGM) was first used as a fast and user-friendly meter for analyzing catechol (CA) based on the reduction of the mediator K3[Fe(CN)6] to K4[Fe(CN)6] in the glucose test strip. Then, an easy, low-cost, and convenient PGM-based method for detecting tyrosinase (TYR) activity and sodium benzoate (SBA) was developed on the basis of the TYR-catalyzed reaction. In this method, CA is oxidized to form o-benzoquinone by TYR, thereby reducing the residual amount of CA and the PGM readout. On the other hand, SBA can inhibit the oxidation of CA catalyzed by TYR and increase the residual amount of CA after the enzymatic reaction. Therefore, the activity of TYR is proportional to the difference in the PGM readout of CA, and the concentration of SBA is positively correlated with the residual amount of CA. After the relevant experimental conditions were systematically optimized, the proposed PGM-based method for the detection of TYR and SBA was successfully validated. The liner ranges are 1.0–103.3 U/mL and 6.25–1000 ppm, and the quantification limits are 1.0 U/mL and 6.25 ppm for TYR and SBA, respectively. Moreover, the spiked recovery tests in normal human serum and carbonate beverages (i.e., Cola, Sprite, and Fanta) were performed, and the recoveries (91.6–106.8%) further confirm the applicability of the PGM-based method in real sample analysis.

In this study, the alkaline phosphatase (ALP)-like activity of zeolitic-imidazolate framework-8 (ZIF-8) is reported for the first time. Then, colorimetric sensors for the ascorbic acid oxidase (AAO) and copper ion (Cu2+) detection were developed based on the acetylcholinesterase (AChE)- and ALP-like activities of ZIF-8. The ZIF-8 has good mimetic enzyme activity and exhibits high affinity to the substrates. Its AChE- and ALP-like activities also have good reusability and storage stability. Good linear dependences are obtained in the range of 1.3−250.0 μM (AChE-like activity-based) and 4.5−454.5 μM (ALP-like activity based) for Cu2+ detection. Furthermore, good linear dependence is also obtained based on the ALP-like activity of ZIF-8 for AAO detection in the range of 2.3−454.5 U/L. Their limits of detection (LODs) are calculated to be 0.7 µM, 2.8 µM, and 1.8 U/L, respectively. Finally, the sample spiked recoveries of Cu2+ in tap water, Cu2+, and AAO in human serum and rabbit plasma were measured, and the results are in the range of 80.0−119.3%. In short, the preparation of ZIF-8 is simple, environmentally friendly, and harmless, and can realize highly selective detection of AAO and Cu2+ in an efficient and fast process.

In this study, a simple colorimetric method was established to detect copper ion (Cu2+), sulfathiazole (ST), and glucose based on the acetylcholinesterase (AChE)-like activity of zeolitic imidazolate framework-8 (ZIF-8). The AChE-like activity of ZIF-8 can hydrolyze acetylthiocholine chloride (ATCh) to thiocholine (TCh), which will further react with 5,5′-dithiobis (2-nitrobenzoic acid) (DTNB) to generate 2-nitro-5-thiobenzoic acid (TNB) that has a maximum absorption peak at 405 nm. The effects of different reaction conditions (buffer pH, the volume of ZIF-8, reaction temperature and time, and ATCh concentration) were investigated. Under the optimized conditions, the value of the Michaelis-Menten constant (Km) is measured to be 0.83 mM, which shows a high affinity toward the substrate (ATCh). Meanwhile, the ZIF-8 has good storage stability, which can maintain more than 80.0% of its initial activity after 30 days of storage at room temperature, and the relative standard deviation (RSD) of batch-to-batch (n = 3) is 5.1%. The linear dependences are obtained based on the AChE-like activity of ZIF-8 for the detection of Cu2+, ST, and glucose in the ranges of 0.021–1.34 and 5.38–689.66 µM, 43.10–517.24 µM, and 0.0054–1.40 mM, respectively. The limit of detections (LODs) are calculated to be 20.00 nM, 9.25 µM, and 5.24 µM, respectively. Moreover, the sample spiked recoveries of Cu2+ in lake water, ST in milk, and glucose in strawberry samples were measured, and the results are in the range of 98.4–115.4% with the RSD (n = 3) lower than 3.3%. In addition, the method shows high selectivity in the real sample analysis.

In this work, indoxyl-glucoside was used as the substrate to develop a cost-effective, paper-based analytical device for the fluorescent and colorimetric dual-mode detection of β-glucosidase activity through a smartphone. The β-glucosidase can hydrolyze the colorless substrate indoxyl-glucoside to release indoxyl, which will be self-oxidized to generate green products in the presence of oxygen. Meanwhile, the green products emit bright blue-green fluorescence under ultraviolet–visible light irradiation at 365 nm. Fluorescent or colorimetric images were obtained by a smartphone, and the red-green-blue channels were analyzed by the Adobe Photoshop to quantify the β-glucosidase activity. Under the optimum conditions, the relative fluorescent and colorimetric signals have a good linear relationship with the activity of β-glucosidase, in the range of 0.01–1.00 U/mL and 0.25–5.00 U/mL, and the limits of detection are 0.005 U/mL and 0.0668 U/mL, respectively. The activities of β-glucosidase in a crude almond sample measured by the fluorescent and colorimetric methods were 23.62 ± 0.53 U/mL and 23.86 ± 0.25 U/mL, respectively. In addition, the spiked recoveries of normal human serum and crude almond samples were between 87.5% and 118.0%. In short, the paper-based device, combined with a smartphone, can provide a simple, environmentally friendly, and low-cost method for the fluorescent and colorimetric dual-mode detection of β-glucosidase activity.

In this study, a paper-based sensor was developed for the detection of hydrogen-peroxide-related biomarkers, with glucose oxidase catalyzing as an example. Potassium iodide can catalyze the oxidation of 3,3′,5,5′-tetramethylbenzidine in the presence of hydrogen peroxide to colorize the paper-based biosensor detection area, which was imaged by a scanner, and the color intensity was analyzed by the Adobe Photoshop. Under the optimal conditions, the color intensity shows a good linear relationship with hydrogen peroxide and glucose concentrations in the ranges of 0.1–5.0 mM and 0.5–6.0 mM, respectively. The detection limit of hydrogen peroxide is 0.03 mM and the limit of quantification of glucose is 0.5 mM. Besides, the method was employed in measuring glucose concentration in fruit samples, and the spiked recoveries are in the range of 95.4–106.1%. This method is cost-effective, environmentally friendly, and easy to be operated, which is expected to realize the point-of-care testing of more hydrogen-peroxide-related biomarkers.

Abstract Differential proteomics, which has been widely used in studying of traditional Chinese medicines (TCMs) during the past 10 years, is a powerful tool to visualize differentially expressed proteins and analyzes their functions. In this paper, the applications of differential proteomics in exploring the action mechanisms of TCMs on various diseases including cancers, cardiovascular diseases, diabetes, liver diseases, kidney disorders and obesity, etc. were reviewed. Furthermore, differential proteomics in studying of TCMs identification, toxicity, processing and compatibility mechanisms were also included. This review will provide information for the further applications of differential proteomics in TCMs studies.

The peroxidase-like activity of vitamin B6 (VB6) was firstly demonstrated by catalyzing the peroxidase chromogenic substrate 3,3′,5,5′-tetramethylbenzidine (TMB) at the existence of H2O2. The influence of different factors on the catalytic property of VB6, including pH, temperature, VB6 concentration, and incubation time, were investigated. The steady-state kinetic study results indicate that VB6 possesses higher affinity to H2O2 than natural horseradish peroxidase and some other peroxidase mimics. Besides, the radical quenching experiment results confirm that hydroxyl radical (•OH) accounts for the catalytic process. Based on the excellent peroxidase-like catalytic activity of VB6, the colorimetric methods for H2O2 and gallic acid (GA) detection were developed by measuring the absorbance variance of the catalytic system. Under the optimal conditions, the linear ranges of the methods for H2O2 and GA determination with good selectivity are 50.0–600.0 μM and 10.0–50.0 μM, respectively. In addition, the developed method was applied in the detection of H2O2 in milk samples and evaluation of total antioxidant capacity of different tea infusions. This study may broaden the application prospect of VB6 in environmental and biomedical analysis fields, contribute to profound insight of the physiological functions of VB6, as well as lay foundation for further excavation of small-molecule peroxidase mimics.

The α-glucosidase inhibitor is regarded as one of the most important drugs for the treatment of diabetes, which can control postprandial blood glucose levels via prolonging the carbohydrate digestion time and retarding the carbohydrates’ absorption. The present work aims to establish a facile bioanalytical method, based on α-glucosidase catalyzing the hydrolysis of 2-O-alpha-D-Glucopyranosyl-L-ascorbic acid (AA-2G), for the quantification of acarbose and migliol using a personal glucose meter (PGM). The hydrolysis products (ascorbic acid and glucose) can trigger the reduction of K3[Fe(CN)6] to K4[Fe(CN)6] in the glucose test strips, which results in the formation of the electron, which can be measured by PGM. Thus, ascorbic acid and glucose can be simultaneously measured by a simplified and miniaturized PGM method. However, the products produced by the hydrolysis of AA-2G will be decreased after the addition of acarbose or migliol to inhibit the activity of α-glucosidase, thereby resulting in a decreased PGM readout. After being incubated with α-glucosidase for 3.0 min and enzymatic reaction for 5.0 min, the quantitative detection of acarbose and migliol can be achieved within the ranges of 1.0–30.0 μM with the limit of detection of 0.33 μM and 3.0–33.3 μM with the limit of detection of 1.0 μM, respectively. IC50 values for acarbose and migliol are calculated to be 10.0 μM and 16.0 μM, respectively. The recoveries of the acarbose and migliol spiked with three different concentrations (final concentrations of 10.0, 20.0, and 30.0 μM) in human serum sample are in the ranges of 89.6–114.5% and 93.9–106.5%, respectively. These results demonstrate that the developed PGM method may be useful in future studies on therapeutic monitoring of acarbose and migliol.

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.

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.

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.

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.

Context: Corydalis yanhusuo W.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 using ortho-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

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.

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.

Multi-origin Chinese herbal medicines, with herbs originating from more than one species of plants, is a common phenomenon but an important issue in Traditional Chinese Medicines (TCMs). In the present study, a gas chromatography-mass spectrometry (GC-MS)—based fatty acid profiling approach to rapidly discriminate multi-origin Chinese medicines in terms of species and medicinal parts was proposed and validated using tuberous roots (Curcumae Radix) and rhizomes (Curcumae Rhizoma and Curcumae Longae Rhizoma) derived from four Curcuma species (e.g., C. wenyujin, C. kwangsiensis, C. phaeocaulis and C. longa) as models. Both type and content of fatty acids varied among different species of either tuberous roots or rhizomes, indicating each species has its own fatty acid pattern. Orthogonal partial least squares discriminant analysis (OPLS-DA) and hierarchical clustering analysis (HCA) based on dataset of global fatty acid profiling showed that either tuberous roots or rhizomes samples could be clearly classified into four clusters according to their species. Furthermore, those tested samples could also be discriminated in terms of their medicinal parts (e.g., tuberous root and rhizome). Our findings suggest that the proposed GC-MS-based fatty acid profiling followed by multivariate statistical analysis provides a reliable platform to discriminate multi-origin Chinese herbal medicines according to species and medicinal parts, which will be helpful for ensuring their quality, safety and efficacy.

Free and polymeric carbohydrates in Cordyceps, a valued edible mushroom and well-known traditional Chinese medicine, were determined using stepwise pressurized liquid extraction (PLE) extraction and GC-MS. Based on the optimized PLE conditions, acid hydrolysis and derivatization, ten monosaccharides, namely rhamnose, ribose, arabinose, xylose, mannose, glucose, galactose, mannitol, fructose and sorbose in 13 samples of natural and cultured Cordyceps were qualitatively and quantitatively analyzed and compared with myo-inositol hexaacetate as internal standard. The results showed that natural C. sinensis contained more than 7.99% free mannitol and a small amount of glucose, while its polysaccharides were usually composed of mannose, glucose and galactose with a molar ratio of 1.00:16.61~3.82:1.60~1.28. However, mannitol in cultured C. sinensis and cultured C. militaris were less than 5.83%, and free glucose was only detected in a few samples, while their polysaccharides were mainly composed of mannose, glucose and galactose with molar ratios of 1.00:3.01~1.09:3.30~1.05 and 1.00:2.86~1.28:1.07~0.78, respectively. Natural and cultured Cordyceps could be discriminated by hierarchical clustering analysis based on its free carbohydrate contents.

Background: Existing studies for ferroptosis and prognosis in colorectal cancer (CRC) were limited. In this study, we aim to investigate the prognostic role of ferroptosis markers in patients with CRC and exploration of its micro‐environmental distributions. Methods: Immunohistochemical staining was performed for CRC patients' tissue microarray. Selection and prognostic validation of markers were based on mRNA data from the cancer genome atlas (TCGA) database. Gene Set Enrichment Analysis (GSEA) was performed to indicate relative immune landmarks and hallmarks. Ferroptosis and immune contexture were examined by CIBERSORT. Survival outcomes were analyzed by Kaplan‐Meier analysis and cox analysis. Results: A panel of 42 genes was selected. Through mRNA expression difference and prognosis analysis, GPX4, NOX1 and ACSL4 were selected as candidate markers. By IHC, increased GPX4, decreased NOX1 and decreased FACL4 indicate poor prognosis and worse clinical characteristics. Ferroptosis score based on GPX4, NOX1 and ACSL4 was constructed and validated with high C‐index. Low ferroptosis score can also demonstrate the better progression free survival and better adjuvant chemotherapy (ACT) responsiveness. Moreover, tumor with low ferroptosis score tend to be infiltrated with more CD4+ T cells, CD8+ T cells and less M1 macrophage. Finally, we found that IFN‐γ was potentially the central molecule at the crossroad between ferroptosis and onco‐immune response. Conclusion: Ferroptosis plays important role on CRC tumor progression, ACT response and prognosis. Ferroptosis contributes to immune‐supportive responses and IFN‐γ was the central molecule for this process. [ABSTRACT FROM AUTHOR]

Inflammation is considered to be critical in the pterygium progression and recurrence. However, the underlying molecular mechanism is not well understood. Herein, we investigated the potential role of RNA binding protein human antigen R (HuR) responsible for the impact of inflammation on pterygium development. The expression of HuR and matrix metallopeptidase‐9 (MMP‐9) in pterygium and normal conjunctiva was detected with immunohistochemistry and quantitative reverse transcription polymerase chain reaction (qRT‐PCR). The influence of interleukin‐1β (IL‐1β) on HuR expression and cellular distribution was determined with western blot and immunofluorescence. The pterygium fibroblast (PTF) migration was determined with scratch wound healing assay and Transwell migration assay. MMP‐9 production was determined with qRT‐PCR and gelatin zymography. The interaction between HuR and MMP‐9 was investigated with RNP immunoprecipitation (IP) followed by RT‐PCR and messenger RNA (mRNA) stability analysis. HuR and MMP‐9 expression are elevated in pterygium, especially progressive pterygium compared with normal conjunctiva. IL‐1β could increase the expression and nucleus–cytoplasm shuttle of HuR in cultured PTFs. HuR mediated the stimulatory effect of IL‐1β on PTF migration and MMP‐9 production. HuR bound to MMP‐9 mRNA and in turn increased it stability. Our results suggest that posttranscriptional regulation of MMP‐9 via stabilizing mRNA by HuR might contribute to the stimulatory effect of inflammatory factor IL‐1β on pterygium progression. These findings shed light on the pathogenesis of pterygium and provide a promising target for adjuvant treatment of pterygium. [ABSTRACT FROM AUTHOR]

Aim: Lymph node ratio (LNR) seems to be more precise than classic N stage in classifying cancer stage. Thus, we aim to construct a modified classification system based on LNR for colon cancer without distant metastasis. Methods: This study enrolled two independent cohorts of patients. The primary cohort enrolled 2,152 patients from 2008 to 2013 in Zhongshan Hospital, Fudan University. The validation cohort consisted of 77,406 patients from the Surveillance, Epidemiology, and End Results (SEER) registry from 2004 to 2014. The inclusion criteria were: pathologically confirmed colon cancer, and American Joint Committee on Cancer (AJCC) stage I/II/III. The exclusion criteria included: incomplete follow-up information, rectal cancer, and multiple primary sites. The prognostic value of LNR for overall survival was evaluated. The cutoff value of LNR was determined by the X-tile. Predictive performance of modified classification was determined by the concordance index. Results: After analysis, 0.05 and 0.50 were determined as the best threshold values of LNR. A value of <0.05, 0.05–0.50 and >0.50 was reclassified as the mN0, mN1 and mN2 stage. A modified classification based on mN0, mN1, and mN2 was further constructed for stage I/II/III colon cancer. C-index of the modified classification was statistically more precise than AJCC classification (0.687 versus 0.605, P<0.001). The same results can also be determined in the validation cohort (0.715 versus 0.640, P<0.001). Furthermore, a prognostic nomogram including independent factors was constructed. The constructed nomogram showed good performance according to the calibration curve. Conclusion: The clinical value of LNR level was preferable to classic N stage in colon cancer patients. Our proposed classification based on LNR and AJCC T category can effectively differentiate patients with varied survival outcomes. [ABSTRACT FROM AUTHOR]

In the present study, the anti-platelet aggregation activity of 14 vegetables and fruits was tested in vitro. The aqueous, 90% ethanol and ethyl acetate extracts, as well as concentrated juices of 14 foods (fruits and vegetables) were prepared, and the anti-platelet aggregation activity of those extracts was analyzed on a platelet aggregation analyzer in vitro with adenosine 5'-diphosphate (ADP), bovine thrombin (THR) and arachidonic acid (AA) as aggregation inducers, respectively. Aspirin (ASP) was used as the positive control. A number of the tested foods had inhibitory effects in concentration-dependent manner on platelet aggregations induced by various agonists. Especially, some foods such as lemon, leek, garlic, scallion, ginger, tomato and grapefruit showed good anti-platelet aggregation effect similar or higher than that of positive control group i.e. aspirin (ASP). The results of present study provide scientific reference for reasonable selection of daily dietary with supplementary curative effects or prevention of cardiovascular diseases (CVD). [ABSTRACT FROM AUTHOR]

Background: Cornea is the outmost structure of the eye and exposed directly to the air pollution. However, little is known about the effect of PM2.5 on corneal epithelium, which is critical for maintenance of cornea homeostasis and visual function. Objective: We investigated the influence of PM2.5 exposure on corneal epithelial migration and the possible mechanisms involved in the process. Methods: We observed wound healing in mouse model of cornea abrasion, evaluated the migration and mobility of cultured corneal epithelial cells with wound scratch assay and Transwell migration assay, detected the phosphorylation and interaction of FAK/paxillin with immunofluorescence and immunoprecipitation, and determined the RhoA activity and actin reorganization, in response to PM2.5 exposure. Results: Exposure to PM2.5 remarkably inhibited corneal epithelial cell migration both in mouse model of corneal abrasion and in cell culture model. We found the phosphorylation and interaction of FAK/paxillin, RhoA activity as well as actin reorganization were suppressed by PM2.5 exposure. Moreover, formation of ROS might play a role in the action of PM2.5. Conclusions: PM2.5 exposure could result in delay of corneal epithelium wound healing by inhibiting cell migration, thus more attention should be paid to the potential risk of corneal infection and effort should be made to protect eyes against impairment induced by PM2.5 [ABSTRACT FROM AUTHOR]

Ganoderma lucidum (Chizhi in Chinese) is one of the most valuable and widely used medicinal fungi in traditional Chinese medicines (TCMs). Most of previous studies were focused on the triterpenoids and polysaccharides of G. lucidum, whereas less attention had been paid on the protein, which is another bioactive compound in it. In the present study, protein maps of fourteen G. lucidum samples were comprehensively analyzed by sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional electrophoresis (2-DE). The results indicated that there were significant differences in protein profiles of G. lucidum samples from different origins. Furthermore, previous reported bioactive proteins from the fruiting bodies of G. lucidum, were mainly distributed in 4 taxa (A, B, C and D) based on their molecular weights on the 2-DE maps. The proteins should be considered as marker for the quality control of G. lucidum, because the proteomic variation may affect on their pharmacological activities. [ABSTRACT FROM AUTHOR]

Background: Asini Corii Colla (ACC) (namely donkey hide gelatin, E'jiao in Chinese) was one of the most valuable tonic traditional Chinese medicines which is an infallible remedy to promote hematopoiesis. It should be produced by fresh or dried donkey hide according to Chinese Pharmacopeia (2015 edition) with a long-time decoction, while as donkey and horse (or mule) all belong to equids so their hides or their hide gelatins are share much in common, that cause the difficult in distinguishing raw materials donkey hide from horse/mule hide for manufacturer, and the challenge in the quality evaluation of ACC for regulatory authority to identify the adulterated with horse hide. Objective: To establish an effective quality evaluation methods for ACC focused on the qualitative-based identification of the raw material's authenticity, mainly to identify the species origin of the gelatins. Materials and Methods: DNA extracted from (1) Raw materials (hides of donkey, horse, mule, bovine and pig); (2) Five hide-glues (bovine, pig, donkey, horse and mule hide-glue); (3) 11 batches of ACC commercial products made by different manufactures from local drug stores. Polymerase chain reaction (PCR) method with newly designed horse-specific primers I and primer pair II. Results: Use the primer pair I, a 234 bp target product could be amplified sensitively from the DNA sample of horse/mule adulterated commercial ACC products, though the DNA in commercial products is severely degraded. A 219 bp product could be amplified specifically from the DNA sample of horse/mule hide, while the results were all negative for the DNA templates of donkey hide, its gelatin and ACC products without adulteration. Conclusion: The developed PCR method based on primer I and II provide an effective approach to identify the species origin of highly processed product ACC (primer pair I) as well as to distinguish the raw material donkey hide (primer pair II), which might enlighten a new strategy to the Quality Evaluation of ACC. [ABSTRACT FROM AUTHOR]

Background: As one of the bioactive components in Cordyceps sinensis (CS), proteins were rarely used as index components to study the correlation between the protein components and producing areas of natural CS. Objective: Protein components of 26 natural CS samples produced in Qinghai, Tibet, and Sichuan provinces were analyzed and compared to investigate the relationship among 26 different producing areas. Materials and Methods: Proteins from 26 different producing areas were extracted by Tris-HCl buffer with Triton X-100, and separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional electrophoresis (2-DE). Results: The SDS-PAGE results indicated that the number of protein bands and optical density curves of proteins in 26 CS samples was a bit different. However, the 2-DE results showed that the numbers and abundance of protein spots in protein profiles of 26 samples were obviously different and showed certain association with producing areas. Conclusions: Based on the expression values of matched protein spots, 26 batches of CS samples can be divided into two main categories (Tibet and Qinghai) by hierarchical cluster analysis. [ABSTRACT FROM AUTHOR]

In this study, three different oil phases include 1-butyl-1-methylpyrrolidinium bis(trifluoromethanesulfonyl) imide (BMPy[NTf2]), 1-butyl-3-methylimidazolium hexafluorophosphate (BMImPF6) and n-octane were compared for the MEEKC analysis (SDS as surfactant and n-butanol as co-surfactant) of eight isoflavones from pueraria. The investigated isoflavones can bewell separated by all of those three microemulsion systems after careful optimization, and the MEEKC with n-octane as the oil phase was the best choice (good symmetry and high resolutions of peaks with short analysis time) for the analysis. The optimum conditions of MEEKC method were as follows: 70 mM SDS, 0.7 M n-butanol and 0.5% (w/v) n-octane in 10 mM sodium tetraborate (STB) at pH 8.5, applied voltage was 23 kV and cassette temperature was set at 30°C. And then the developed method was fully validated (limit of detection, limit of quantification, intraday precision, interday precision and recovery) and successfully applied to determine the eight analytes in three Radix Puerariae samples. In addition, although the MEEKC with classic oil phase (n-octane) showed better results for isoflavones analysis in this study, the MEEKC with ionic liquids (BMPy[NTf2] and BMImPF6) also showed great separation potential for analytes, which may be further applied in the analysis of other natural products. [ABSTRACT FROM AUTHOR]

Background: Chinese medicinal herbs may use more than one species of Curcumae Radix (Yujin) is the tuberous roots of Curcumae wenyujin, C kwangsiensis, C phaeocaulis and C longa. This study aimed to characterize the chemical profiles of these different species of Curcumae Radix, and develop a method for rapid discrimination of these species by ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC/Q-TOFMS) combined with multivariate statistical analysis. Methods: The metabolomes of 33 different batches of Curcumae Radix derived from four Curcumae species were profiled by UHPLC/Q-TOFMS. The resulting sample codes, tR-m/z pairs and ion intensities were processed by unsupervised principal component analysis (PCA) and supervised orthogonal partial least squared discriminant analysis (OPLS-DA) to characterize the chemical composition of Curcumae Radix across the four different species. Results: Obvious differences were observed in the chemical compositions of the Curcumae Radix samples derived from the four different species according to PCA and OPLS-DA. These results suggested that curcumin, curcumenone, curcumenol and zederone could be used as unique chemical markers for C longa, C wenyujin, C phaeocaulis and C. kwangsiensis, respectively. Conclusions: This study developed a UHPLC/Q-TOFMS method coupled with multivariate statistical analysis to discriminate between Curcumae Radix samples from four different Curcumae species, i.e., C longa, C wenyujin, C phaeocaulis and C kwangsiensis. Notably, this new approach resulted in the identification of curcumin (a), curcumenone (b), curcumenol (c) and zederone (d) as unique chemical markers for the identification. [ABSTRACT FROM AUTHOR]

Thrombosis is considered to be closely related to several diseases such as atherosclerosis, ischemic heart disease and stroke, as well as rheumatoid arthritis, hyperuricemia, and various inflammatory conditions. More and more studies have been focused on understanding the mechanism of molecular and cellular basis of thrombus formation as well as preventing thrombosis for the treatment of thrombotic diseases. In reality, there is considerable interest in the role of natural products and their bioactive components in the prevention and treatment of thrombosis related disorders. This paper briefly describes the mechanisms of thrombus formation on three aspects, including coagulation system, platelet activation, and aggregation, and change of blood flow conditions. Furthermore, the natural products for antithrombosis by anticoagulation, antiplatelet aggregation, and fibrinolysis were summarized, respectively. [ABSTRACT FROM AUTHOR]

A double-development TLC method has been developed for simultaneous qualitative and quantitative analysis of hydrophilic and lipophilic constituents of Salvia miltiorrhiza (Danshen). The optimized mobile phases dichloromethane-ethyl acetate-formic acid 22:24:10 (v/v) and petroleum ether-ethyl acetate-cyclohexane 25:11:14 (v/v) were used for the double development on nano-silica gel 60F254 plates. Their characteristic TLC profiles were observed under UV light at 254 and 365 nm and the bands were then revealed by reaction with 5% H2SO4 in EtOH. Quantification of twelve compounds was achieved by densitometry at 260 or 290 nm, with reference at 400 nm. Linearity was quite good (R² > 0.99) within the ranges tested. This method could be used for quality control of Danshen. [ABSTRACT FROM AUTHOR]

The Late Permian Shaiwa Group of the Ziyun area of Guizhou, South China is a deep-water facies succession characterized by deep-water assemblages of pelagic radiolarians, foraminifers, bivalves, ammonoids and brachiopods. Here we report 20 brachiopod species in 18 genera from the uppermost Shaiwa Group. This brachiopod fauna is latest Changhsingian in age and dominated by productides. The palaeoecologic and taphonomic analysis reveals that the brachiopod fauna is preserved in situ . The attachment modes and substratum preference demonstrate that the Shaiwa brachiopod fauna comprises admixed elements of deep-water and shallow-water assemblages. The presence of the shallow-water brachiopods in the Shaiwa faunas indicates the involuntary settlement of shallow-water brachiopods. The stressed ecologic pressure, triggered by warming surface waters, restricted ecospace and short food sources, may have forced some shallow-water elements to move to hospitable deep-water settings and others to modify their habiting behaviours and exploit new ecospace in deep-water environments. We infer that the end-Permian global warming and subsequent transgression event may have accounted for the stressed environmental pressure in the shallow-water communities prior to the end-Permian mass extinction. [ABSTRACT FROM AUTHOR]