Cell membrane chromatography coupled online with LC‐MS to screen anti‐anaphylactoid components from Magnolia biondii Pamp. targeting on Mas‐related G protein‐coupled receptor X2

Mas‐related G protein‐coupled receptor X2 was a mast cell–specific receptor mediating anaphylactoid reactions by activating mast cells degranulation, and it was also identified as a target for modulating mast cell–mediated anaphylactoid and inflammatory diseases. The anti‐anaphylactoid drugs used clinically disturb the partial effect of partial mediators released by mast cells. The small molecule of Mas‐related G protein‐coupled receptor X2 specific antagonists may provide therapeutic action for the anaphylactoid and inflammatory diseases in the early stage. In this study, the Mas‐related G protein‐coupled receptor X2 high expression cell membrane chromatography was coupled online with liquid chromatography and mass spectrometry and successfully used to screen anti‐anaphylactoid components from Magnolia biondii Pamp. Fargesin and pinoresinol dimethyl ether were identified as potential anti‐anaphylactoid components. Bioactivity of these two components were investigated by β hexosaminidase and histamine release assays on mast cells, and it was found that these two components could inhibit β hexosaminidase and histamine release in a concentration‐dependent manner. This Mas‐related G protein‐coupled receptor X2 high expression cell membrane chromatography coupled online with liquid chromatography and mass spectrometry system could be applied for screening potential anti‐anaphylactoid components from natural medicinal herbs. This study also provided a powerful system for drug discovery in natural medicinal herbs.     

Comparative pharmacokinetics of prim-O-glucosylcimifugin and cimifugin by liquid chromatography-mass spectrometry after oral administration of Radix Saposhnikoviae extract, cimifugin monomer solution and prim-O-glucosylcimifugin monomer solution to rats

A sensitive and reliable liquid chromatography–mass spectrometry method has been developed and validated for simultaneous determination of cimifugin and prim‐O‐glucosylcimifugin in rat plasma after oral administration of Radix Saposhnikoviae (RS) extract, prim‐O‐glucosylcimifugin monomer solution and cimifugin monomer solution. Plasma samples were pretreated by protein precipitation with acetonitrile containing the internal standards puerarin and daidzein. LC separation was achieved on a Zorbax SB‐C₁₈ column (150 × 4.6 mm i.d., 5 µm) with 0.1% formic acid in water and methanol by isocratic elution. The detection was carried out in select‐ion‐monitoring mode with a positive electrospray ionization interface. The fully validated method was successfully applied to the pharmacokinetic study of the analytes in rats. A bimodal phenomenon appeared in the concentration–time curve of prim‐O‐glucosylcimifugin and cimifugin after oral administration of RS extract. Prim‐O‐glucosylcimifugin mainly transformed to cimifugin when it was absorbed into blood. Both absorption and elimination of cimifugin after oral administration of RS were longer than after administration of single cimifugin. The pharmacokinetic parameters (AUC_0–t, AUC_0–∞ and t_1/2) of prim‐O‐glucosylcimifugin and cimifugin by giving cimifugin monomer solution, prim‐O‐glucosylcimifugin monomer solution and RS extract had significant differences (P < 0.05). Copyright © 2012 John Wiley & Sons, Ltd.     

Screening anti‐allergic components of Astragali Radix using LAD2 cell membrane chromatography coupled online with UHPLC‐ESI‐MS/MS method

Huangqi (Astragali Radix), a traditional Chinese herb, is widely used in clinical therapy in China. In addition, an anti‐allergic effect of constituents in Huangqi has been reported in the scientific literature. In the present study, cell membrane chromatography coupled online with UHPLC‐ESI‐MS/MS method was developed to screen, analyze and identify the anti‐allergic components of Huangqi. The Laboratory of Allergic Disease 2 (LAD2) cell was used to establish cell membrane chromatography, which was combined with UHPLC‐ESI‐MS/MS. The coupled system was then used to screen anti‐allergic components from Huangqi. Effects of active components were verified by histamine release assay. A component retained on the LAD2 cell membrane chromatography was identified as formononetin. Bioactivity of formononetin was investigated by histamine release assay in LAD2 cells, and it was found that formononetin could inhibit histamine release in a dose‐dependent manner from 1 to 100 μm . The LAD2 cell membrane chromatography online with UHPLC‐ESI‐MS/MS method is an effective technique for screening the anti‐allergic components of Huangqi.     

Metabolism studies on prim‐O‐glucosylcimifugin and cimifugin in human liver microsomes by ultra‐performance liquid chromatography quadrupole time‐of‐flight mass spectrometry

Prim‐O‐glucosylcimifugin (PGCN) and cimifugin (CN) are major constituents of Radix Saposhnikoviae that have antipyretic, analgesic and anti‐inflammatory pharmacological activities. However, there were few reports with respect to the metabolism of PGCN and CN in vitro. In this paper, we describe a strategy using ultra‐performance liquid chromatography quadrupole time‐of‐flight mass spectrometry (UPLC‐Q‐TOF‐MS) for fast analysis of the metabolic profile of PGCN and CN in human liver microsomes. In total, five phase I metabolites of PGCN, seven phase I metabolites and two phase II metabolites of CN were identified in the incubation of human liver microsomes. The results revealed that the main phase I metabolic pathways of PGCN were hydroxylation and hydrolysis reactions. The phase I metabolic pathways of CN were found to be hydroxylation, demethylation and dehydrogenation. Meanwhile, the results indicated that O‐glucuronidation was the major metabolic pathway of CN in phase II metabolism. The specific UDP‐glucuronosyltransferase (UGT) enzymes responsible for CN glucuronidation metabolites were identified using recombinant UGT enzymes. The results indicated that UGT1A1, UGT1A9, UGT2B4 and UGT2B7 might play major roles in the glucuronidation of CN. Overall, this study may be useful for the investigation of metabolic mechanism of PGCN and CN, and it can provide reference and evidence for further pharmacodynamic experiments. Copyright © 2016 John Wiley & Sons, Ltd.     

Separation of five compounds from leaves of Andrographis paniculata (Burm. f.) Nees by off‐line two‐dimensional high‐speed counter‐current chromatography combined with gradient and recycling elution

An off‐line two‐dimensional high‐speed counter‐current chromatography method combined with gradient and recycling elution mode was established to isolate terpenoids and flavones from the leaves of Andrographis paniculata (Burm. f.) Nees. By using the solvent systems composed of n‐hexane/ethyl acetate/methanol/water with different volume ratios, five compounds including roseooside, 5,4′‐dihydroxyflavonoid‐7‐O‐β‐d ‐pyranglucuronatebutylester, 7,8‐dimethoxy‐2′‐hydroxy‐5‐O‐β‐d ‐glucopyranosyloxyflavon, 14‐deoxyandrographiside, and andrographolide were successfully isolated. Purities of these isolated compounds were all over 95% as determined by high‐performance liquid chromatography. Their structures were identified by UV, mass spectrometry, and ¹H NMR spectroscopy. It has been demonstrated that the combination of off‐line two‐dimensional high‐speed counter‐current chromatography with different elution modes is an efficient technique to isolate compounds from complex natural product extracts.     

Target‐guided isolation of polar antioxidants from Abelmoschus esculentus (L). Moench by high‐speed counter‐current chromatography method coupled with wavelength switching and extrusion elution mode

An off‐line two‐dimensional high‐speed counter‐current chromatography strategy combined with the wavelength switching technique and extrusion elution mode was successfully developed and applied to the isolation of polar antioxidants from Abelmoschus esculentus (L).Moench. Target‐guided by the result of 2,2‐diphenyl‐1‐picrylhydrazyl screening assay, four antioxidants were obtained with purities over 90% through orthogonal high‐speed counter‐current chromatography separation. UV spectroscopy, mass spectrometry and ¹H NMR spectroscopy were employed to identify their structures, which were assigned as l ‐tryptophan, quercetin‐3‐O‐sophoroside, 5,7,3′,4′‐tetrahydroxyflavonol‐3‐O‐[β‐d ‐rhamnopyranosil‐(1→2)]‐β‐d ‐glucopyranoside, and quercetin‐3‐O‐glucoside. Each monomer exhibited significant antioxidant activity. The results demonstrated that proposed method could be an effective approach to isolate bioactive compounds from complex natural products.     

Conformational analysis of the disaccharide methyl α‐d‐mannopyranosyl‐(1→3)‐2‐O‐acetyl‐β‐d‐mannopyranoside monohydrate

The crystal structure of methyl α‐d ‐mannopyranosyl‐(1→3)‐2‐O‐acetyl‐β‐d ‐mannopyranoside monohydrate, C₁₅H₂₆O₁₂·H₂O, ( II ), has been determined and the structural parameters for its constituent α‐d ‐mannopyranosyl residue compared with those for methyl α‐d ‐mannopyranoside. Mono‐O‐acetylation appears to promote the crystallization of ( II ), inferred from the difficulty in crystallizing methyl α‐d ‐mannopyranosyl‐(1→3)‐β‐d ‐mannopyranoside despite repeated attempts. The conformational properties of the O‐acetyl side chain in ( II ) are similar to those observed in recent studies of peracetylated mannose‐containing oligosaccharides, having a preferred geometry in which the C2—H2 bond eclipses the C=O bond of the acetyl group. The C2—O2 bond in ( II ) elongates by ～0.02 Å upon O‐acetylation. The phi (?) and psi (ψ) torsion angles that dictate the conformation of the internal O‐glycosidic linkage in ( II ) are similar to those determined recently in aqueous solution by NMR spectroscopy for unacetylated ( II ) using the statistical program MA′AT, with a greater disparity found for ψ (Δ = ～16°) than for ? (Δ = ～6°).     

Rapid preparative separation of six bioactive compounds from Gentiana crassicaulis Duthie ex Burk. using microwave‐assisted extraction coupled with high‐speed counter‐current chromatography

A rapid method combining microwave‐assisted extraction (MAE) and high‐speed counter‐current chromatography (HSCCC) was applied for preparative separation of six bioactive compounds including loganic acid ( I ), isoorientin‐4′‐O‐glucoside ( II ), 6′‐O‐β‐d ‐glucopyranosyl gentiopicroside ( III ), swertiamarin ( IV ), gentiopicroside ( V ), sweroside ( VI ) from traditional Tibetan medicine Gentiana crassicaulis Duthie ex Burk. MAE parameters were predicted by central composite design response surface methodology. That is, 5.0 g dried roots of G. crassicaulis were extracted with 50 mL 57.5% aqueous ethanol under 630 W for 3.39 min. The extract (gentian total glycosides) was separated by HSCCC with n‐butanol/ethyl acetate/methanol/1% acetic acid water (7.5:0.5:0.5:3.5, v/v/v/v) using upper phase mobile in tail‐to‐head elution mode. 16.3, 8.8, 12., 25.1, 40.7, and 21.8 mg of compounds I–VI were obtained with high purities in one run from 500 mg of original sample. The purities and identities of separated components were confirmed using HPLC with photo diode array detection and quadrupole TOF‐MS and NMR spectroscopy. The study reveals that response surface methodology is convenient and highly predictive for optimizing extraction process, MAE coupled with HSCCC could be an expeditious method for extraction and separation of phytochemicals from ethnomedicine.     

Synthesis of 2‐(sulfamoylphenyl)‐4′‐(iminoaryl/hetroaryl)‐4‐(4′′‐hydroxyphenyl)‐thiazoles and their O‐glucosides

In continuation of our work, we synthesized 2‐(sulfamoylphenyl)‐4′‐amino‐4‐(4″‐hydroxyphenyl)‐thiazole ( 3a ), which were reacted with various (aryl/hetroaryl) aldehyde to form 2‐(sulfamoylphenyl)‐4′‐(iminoaryl/hetroaryl)‐4‐(4″‐hydroxyphenyl)‐thiazoles ( 4a , 4b , 4c , 4d , 4e , 4f ). Glucosylation of compounds ( 4a , 4b , 4c , 4d , 4e , 4f ) have been done by using acetobromoglucose as a glucosyl donor to afford 2‐(sulfamoylphenyl)‐4′‐(iminoaryl/hetroaryl)‐4‐(2,3,4,6‐tetra‐O‐acetyl‐4″‐O‐β‐D ‐glucosidoxyphenyl)‐thiazoles ( 5a , 5b , 5c , 5d , 5e , 5f ), further on deacetylation to produce 2‐(sulfamoylphenyl)‐4′‐(iminoaryl/hetroaryl)‐4‐(4″‐O‐β‐D ‐glucosidoxyphenyl)‐thiazoles ( 6a , 6b , 6c , 6d , 6e , 6f ). The compounds are confirmed by FTIR, ¹H‐NMR, ¹³C‐NMR, and ES‐Mass spectral analysis. J. Heterocyclic Chem., (2011).     

Rapid characterization and determination of multiple components in Bu‐Shen‐Yi‐Qi‐Fang by high‐performance liquid chromatography coupled to electrospray ionization and quadrupole time‐of‐flight mass spectrometry

In this study, a qualitative and quantitative analysis using high‐performance liquid chromatography coupled to electrospray ionization and quadrupole time‐of‐flight mass spectrometry was performed for the quality control of Bu‐Shen‐Yi‐Qi‐Fang, a traditional Chinese formula used for asthma. Thirty‐four compounds, including flavonoids, isoflavonoids, triterpenoid saponins, and iridoid glycosides were identified or tentatively characterized by comparing their retention times and mass spectra with those of authentic standards or literature data. Sixteen components were considered as the main bioactive constituents of Bu‐Shen‐Yi‐Qi‐Fang and they were chosen as the chemical markers in quantitative analysis, including catalpol, leonuride, calycosin‐7‐O‐β‐d ‐glucoside, hyperoside, acteoside, formononetin‐7‐O‐β‐d ‐glucoside, epimedin A, calycosin, icariin, epimedin B, epimedin C, formononetin, astragaloside IV, astragaloside II, baohuoside‐I, and astragaloside I. The total run time was 20 min. It was found that the calibration curves for all analytes showed good linearity (R² > 0.99) within the test ranges. The relative standard deviations for intra‐ and inter‐day precisions were below 3.9 and 11.7%, respectively. The accuracy was evaluated by the recovery test within the range of 89.20–110.71% with the relative standard deviation < 4.8%. The sample was stable for at least 48 h at 4°C. The results showed that the new approach was effective for the quality control of Bu‐Shen‐Yi‐Qi‐Fang.     

Simultaneous identification of three pseudoallergic components in Danshen injection by using high‐expression Mas‐related G protein coupled receptor X2 cell membrane chromatography coupled online to HPLC–ESI‐MS/MS

Adverse drug reactions of Danshen injection mainly manifested as pseudoallergic reactions. In the present study, salvianolic acid A and a pair of geometric isomers (isosalvianolic acid C and salvianolic acid C) were identified as pseudoallergic components in Danshen injection by a high‐expression Mas‐related G protein coupled receptor X2 cell membrane chromatography coupled online with high‐performance liquid chromatography with electrospray ionization tandem mass spectrometry. Their pseudoallergic activities were evaluated by in vitro assay, which were consistent with the retention times on the cell membrane chromatography column. Salvianolic acid C, the most outstanding compound, was further found to induce pseudoallergic reaction through Mas‐related G protein coupled receptor X2. All the results above indicated that the system developed in this study is an effective method for simultaneously analyzing pseudoallergic components, even those with similar structures and the microcomponents in complex samples (salvianolic acid C in Danshen injection).     

Ring‐opening polymerization of benzylated 1,6‐anhydro‐β‐D‐lactose and specific biological activities of sulfated (1→6)‐α‐D‐lactopyranans

A new anhydro disaccharide monomer, 1,6‐anhydro‐2,3‐di‐o‐benzyl‐4‐o‐(2′,3′,4′,6′‐tetra‐o‐benzyl‐β‐D ‐galactopyranosyl)‐β‐D ‐glucopyranose (benzylated 1,6‐anhydro lactose (LSHBE)), was synthesized from D ‐lactose to investigate the polymerizability and biological activities of the resulting branched polysaccharides. The ring‐opening polymerization of LSHBE was carried out with phosphorus pentafluoride as a catalyst under high vacuum to give a stereoregular benzylated (1 → 6)‐α‐D ‐lactopyranan. The molecular weights of poly(LSHBE)s increased with an increase in the amount of CH₂Cl₂ solvent, and polymerization temperatures were affected in both molecular weights and yields of the polymers. The copolymerization of LSHBE with benzylated 1,6‐anhydro‐β‐D ‐glucopyranose (LGTBE) gave the corresponding copolysacchrides having different proportions of lactose and glucose units in good yields. After debenzylation to recover hydroxyl groups and then sulfation, sulfated homopoly(lactose)s and copoly(lactose and glucose)s were obtained. Sulfated homopoly(lactose)s had moderate anti‐HIV (EC₅₀ = 5.9 and 1.3 μg/mL) and blood anticoagulant activities (AA = 18 and 13 unit/mg), respectively. Sulfated copoly(lactose and glucose) having 15 mol % lactose units gave high anti‐HIV and blood anticoagulant activities of 0.3 μg/mL and 54 unit/mg, respectively. These biological results suggest that the distance between branched units on the main chain plays an important role in the anti‐HIV and blood anticoagulant activities. © 2008 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 47: 913–924, 2009     

Simultaneous quantification and rat pharmacokinetics of formononetin‐7‐O‐β‐d‐glucoside and its metabolite formononetin by high‐performance liquid chromatography–tandem mass spectrometry

Formononetin‐7‐O‐β‐d ‐glucoside has been proved to have significant anti‐inflammatory effect. To evaluate its rat pharmacokinetics, a rapid, sensitive, and specific liquid chromatography–tandem mass spectrometry method has been developed and validated for the quantification of formononetin‐7‐O‐β‐d ‐glucoside and its main metabolite formononetin in rat plasma. Samples were pretreated using a simple protein precipitation and the chromatographic separation was performed on a C₁₈ column by a gradient elution using a mobile phase consisting of water and acetonitrile both containing 0.1% formic acid. Both analytes were detected using a tandem mass spectrometer in positive multiple reaction monitoring mode. The assay showed wide linear dynamic ranges of both 0.10–100 ng/mL, with acceptable intra‐ and inter‐batch accuracy and precision. The lower limits of quantification were both 0.10 ng/mL using 50 μL of rat plasma for two analytes. The method has been successfully used to investigate the oral pharmacokinetic profiles of both analytes in rats. After oral administration of formononetin‐7‐O‐β‐d ‐glucoside at the dose of 50 mg/kg, it was rapidly absorbed in vivo and metabolized to its metabolite formononetin. The plasma concentration‐time profiles both showed double‐peak phenomena, which would be attributed to the strong enterohepatic circulation of formononetin‐7‐O‐β‐d ‐glucoside.     

Pyranoside‐into‐Furanoside Rearrangement: New Reaction in Carbohydrate Chemistry and Its Application in Oligosaccharide Synthesis

Great interest in natural furanoside‐containing compounds has challenged the development of preparative methods for their synthesis. Herein a novel reaction in carbohydrate chemistry, namely a pyranoside‐into‐furanoside (PIF) rearrangement permitting the transformation of selectively O‐substituted pyranosides into the corresponding furanosides is reported. The discovered process includes acid‐promoted sulfation accompanied by rearrangement of the pyranoside ring into a furanoside ring followed by solvolytic O‐desulfation. This process, which has no analogy in organic chemistry, was shown to be a very useful tool for the synthesis of furanoside‐containing complex oligosaccharides, which was demonstrated by synthesizing disaccharide derivatives α‐D ‐Galp‐(1→3)‐β‐D ‐ Galf ‐OPr, 3‐O‐s ‐lactyl‐β‐D ‐ Galf ‐(1→3)‐β‐D ‐Glcp‐OPr, and α‐L ‐ Fucf ‐(1→4)‐β‐D ‐GlcpA‐OPr related to polysaccharides from the bacteria Klebsiella pneumoniae and Enterococcus faecalis and the brown seaweed Chordaria flagelliformis.     

Anti‐inflammatory bioactive equivalence of combinatorial components β‐carboline alkaloids identified in Arenaria kansuensis by two‐dimensional chromatography and solid‐phase extraction coupled with liquid–liquid extraction enrichment technology

Bioactive equivalent combinatorial components play a critical role in herbal medicines. However, how to discover and enrich them efficiently is a question for herbal pharmaceuticals researchers. In our work, a novel two‐dimensional reversed‐phase/hydrophilic interaction high‐performance liquid chromatography method was established to perform real‐time components trapping and combining for preparation and isolation of coeluting components. Arenaria kansuensis was taken as an example, and solid‐phase extraction coupled with liquid–liquid extraction as a simple and efficient method for enriching trace components, reversed phase column coupled with hydrophilic interaction liquid chromatography XAmide column as two‐dimensional chromatography technology for isolation and preparation of coeluting constituents, enzyme‐linked immune‐sorbent assay as bio‐guided assay, and anti‐inflammatory bioactivity evaluation for bioactive constituents. A combination of 12 β‐carboline alkaloids was identified as anti‐inflammatory bioactive equivalent combinatorial components from A. kansuensis , which accounts for 1.9% w/w of original A. kansuensis . This work answers the key question of which are real anti‐inflammatory components from A. kansuensis and provides a fast and efficient approach for discovering and enriching trace β‐carboline alkaloids from herbal medicines for the first time. More importantly, the discovery of bioactive equivalent combinatorial components could improve the quality control of herbal products and inspire a herbal medicine based on combinatorial therapeutics.     

The human mast cell line‐1 cell membrane chromatography coupled with HPLC‐ESI‐MS/MS method for screening potentical anaphylactic components from chuanxinlian injection

Chuanxinlian injection is a traditional Chinese medicine injection widely used in China to treat sore throat, cough and dysentery, although a high occurrence of severe adverse reactions has been reported in clinical practice in recent years. In the present study, a human mast cell line‐1 cell membrane chromatography coupled with HPLC‐ESI‐MS/MS method was established to screen and identify potentical anaphylactic components in chuanxinlian injection, and the dehydroandrographolide was identified as a potential anaphylactic component. In vitro anaphylactic assay showed that intracellular Ca²⁺ concentration clearly increased under dehydroandrographolide (100 μm ) treatment. β ‐Hexosaminidase and histamine release in human mast cell line‐1 cells were both markedly enhanced with increased concentrations of dehydroandrographolide, confirming the anaphylactic activity of dehydroandrographolide. The application for chuanxinlian injection in this study suggested that the developed human mast cell line‐1 cell membrane chromatography coupled with HPLC‐ESI‐MS/MS system may be effective and rapid for screening the potentical anaphylactic components from complex samples.     

Development of a high‐performance liquid chromatography procedure to identify known and detect novel C‐13 oligosaccharide moieties in diterpene glycosides from Stevia rebaudiana (Bertoni) Bertoni (Asteraceae): Structure elucidation of rebaudiosides V and W

As an aid for structure elucidation of new steviol glycosides, reversed‐phase C18 high‐performance liquid chromatography method was developed with several previously characterized diterpene glycosides, to identify known and detect novel aglycone‐C13 oligosaccharide moieties and indirectly identify C‐19 interlinkages. Elution order of several diterpene glycosides and their aglycone‐C13 oligosaccharide substituted with different sugar arrangements were also summarized. Comparison of the retention time of a product obtained after alkaline hydrolysis with the aglycone‐C‐13 portions of known compounds reported herein allowed us to deduce the exact positions of the sugars in the C‐13 oligosaccharide portion. The elution position of several steviol glycosides with an ent‐kaurene skeleton was helpful to describe an identification key. Two previously uncharacterized diterpene glycosides together with two known compounds were isolated from a commercial Stevia rebaudiana leaf extract. One was found to be 13‐[(2‐O‐β‐d ‐xylopyranosyl‐ 3‐O‐β‐d ‐glucopyranosyl‐β‐d ‐glucopyranosyl)oxy]ent‐kaur‐16‐en‐19‐oic acid‐(2‐O‐β‐d ‐glucopyranosyl‐β‐d ‐glucopyranosyl) ester (rebaudioside V), whereas the other was determined to be 13‐[(2‐O‐β‐d ‐xylopyranosyl‐ 3‐O‐β‐d ‐glucopyranosyl‐β‐d‐ glucopyranosyl)oxy]ent‐kaur‐16‐en‐19‐oic acid‐(2‐O‐α‐l ‐rhamnopyranosyl‐3‐O‐β‐d ‐glucopyranosyl‐β‐d ‐glucopyranosyl) ester (rebaudioside W). Previously reported compounds were isolated in gram quantities and identified as rebaudioside J and rebaudioside H. In addition, a C‐19 sugar‐free derivative was also prepared from rebaudioside H to afford rebaudioside H₁. Chemical structures were partially determined by the high‐performance liquid chromatography method and unambiguously characterized by using one‐dimensional and two‐dimensional nuclear magnetic resonance experiments.     

Separation of α‐amylase inhibitors from Abelmoschus esculentus (L).Moench by on‐line two‐dimensional high‐speed counter‐current chromatography target‐guided by ultrafiltration‐HPLC

In this study, an on‐line two‐dimensional high‐speed counter‐current chromatography system based on a six‐port valve was developed. Target‐guided by ultrafiltration with high‐performance liquid chromatography, the one‐step isolation of three potential α‐amylase inhibitors from Abelmoschus esculentus (L).Moench was achieved by employing the developed orthogonal system and extrusion elution mode. The purities of three potential α‐amylase inhibitors were all over 95% as determined by high‐performance liquid chromatography. Furthermore, UV, mass spectrometry and ¹H NMR spectroscopy were applied to the structural identification of the isolated three target compounds, their structures were assigned as quercetin‐3‐O‐sophoroside (i), 5,7,3′,4′‐tetrahydroxy flavonol‐3‐O‐[β‐d ‐rhamnopyranosil‐(1→2)]‐β‐d ‐glucopyranoside (ii ) and isoquercitrin (iii), respectively. The Results demonstrated that the proposed method was highly efficient to screen and isolate enzyme inhibitors from complex natural products extracts, and on‐line two‐dimensional high‐speed counter‐current chromatography can effectively increase the peak resolution of target compounds.     

Screening antiallergic components from Carthamus tinctorius using rat basophilic leukemia 2H3 cell membrane chromatography combined with high‐performance liquid chromatography and tandem mass spectrometry

Carthamus tinctorius, used in traditional Chinese medicine, has many pharmacological effects, such as anticoagulant effects, antioxidant effects, antiaging effects, regulation of gene expression, and antitumor effects. However, there is no report on the antiallergic effects of the components in C. tinctorius. In the present study, we investigated the antiallergic components of C. tinctorius and its mechanism of action. A rat basophilic leukemia 2H3/cell membrane chromatography coupled online with high‐performance liquid chromatography and tandem mass spectrometry method was developed to screen antiallergic components from C. tinctorius. The screening results showed that Hydroxysafflor yellow A, from C. tinctorius, was the targeted component that retained on the rat basophilic leukemia 2H3/cell membrane chromatography column. We measured the amount of β‐hexosaminidase and histamine released in mast cells and the key markers of degranulation. The release assays showed that Hydroxysafflor yellow A could attenuate the immunoglobulin E induced release of allergic cytokines without affecting cell viability from 1.0 to 50.0 μM. In conclusion, the established rat basophilic leukemia 2H3 cell membrane chromatography coupled with online high‐performance liquid chromatography and tandem mass spectrometry method successfully screened and identified Hydroxysafflor yellow A from C. tinctorius as a potential antiallergic component. Pharmacological analysis elucidated that Hydroxysafflor yellow A is an effective natural component for inhibiting immunoglobulin E–antigen‐mediated degranulation.     

3‐Deoxy‐β‐d‐ribo‐hexopyranose (3‐deoxy‐β‐d‐glucopyranose)

The β‐pyranose form, (III), of 3‐deoxy‐d ‐ribo‐hexose (3‐deoxy‐d ‐glucose), C₆H₁₂O₅, crystallizes from water at 298 K in a slightly distorted ⁴C₁ chair conformation. Structural analyses of (III), β‐d ‐glucopyranose, (IV), and 2‐deoxy‐β‐d ‐arabino‐hexopyranose (2‐deoxy‐β‐d ‐glucopyranose), (V), show significantly different C—O bond torsions involving the anomeric carbon, with the H—C—O—H torsion angle approaching an eclipsed conformation in (III) (−10.9°) compared with 32.8 and 32.5° in (IV) and (V), respectively. Ring carbon deoxygenation significantly affects the endo‐ and exocyclic C—C and C—O bond lengths throughout the pyranose ring, with longer bonds generally observed in the monodeoxygenated species (III) and (V) compared with (IV). These structural changes are attributed to differences in exocyclic C—O bond conformations and/or hydrogen‐bonding patterns superimposed on the direct (intrinsic) effect of monodeoxygenation. The exocyclic hydroxymethyl conformation in (III) (gt) differs from that observed in (IV) and (V) (gg).