Screening Of Bioactive Components Research Articles

A green, efficient and stable platform based on hyperbranched quaternized hydrothermal magnetic chitosan nanospheres integrated cytomembranes for screening drug candidates from natural products

Compared with traditional tedious organic solvent-assisted separation process in natural medicinal chemistry, cytomembrane (CM) fishing technique became a more appealing and greener choice for screening bioactive components from natural products. However, its large-scale practical value was greatly weakened by the easy fall-off of CMs from magnetic supports, rooted in the instability of common Fe3O4 particles and their insufficient interaction with CMs. In this research, a new green biostable platform was developed for drug screening through the integration of hyperbranched quaternized hydrothermal magnetic carbon spheres (HQ-HMCSs) and CMs. The positive-charged HQ-HMCSs were constructed by chitosan-based hydrothermal carbonization onto Fe3O4 nanospheres and subsequent aqueous hyperbranching quaternization with 1,4-butanediol diglycidyl ether and methylamine. The strong interaction between HQ-HMCSs and CMs was formed via electrostatic attraction of HQ-HMCSs to negative-charged CMs and covalent linkage derived from the epoxy-amine addition reactions. The chemically stable HMCSs and its integration with CMs contributed to dramatically higher stability and recyclability of bionic nanocomposites. With the fishing of osteoblast CMs integrated HQ-HMCSs, two novel potential anti-osteoporosis compounds, narcissoside and beta-ionone, were discovered from Hippophae rhamnoides L. Enhanced osteoblast proliferation, alkaline phosphatase, and mineralization levels proved their positive osteogenesis effects. Preliminary pharmacological investigation demonstrated their potential action on membrane proteins of estrogen receptor alpha and insulin-like growth factor 1. Furthermore, beta-ionone showed apparent therapeutic effects on osteogenic lesions in zebrafish. These results provide a green, stable, cost-efficient, and reliable access to rapid discovery of drug leads, which verifiably benefits the design of nanocarbon-based biocomposites with increasingly advanced functionality.

A screening strategy for bioactive components of Bu-Zhong-Yi-Qi-Tang regulating spleen-qi deficiency based on “endobiotics-targets-xenobiotics” association network

Ethnopharmacological relevanceBu-Zhong-Yi-Qi-Tang is a famous traditional Chinese medicine formula that has been prevalent in China for over 700 years to treat spleen-qi deficiency related diseases, such as gastrointestinal and respiratory disorders. However, the bioactive components responsible for regulating spleen-qi deficiency remain unclear and have puzzled many researchers. Aim of the studyThe current study focuses on efficacy evaluation of regulating spleen-qi deficiency and screening the bioactive components of Bu-Zhong-Yi-Qi-Tang. Materials and methodsThe effects of Bu-Zhong-Yi-Qi-Tang were evaluated through blood routine examination, immune organ index, and biochemical analysis. Metabolomics was utilized to analyze the potential endogenous biomarkers (endobiotics) in the plasma, and the prototypes (xenobiotics) of Bu-Zhong-Yi-Qi-Tang in the bio-samples were characterized using ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry. Then, these endobiotics were used as “bait” to predict targets based on network pharmacology and to screen potential bioactive components from the absorbed prototypes in the plasma by constructing an “endobiotics-targets-xenobiotics” association network. Further, the anti-inflammatory activities of representative compounds (calycosin and nobiletin) were validated through poly(I:C)-induced pulmonary inflammation mice model. ResultsBu-Zhong-Yi-Qi-Tang exhibited immunomodulatory and anti-inflammatory activities in spleen-qi deficiency rat, as supported by the observation of increased levels of D-xylose and gastrin in serum, an increase in the thymus index and number of lymphocytes in blood, as well as a reduction in the level of IL-6 in bronchoalveolar lavage fluid. Furthermore, plasma metabolomic analysis revealed a total of 36 Bu-Zhong-Yi-Qi-Tang related endobiotics, which were mainly enriched in primary bile acids biosynthesis, the metabolism of linoleic acid, and the metabolism of phenylalanine pathways. Meanwhile, 95 xenobiotics were characterized in plasma, urine, small intestinal contents, and tissues of spleen-qi deficiency rat after Bu-Zhong-Yi-Qi-Tang treatment. Using an integrated association network, six potential bioactive components of Bu-Zhong-Yi-Qi-Tang were screened. Among them, calycosin was found to significantly reduce the levels of IL-6 and TNF-α in the bronchoalveolar lavage fluid, increase the number of lymphocytes, while nobiletin dramatically decreased the levels of CXCL10, TNF-α, GM-CSF, and IL-6. ConclusionOur study proposed an available strategy for screening bioactive components of BYZQT regulating spleen-qi deficiency based on “endobiotics-targets-xenobiotics” association network.

Screening of trypsin inhibitors in Cotinus coggygria Scop. extract using at-line nanofractionation coupled with semi-preparative reverse-phase liquid chromatography

In this study, an at-line nanofractionation (ANF) platform was successfully fabricated in parallel with mass spectrometry and trypsin inhibitory bioactivity assessment for rapid screening of trypsin inhibitors (TIs) from natural products for the first time. After systematic optimization, the ANF platform was applied to screen and identify TIs in the extract of a traditional Chinese herb, i.e., Cotinus coggygria Scop. The semi-preparative reverse-phase liquid chromatography was used subsequently to further simplify and enrich the insufficiently separated components. After comprehensive evaluation and validation, the ANF platform successfully identified 12 compounds as potential TIs, including 8 flavonoids and 2 organic acids. Additionally, a comparison study was conducted using two other ligand fishing approaches, i.e., capillary monolithic and magnetic beads-based trypsin-immobilized enzyme microreactors, which successfully identified 8 identical flavonoids as TIs. Importantly, the molecular docking study showed the molecular interactions between enzymes and inhibitors, thus strongly supporting the experimental results. Overall, this work has fully demonstrated the feasibility of the established ANF platform for screening TIs from Cotinus coggygria Scop., and proved its great prospects for screening bioactive components from natural products.

Two-dimensional chromatography in screening of bioactive components from natural products.

Screening and analysis of bioactive components from natural products is a fundamental part of new drug development and innovation. Two-dimensional (2D) chromatography has been demonstrated to be an effective method for screening and preparation of specific bioactive components from complex natural products. To collect details of application of 2D chromatography in screening of natural product bioactive components and to outline the research progress of different separation mechanisms and strategies. Three screening strategies based on 2D chromatography are reviewed, including traditional separation-based screening, bioactivity-guided screening and affinity chromatography-based screening. Meanwhile, in order to cover these aspects, selections of different separation mechanisms and modes are also presented. Compared with traditional one-dimensional (1D) chromatography, 2D chromatography has unique advantages in terms of peak capacity and resolution, and it is more effective for screening and identifying bioactive components of complex natural products. Screening of natural bioactive components using 2D chromatography helps separation and analysis of complex samples with greater targeting and relevance, which is very important for development of innovative drug leads.

Screening of hypoglycemic components in Platycladi Cacumen by phytochemical investigation, spectrum-effect relationship, and chemometric methods.

In this work, the hypoglycemic components in Platycladi Cacumen, an essential traditional Chinese medicine, were evaluated by combining phytochemical investigation, spectrum-effect relationship analysis, and chemometric methods. The phytochemical studies on Platycladi Cacumen extract lead to the isolation of 21 potential bioactive compounds. The chromatographic fingerprints of Platycladi Cacumen samples were established by high-performance liquid chromatography. The hypoglycemic effects of Platycladi Cacumen samples were further evaluated by inhibition of α-glucosidase and detected by the high-performance liquid chromatography method. The spectrum-effect relationship study by bivariate correlations analysis and orthogonal partial least squares regression revealed that myricitrin (P9), quercitrin (P13), afzelin (P18), and amentoflavone (P24) were more relevant to the α-glucosidase inhibitory activity. The results of α-glucosidase inhibitory activity of 21 isolated compounds and molecular docking studies also indicated these flavonoids had potent α-glucosidase inhibitory activity. Collectively, the present study established the spectrum-effect relationship mode of Platycladi Cacumen and discovered the major hypoglycemic components, which provides a feasible method for screening bioactive components.

Enhanced stability designs of cell membrane chromatography for screening drug leads

Cell membrane chromatography is an effective method for screening bioactive components acting on specific receptors in complex systems, which maintains the biological activity of the membrane receptors and improves screening efficiency. However, traditional cell membrane chromatography suffers from poor stability, resulting in a limited life span and low reproducibility, greatly limiting the application of this method. To address this problem, cyanuric chloride-decorated silica gel was used for the covalent immobilization of the cell membranes. Cyanuric chloride reacts with amino groups on the cell membranes and membrane receptors to form covalent bonds. In this way, the cell membranes are not easy to fall off. The column life of the cyanuric chloride-decorated epidermal growth factor receptor/cell membrane chromatography column was extended to more than 8 days, whereas the column life of the normal cell membrane chromatography column dropped sharply in the first 3 days. A cyanuric chloride-decorated epidermal growth factor receptor/cell membrane chromatography online HPLC-IT-TOF-MSn system was applied for screening drug leads from Trifolium pratense L. One potential drug lead, formononetin, which acts on the epidermal growth factor receptor, was screened. Our strategy of covalently immobilizing cell membrane receptors also improved the stability of cell membrane chromatography.

Ligand fishing based on tubular microchannel modified with monoamine oxidase B for screening of the enzyme's inhibitors from Crocus sativus and Edgeworthia gardneri.

A novel strategy of performing ligand fishing with enzyme-modified open tubular microchannel was proposed for screening bioactive components present in medicinal plants. Monoamine oxidase B was immobilized onto the surface of the microchannel for the first time to specifically extract its ligands when the plant's extracts solution flows through the channel. The thermal and the storage stability of immobilized monoamine oxidase B were significantly enhanced after immobilization. Crocin I and Ⅱ were extracted from Crocus sativus, and tiliroside was extracted from Edgeworthia gardneri. All the three compounds were inhibitors of the enzyme with the half-maximal inhibitory concentration values of 26.70 ± 0.91, 19.88 ± 2.78, and 15.65 ± 0.85μM, respectively. The enzyme inhibition kinetics and molecular docking were investigated. This is the first report on the inhibitory effects of tiliroside and crocin Ⅱ. The novel ligand fishing method proposed in this work possesses advantages of rapidness, high efficiency, and tiny sample consumption compared to routine ligand fishing, with promising potential for screening active natural products in complex mixtures.

Biochemical potential and screening of bioactive components of Ocimum basilicum

Biochemical potential and screening of bioactive components of Ocimum basilicum

None-Loss Target Release of Biomimetic CaCO3 Nanocomposites for Screening Bioactive Components and Target Proteins

Many traditional Chinese medicines have been clinically proven to have good anticancer effects in recent years. Tripterygium wilfordii, as a classic anticancer herbal medicine, has good effects, bu...

Screening of bioactive components from traditional Chinese medicine by immobilized β2 adrenergic receptor coupled with high performance liquid chromatography/mass spectrometry

Traditional Chinese medicine (TCM) represents a valuable resource for lead compounds discovery. Given the complexity of TCM components, analytical methods play a key role in novel drug development. In our study, we established a high specific and reliable bio-active components screen system, where β2 adrenergic receptor (β2-AR) was immobilized on silica by non-covalent bonds and packed into a stainless steel column (4.6 × 50 mm, 7 μm) to form β2-AR chromatography column. The column was further coupled with high performance liquid chromatography-time of flight tandem mass spectrometry (TOF-MS/MS). By utilizing this strategy, we successfully identified four β2-AR-targeting compounds: tetrahydroberberine, tetrahydrocolumbamine, fumarine and corydaline from Corydalis Rhizome. The association constants between β2-AR and tetrahydroberberine (9.04 × 104/M) as well as fumarine (4.30 × 104/M) were determined by frontal chromatography. We also found that these two compounds shared the identical binding site on immobilized β2-AR with corresponding concentrations of 6.67 × 10−4 M and 5.88 × 10−4 M, respectively. The newly established method represents an efficient tool to identify the target specific natural compounds.

Study the effect of trypsin enzyme activity on the screening of applying frontal affinity chromatography

Frontal affinity chromatography (FAC) combined with enzyme has been widely used for drug screening. In this paper, the effect of target enzyme activity on screening of bioactive compounds was studied through applying FAC. Trypsin with different degree of inactivation were prepared as target enzyme by thermal denaturation. Their primary structure was identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and use Fourier transform infrared (FTIR) and ultraviolet-visible (UV–vis) spectroscopy to detect group structure. Ultimately, it was found that the main structure of enzyme with decreased activity remained unchanged. The oxymatrine and matrine which can interact with trypsin were selected to study their binding to trypsin with different activities in FAC. The results showed that oxymatrine and matrine had a significant difference in the breakthrough volume among seven kinds of columns prepared by trypsins with different activities, at the different concentration. It indicated that trypsins with different activities in FAC could combine with oxymatrine and matrine. The binding constant (Kd) variation between oxymatrine, matrine and trypsin with different activities are 5.520 ± 0.038 and 3.577 ± 0.071, within error range, which indicated that the activity of target enzyme with primary unchanged structure has no effect on screening of bioactive components by FAC.

Quality evaluation of traditional Chinese medicines based on fingerprinting.

The usage of traditional Chinese medicines has expanded globally, but the data about authentication, efficacy, and safety is far from sufficient to meet the criteria supporting their use worldwide due to complexity in the composition. Fingerprinting describes integral characterization and reflects interactive aspects of complex components; therefore, it can offer the possibility of evaluating quality of traditional Chinese medicines following the overall principle. Chemometric techniques introduce multivariate analytical methods into fingerprinting to obtain more information that is useful, which is consistent with the holistic thought and plays an important role in research on the substantial basis. In this review, we will start with three aspects to expound the quality evaluation of traditional Chinese medicines based on fingerprints. The analytical techniques used in developing fingerprints including chromatographic methods, spectroscopic methods, and capillary electrophoresis are introduced. Strategies for fingerprints analysis usually based on chemometric methods including unsupervised and supervised pattern recognition are described. Applications of fingerprints for multi-component quantification, quality control, screening of bioactive components, and fingerprint-efficacy relationship study are also outlined. Finally, we propose challenges and future perspectives of fingerprints in quality evaluation to promote the development of modernization and internationalization of traditional Chinese medicines.

A strategy for screening bioactive components from natural products based on two-dimensional cell membrane chromatography and component-knockout approach

Cell membrane chromatography (CMC) is a bioaffinity chromatographic method used to screen active compounds from natural products. However, since the receptor capacity of CMC column is limited, high content/affinity compounds may cause column overloading and thus lead to ignorance of other positive candidates. For avoiding this effect and comprehensively discovering bioactive components, a strategy based on two-dimensional CMC and component-knockout approach was proposed. As an illustrative case study, red yeast rice (RYR), a rice product with good myocardial protective effect in clinical studies, was selected as the model experimental sample. For discovering its potential cardioprotective compounds, a CMC model with H9c2 rat cardiac myoblasts (H9c2/CMC) with good selectivity, stability and reproducibility was established. By using two-dimensional H9c2/CMC-HPLC coupled with QTOF MS system, three components were firstly screened out. After knocking out high content/affinity compound, another four bioactive compounds were then found. By this two-round screening, column overloading caused by high concentration or infinity compounds was avoided, and trace compounds were enriched. As a result, one pigment and six monacolins from RYR were fished out. The results indicate the proposed strategy might be used to discover active compounds from complex matrix.

Screening bioactive components of Glycyrrhiza uralensis Fisch. with isolated perfused lung extraction and HPLC-ESI-MSn analysis

The isolated perfused rat lung (IPL), coupled with high performance liquid chromatography\ andem mass spectrometry analysis (HPLC–ESI-MSn), has been developed as a tool for screening bioactive components in Glycyrrhiza uralensis Fisch. (GU). First, IPL was perfused with the water extract of GU (EGU), the bioactive components in the EGU would selectively combine to the receptors or channels of lung. By changing the pH of perfused solution, the combined components were eluated and then detected by HPLC-ESI-MSn. Four compounds were detected in the desorption eluate of IPL, among these compounds, liquiritin (1), ononin (2) and glycyrrhizic acid (4) were identified by comparing with the chromatography of the standards, while licorice-saponin G2 (3) were determined by analysis of the structure clearage characterization of mass spectrometry. Then, due to the lack of compound 3 sample, compounds 1, 2 and 4 with respective concentrations of 50 μM, 5 μM, 500 nM, 50 nM and 5 nM were applied to evaluate the protective effect of pulmonary epithelial cells (PEC, A549 cell) injury induced by lipopolysaccharide (LPS) for anti-inflammatory activity assessment. The results showed that except the 5 nM group of compound 1, 5 nM and 50 nM groups of compound 2, all other groups could remarkably inhibit the PEC injury (vs LPS group, 2-500 nM groups: p < 0.05; other groups: p < 0.01), all compound showed the dose-dependent effect. In conclusion, IPL coupled with HPLC-ESI-MSn was successfully used to screen the anti-inflammatory components of GU for the first time. The application of IPL coupled with HPLC-ESI-MSn for screening bioactive components of TCMs is rapid, convenient and reliable, and the isolated perfused technology could be extended to isolated heart, liver, kidney, and so on.

Rapid Investigation and Screening of Bioactive Components in Simo Decoction via LC-Q-TOF-MS and UF-HPLC-MD Methods.

Simo decoction (SMD), as a traditional medicine, is widely used in the treatment of gastrointestinal dysmotility in China. In this study, a combined method of liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS) and ultrafiltration high-performance liquid chromatography molecular docking (UF-HPLC-MD) was efficiently employed to identify and screen bioactive ingredients in SMD. Ninety-four major constituents were identified or tentatively characterized by comparing their retention times and mass spectra with standards or literature data by using LC-Q-TOF-MS, and the ascription of those compounds were classified for the first time. Among them, 13 bioactive ingredients, including norisoboldine, eriocitrin, neoeriocitrin, narirutin, hesperidin, naringin, neohesperidin, hesperitin-7-O-glucoside, linderane, poncirin, costunolide, nobiletin, and tangeretin, were primarily identified as the human serum albumin (HSA) ligands at a range of docking scores from −29.7 to −40.6 kJ/mol by UF-HPLC-MD. The results indicate the systematic identification and screening of HSA ligands from Simo decoction guided by LC-Q-TOF-MS and UF-HPLC-MD represents a feasible and efficient method that could be extended for the identification and screening of other bioactive ingredients from natural medicines.

Screening of bioactive components from traditional Chinese medicines using cell membrane chromatography coupled with mass spectrometry.

Cell membrane chromatography (CMC), as a highly selective type of affinity chromatography, has been demonstrated as an effective method to screen bioactive components acting on specific receptor from a complicated biological system. To review the recent research progress and the technical applications of these analytical methods using CMC combined with gas chromatography-mass spectrometry, (GC/MS) and liquid chromatography-mass spectrometry (LC/MS). In this review, we briefly introduce the CMC offline GC/MS, CMC online GC/MS, CMC offline LC/MS, and CMC online LC/MS system. And the practical application of these technologies is also enumerated. Then the future of these technologies and research methods were discussed. Many bioactive components interacting with specific receptors have been screened and identified in traditional Chinese medicines. CMC technique has been combined with GC/MS and HPLC/MS and these combined systems have been successfully used to screen bioactive components acting on specific receptors from a complicated biological system.

Screening and identification of neuroprotective compounds from Scrophularia buergeriana using cell extraction coupled with LC–MS

In the cell extraction_LC–MS method, when cells are incubated with natural product extracts, bioactive compounds selectively bind to extracellular or intracellular targets. The extracts and major compounds (phenylpropanoids and iridoid glycosides) of Scrophularia buergeriana Miquel have been reported to show neuroprotective effects both in vitro and in vivo. In this study, the cell extraction_LC–MS strategy was applied to screen and identify potential neuroprotective compounds from S. buergeriana by using immortalized mouse hippocampal HT22 cells. The results showed that two known compounds from S. buergeriana selectively bound HT22 cells. Additionally, metabolomics analyses were performed using the Mass Profiler Professional and Limma differential expression package of R to identify significant differences between HT22 cells treated with S. buergeriana and untreated cells. The cell extraction approach more accurately reflects in vivo conditions compared with other methods and can be readily used for screening bioactive components from natural products.

Rapid Screening of Active Components with an Osteoclastic Inhibitory Effect in Herba epimedii Using Quantitative Pattern-Activity Relationships Based on Joint-Action Models.

Screening of bioactive components is important for modernization and quality control of herbal medicines, while the traditional bioassay-guided phytochemical approach is time-consuming and laborious. The presented study proposes a strategy for rapid screening of active components from herbal medicines. As a case study, the quantitative pattern–activity relationship (QPAR) between compounds and the osteoclastic inhibitory effect of Herba epimedii, a widely used herbal medicine in China, were investigated based on joint models. For model construction, standard mixtures data showed that the joint-action models are better than the partial least-squares (PLS) model. Then, the Good2bad value, which could reflect components’ importance based on Monte Carlo sampling, was coupled with the joint-action models for screening of active components. A compound (baohuoside I) and a component composed of compounds with retention times in the 6.9–7.9 min range were selected by our method. Their inhibition rates were higher than icariin, the key bioactive compound in Herba epimedii, which could inhibit osteoclast differentiation and bone resorption in a previous study. Meanwhile, the half-maximal effective concentration, namely, EC50 value of the selected component was 7.54 μg/mL, much smaller than that of baohuoside I—77 μg/mL—which indicated that there is synergistic action between compounds in the selected component. The results clearly show our proposed method is simple and effective in screening the most-bioactive components and compounds, as well as drug-lead components, from herbal medicines.

GC-MS Fingerprinting Combined with Chemometric Methods Reveals Key Bioactive Components in Acori Tatarinowii Rhizoma.

This present study aims to identify the key bioactive components in acorus tatarinowii rhizoma (ATR), a traditional Chinese medicine (TCM) with various bioactivities. Partial least squares regression (PLSR) was employed to describe the relationship between the radical scavenging activity and the volatile components. The PLSR model was improved by outlier elimination and variable selection and was evaluated by 10-fold cross-validation and external validation in this study. Based on the PLSR model, eleven chemical components were identified as the key bioactive components by variable importance in projection. The final PLS regression model with these components has good predictive ability. The Q2 was 0.8284, and the root mean square error for prediction was 2.9641. The results indicated that the eleven components could be a pattern to predict the radical scavenging activity of ATR. In addition, we did not find any specific relationship between the radical scavenging ability and the habitat of the ATRs. This study proposed an efficient strategy to predict bioactive components using the combination of quantitative chromatography fingerprints and PLS regression, and has potential perspective for screening bioactive components in complex analytical systems, such as TCM.

Screening of bioactive components in grape and apple vinegars: Antioxidant and antimicrobial potential

Vinegars contain several bioactive compounds that are characterized according to the type of the raw material, such as grape vinegars and apple vinegars. Liquid chromatography coupled to diode array detection and electrospray ionization tandem mass spectrometry was used for identification and quantification of phenolic compounds. Antioxidant properties of vinegars were determined by 2,2-diphenyl-1-picrylhydrazyl and 2,2′-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid assays. Antimicrobial activities of vinegars were examined with an agar disc diffusion method with Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa. Gallic acid and chlorogenic acid were found to be the major phenolic acids accounting for the largest proportion of the total phenolic acid contents in grape vinegars and apple vinegars. Within the flavonols, quercetin-3-O-galactoside and quercetin were detected as the major compounds in grape vinegars. Apple vinegars were characterized by phloridzin, phloretin and high chlorogenic acid content. Antimicrobial activity results indicated that grape vinegars exhibited higher antimicrobial activity against tested bacterial strains correlated with their higher antioxidant capacity. Copyright © 2017 The Institute of Brewing & Distilling