Corybungines A-K: Isoquinoline alkaloids from Corydalis bungeana with dopamine D2 receptor activity.

Eleven undescribed isoquinoline alkaloids corybungines A-K including a protoberberine-type alkaloid, an isoquinoline alkaloid with a unique 6-norprotoberberine skeleton, one 13,14-seco-protoberberine-type alkaloid, two 1a,14-seco-protoberberine-type alkaloids with a 4-(hydroxymethyl)phenoxy moiety and six aporphine alkaloids, together with seven known alkaloids, have been isolated from the whole herb extract of Corydalis bungeana Turcz. Their structures and absolute configurations were elucidated based on an analysis of spectroscopic data and electronic circular dichroism (ECD) spectra. (R)-stephanine displayed high antagonistic activity against the dopamine D2 receptor with an IC50 value of 0.85 ± 0.09 µM in CHO-D2 cells. Additionally, corybungines D, F, H, (R)-roemerine, (R)-vireakine and (R)-tuduranine showed moderate D2 antagonism (IC50 5.20-26.07 µM). The preliminary structure-activity relationships (SARs) of aporphine alkaloids were discussed.

Structurally diverse isoquinoline and amide alkaloids with dopamine D2 receptor antagonism from Corydalis bungeana.

Four new isoquinoline alkaloids including a benzophenanthridine alkaloid (1), a morphine derivative (2), a narceine-type alkaloid (3) and a simple isoquinoline alkaloid (4), a new amide alkaloid (5) and a new phthalic acid derivative (6), together with eleven known alkaloids (7-17) were obtained from the whole herbs extract of Corydalis bungeana Turcz. Their structures and absolute configurations were elucidated by extensive spectroscopic data analysis including HRESIMS, NMR and electronic circular dichroism (ECD) and ECD calculation. Compounds 1-17 were evaluated for dopamine D2 receptor activity in CHO-D2 cells. Among them, 16 showed the highest antagonistic activity on the D2 receptor with an IC50 value of 2.04 ± 0.01 µM. Compounds 14 and 15 exhibited moderate antagonism with IC50 values of 13.66 ± 2.28 and 31.72 ± 2.52 µM, respectively.

Offline preparative 2-D polar-copolymerized reversed-phase chromatography × zwitterionic hydrophilic interaction chromatography for effective purification of polar compounds from Caulis Polygoni Multiflori.

Polar compounds are abundant in the water decoction of Traditional Chinese Medicines (TCMs) and possess important biological activities. However, purifying these compounds has faced great difficulties largely due to poor retention and insufficient selectivity. To solve this problem, an offline orthogonal 2-D RPLC coupled with hydrophilic interaction chromatography (HILIC) method was developed to achieve purification of polar compounds from Caulis Polygoni Multiflori. A polar-copolymerized XAqua C18 column and a zwitterionic Click XIon column exhibited satisfactory retention and separation for polar compounds. Therefore, they were adopted to construct the offline 2-D LC system. Furthermore, the method presented a high orthogonality, which was calculated to be 72%. The XAqua C18 column was used in the first dimension to fractionate the 5.1 g polar fraction with a recovery of 85% within 10 h. In the second dimension, three representative fractions were purified using the Click XIon column. Finally, three compounds with purity higher than 95% were identified for the first time from this plant. This offline 2-D RPLC/HILIC method was shown to be an effective approach to purifying polar compounds from Caulis Polygoni Multiflori.

The expression level of miR-155 in plasma and peripheral blood mononuclear cells in coronary artery disease patients and the associations of these levels with the apoptosis rate of peripheral blood mononuclear cells.

The aim of the study was to investigate the expression of miR-155 in plasma and peripheral blood mononuclear cells (PBMCs), the effects of miR-155 on the apoptosis rate of PBMCs and the extent of coronary stenosis in coronary artery disease patients. Seventy chest pain patients were divided into three groups by symptoms, signs, auxiliary examination and coronary arteriography: 21 cases in the acute myocardial infarction group (AMI), 23 cases in the Angor pectoris group (AP) and 26 cases in the control group (CT). The peripheral blood mononuclear cells of the patients in the three groups were separated for cell culture. Annexin V/propidium iodide (PI) was performed to analyze cell apoptosis. RT-qPCR was used to evaluate the expression level of miR-155 in plasma and PBMCs. There were significant differences on the expression of miR-155 in plasma and PBMCs, the apoptosis rate of PBMCs, Gensini score and the extent of coronary stenosis among the three groups (P<0.05). The expression of miR-155 in plasma and PBMCs in AMI and AP group were lower than CT group while the AMI group was lower than the AP group (P<0.05). The apoptotic rate of PBMCs, Gensini score and the extent of coronary stenosis of the AMI and AP groups were higher than CT group while the AMI group was higher than the AP group (P<0.05). The expression of miR-155 in plasma was positively correlated with PBMCs, but negatively correlated with the apoptosis rate of PBMCs, Gensini score and the extent of coronary stenosis. The apoptosis rate of PBMCs in patients with coronary heart disease was positively correlated with the degree of coronary artery stenosis and Gensini score. In conclusion, in the patients with coronary heart disease the apoptosis rate of PBMCs was increased and the expression of miR-155 in plasma and PBMCs cells was decreased, which were correlated with the severity of coronary heart disease.

Discovery of novel antagonists on ß2-adrenoceptor from natural products using a label-free cell phenotypic assay.

Label-free cell phenotypic assays were performed to establish a ß2-adrenoceptor (ß2-AR) target model in A431 cells and a ß1-AR target model in transfected HEK293-ß1 cells, using known ß2-AR and ß1-AR agonists and antagonists. A list of natural compounds was screened on the target models, among which seven new compounds were found to be antagonistically active against ß2-AR. After receptor specificity evaluations on hydroxyl carboxylic acid receptor-2 (ΗΧΑ-2), histamine receptor (H1R), and ß1-adrenoceptor (ß1-AR), six out of the seven compounds, including nuciferine, epiberberine, harmaline, harmine, palmatine, and columbamine, exhibited specific antagonistic activity against ß2-AR. Epiberberine and palmatine showed the strongest antagonistic activities against ß2-AR with IC50 values of 2.3 ± 0.2 µM and 2.6 ± 0.3 µM, respectively. Docking palmatine to the crystal structure of human ß2-AR (PDB 5X7D) suggested that the ligand forms a hydrogen bond with N312 and hydrophobic interaction with several amino acid residues in the binding pocket, such as D113 and V114. The kinetic binding profile of palmatine was further investigated using co-stimulation assays. Results suggested that palmatine was a competitive antagonist for ß2-AR. The six novel ß2-AR antagonists provide a promising chemical starting point for identification and optimization of drugs used for treating hypertension, glaucoma, and infantile hemangiomas. This study also lays the foundation for the in-depth investigation of biochemical mechanisms and pharmacological properties of natural compounds.

On-line comprehensive two-dimensional liquid chromatography tandem mass spectrometry for the analysis of Curcuma kwangsiensis.

A novel online comprehensive two-dimensional liquid chromatography (2DLC) coupled with quadrupole time-of-flight (Q-TOF) mass spectrometry (MS) method is developed for the analysis of Curcuma kwangsiensis (C. kwangsiensis) extract. In this system, a newly developed phenyl/tetrazole sulfoether (PTAS) bonded stationary phase was introduced to construct RPLC×RPLC combined with C18. The unique structure endowed PTAS with very different selectivity from C18, reaching a high orthogonality of 93.2%. Moreover, such a combination settled compatibility issues because of the weaker hydrophobic retaining property of PTAS, thus allowing direct interfacing in online configuration. As a result of coupling with the mass spectrometry, a four-dimensional (4D) data plot was presented, in which 439 peaks (containing positive mode and negative mode) were counted, and 105 compounds were grouped and tentatively identified in C. kwangsiensis extract, including 73 unreported ones. Some novel types of compounds with masses exceeding 500 were discovered for the first time. Besides, compared to one-dimensional liquid chromatography (1DLC), the great resolution power of this system allowed separation of more isomers. These results provide supplementary to the material basis of C. Kwangsiensis and in-depth research should be conducted. The configuration of RPLC×RPLC-Q-TOF MS can be a powerful and efficient tool for separation and characterization of chemical substances in complicated herbal extracts.

SAR Studies of N-[2-(1H-Tetrazol-5-yl)phenyl]benzamide Derivatives as Potent G Protein-Coupled Receptor-35 Agonists.

G protein-coupled receptor-35 (GPR35) has emerged as a potential target in the treatment of pain and inflammatory and metabolic diseases. We have discovered a series of potent GPR35 agonists based on a coumarin scaffold and found that the introduction of a 1H-tetrazol-5-yl group significantly increased their potency. We designed and synthesized a new series of N-[2-(1H-tetrazol-5-yl)phenyl]benzamide derivatives through a two-step synthetic approach, and characterized their agonistic activities against GPR35 using a dynamic mass redistribution (DMR) assay. N-(5-bromo-2-(1H-tetrazol-5-yl)phenyl)-4-methoxybenzamide (56) and N-(5-bromo-2-(1H-tetrazol-5-yl)phenyl)-2-fluoro-4-methoxybenzamide (63) displayed the highest agonistic potency agonist GPR35 with an EC50 of 0.059 µM and 0.041 µM, respectively. The physicochemical properties of selected compounds were calculated to evaluate their druglikeness, suggesting that compounds 56 and 63 have good druglike properties. Together, N-[2-(1H-tetrazol-5-yl)phenyl]benzamide derivatives are potentially great candidates for developing potent GPR35 agonists.

Mesoporous silica-carbon composites fabricated by a universal strategy of hydrothermal carbonization: controllable synthesis and applications.

Mesoporous silica-carbon composite materials, with homogeneous and thickness-controllable carbon coating, were synthesized by using a universal strategy of hydrothermal carbonization, and the carbon layer could be coated on the surface of ordered and disordered mesoporous silica. The electrostatic interaction between amino-modified silica and hydrothermal carbon was regarded as the main driving force for the formation of homogeneous carbon coverage on the silica surface. The obtained composites showed high graphitization degree, and controlled morphology (shape and particle size) and pore size by adjusting the species of carriers and hydrothermal conditions. The application results demonstrated that a thin carbon layer possessed high adsorption capacities for dyes, and the composite could be rapidly recovered by sedimentation (10 min) after adsorption with 30 µm spherical silica gel as the carrier. Besides, baseline chromatographic separation of oligosaccharide isomers could be achieved on the silica-carbon column. These results indicated that the silica-carbon composites should be promising functional materials for the large-molecule-involving processes such as adsorption and chromatographic separation.

[Separation and purification of the components in Trachelospermum jasminoides by two dimensional hydrophilic interaction liquid chromatography- reversed-phase liquid chromatography].

A preparative two dimensional hydrophilic interaction liquid chromatography/reversed-phase liquid chromatography (Pre-2D-HILIC/RPLC) method was established to separate and purify the components in Trachelospermum jasminoides. The pigments and strongly polar components were removed from the crude extract after the active carbon decolorization and solid phase extraction processes. A Click XIon column (250 mm×20 mm, 10 µm) was selected as stationary phase and water-acetonitrile as mobile phases in the first dimensional HILIC. Finally, 15 fractions were collected under UV-triggered mode. In the second dimensional RPLC, a C18 column (250 mm×20 mm, 5 µm) was selected and water-acetonitrile was used as mobile phases. As a result, 14 compounds with high purity were obtained, which were further identified by mass spectrometry (MS) and nuclear magnetic resonance (NMR). Finally, 11 lignan compounds and three flavonoid compounds were obtained. The method has a good orthogonality, and can improve the resolution and the peak capacity. It is significant for the separation of complex components from Trachelospermum jasminoides.

Preparative separation of the polar part from the rhizomes of Anemarrhena asphodeloides using a hydrophilic C18 stationary phase.

The goal of this study was to develop a method that utilized a hydrophilic C18 stationary phase in the preparative high performance liquid chromatography to isolate the polar part from the rhizomes of Anemarrhena asphodeloides. The results showed that an initial mobile phase of pure water for the separation could greatly increase the retention and solubility of the polar compounds at the preparative scale. Introducing polar groups on the surface of the hydrophilic C18 column together with the use of optimized mobile phase compositions improved the column separation selectivity for polar compounds. Eleven previously undescribed compounds in Anemarrhena asphodeloides were obtained, indicating that the method developed in this study would facilitate the purification and separation of the polar part of traditional Chinese medicines.

Efficient purification of high-purity compounds from the stem of Lonicera japonica Thunb using two-dimensional preparative chromatography.

Purification of high-purity compounds from traditional Chinese medicines (TCMs) plays an important role in investigating their bioactivity. Nevertheless, it is often quite difficult to isolate compounds with high purity because of the complexity of TCMs in chemical composition. In this work, a two-dimensional preparation method was successfully developed for the preparation of high-purity compounds from the stem of Lonicera japonica Thunb, based on two novel polar copolymerized RP stationary phases, XAqua C3 and XAqua C18. An XAqua C3 prep column was used to separate the sample in the first-dimensional preparation, and 14 g of sample was fractionated into eight fractions with a recovery of 82%. An XAqua C18 prep column was selected to prepare high-purity compounds in the second-dimensional preparation for its good orthogonality with the XAqua C3 stationary phase. As a result, major compounds in the sample were isolated with more than 99% purity. This method is a potent method to realize the efficient purification of compounds with high purity from the stem of L. japonica Thunb and it shows great potential in the separation of high-purity compounds from complex samples.

Purification of saponins from leaves of Panax notoginseng using preparative two-dimensional reversed-phase liquid chromatography/hydrophilic interaction chromatography.

Saponins are widely distributed in the plant kingdom and have been shown to be active components of many medicinal herbs. In this study, a two-dimensional purification method based on reversed-phase liquid chromatography coupled with hydrophilic interaction liquid chromatography was successfully applied to purify saponins from leaves of Panax notoginseng. Nine saponin reference standards were used to test the separation modes and columns. The standards could not be resolved using C18 columns owing to their limited polar selectivity. However, they were completely separated on a XAmide column in hydrophilic interaction liquid chromatography mode, including two pairs of standards that were coeluted on a C18 column. The elution order of the standards on the two columns was sufficiently different, with a correlation coefficient between retention times on the C18 and XAmide columns of 0.0126, indicating good column orthogonality. Therefore, the first-dimension preparation was performed on a C18 column, followed by a XAmide column that was used to separate the fractions in the second dimension. Fifty-four fractions were prepared in the first dimension, with 25 fractions rich in saponins. Eight saponins, including two pairs of isomeric saponins and one new saponin, were isolated and identified from three representative fractions. This procedure was shown to be an effective approach for the preparative isolation and purification of saponins from leaves of P. notoginseng. Moreover, this method could possibly be employed in the purification of low-content and novel active saponins from natural products.

[Effects of astragali radix extract on matrix metalloproteinase 9 expression and atherosclerotic plaque formation in aorta of apolipoprotein E deficient mice fed with high fat diet].

OBJECTIVE: To explore the effects of astragali radix extract on the expressions of matrix metalloproteinase 9 (MMP-9) and the formation of atherosclerotic plaque in aortic atherosclerotic plaques of apolipoprotein E-deficient mice (ApoE-/-). METHODS: Male 8-week-old ApoE-/- mice fed with high fat diet were randomly divided into four groups (n=12 each): control group (saline 0.2 ml/d), atorvastatin group (atorvastatin 10 mg×kg(-1)×d(-1)), low-dose astragali radix extract group (1.25 g×kg(-1)×d(-1)) and high-dose astragali radix extract group (5 g×kg(-1)×d(-1)). After 12 weeks, serum oxLDL was measured by the method of ELISA. The formation of atherosclerotic plaque was determined in HE and oil red O stained aortic slice. The expressions of macrophage and MMP-9 in the aortic plaque were detected by immune fluorescence and immunohistochemistry staining method. RESULTS: Similarly as atorvastatin, astragali radix extract significantly decreased the level of serum oxLDL in ApoE-/-1 mice in a dose-dependent manner. The level of oxLDL in the high-dose astragali radix extract group [(5.2±6.1) µg/ml] was significantly lower than that in the control group [(15.8±5.4) µg/ml, P<0.01]. The area of atherosclerosis plaques was smaller (17.24%±4.22% vs. 49.87%±9.37%, P<0.01) and the penetration degree of plaques in the arterial wall was relieved in the high-dose astragali radix extract group compared to those in the control group (P<0.01). The expressions of Mac3 in atherosclerosis plaques of the high-dose astragali radix extract group was also significantly lower than in the control group (P<0.01). The mean absorbance value of the expression of MMP-9 in the high-dose astragali radix extract group (0.0154±0.0014)was significantly lower than that in the control group (0.0263±0.0065) (P<0.01). CONCLUSIONS: Similar as atorvastatin, astragali radix extract can dose-dependently inhibit the expression of MMP-9 and the formation of the atherosclerotic plaque in ApoE-/- mouse, probably by reducing the serum oxLDL, inhibiting macrophage infiltration, migration and secretion of MMP-9.

Anti-atherosclerotic function of Astragali Radix extract: downregulation of adhesion molecules in vitro and in vivo.

BACKGROUND: Atherosclerosis is considered to be a chronic inflammatory disease. Astragali Radix extract (ARE) is one of the major active ingredients extracted from the root of Astragalus membranaceus Bge. Although ARE has an anti-inflammatory function, its anti-atherosclerotic effects and mechanisms have not yet been elucidated. METHODS: Murine endothelial SVEC4-10 cells were pretreated with different doses of ARE at different times prior to induction with tumor necrosis factor (TNF)-α. Cell adhesion assays were performed using THP-1 cells and assessed by enzyme-linked immunosorbent assay, western blotting and immunofluorescence analyses to detect the expression of vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), phosphorylated inhibitor of κB (p-iκB) and nuclear factor (NF)-κB. We also examined the effect of ARE on atherosclerosis in the aortic endothelium of apolipoprotein E-deficient (apoE(-/-)) mice. RESULTS: TNF-α strongly increased the expression of VCAM-1 and ICAM-1 accompanied by increased expression of p-iκB and NF-κB proteins. However, the expression levels of VCAM-1 and ICAM-1 were reduced by ARE in dose- and time-dependent manners, with the strongest effect at a dose of 120 µg/ml incubated for 4 h. This was accompanied by significantly decreased expression of p-iκB and inhibited activation of NF-κB. Immunofluorescence analysis also revealed that oral administration of ARE resulted in downregulation of adhesion molecules and decreased expression of macrophages in the aortic endothelium of apoE(-/-) mice. ARE could suppress the inflammatory reaction and inhibit the progression of atherosclerotic lesions in apoE(-/-) mice. CONCLUSION: This study demonstrated that ARE might be an effective anti-inflammatory agent for the treatment of atherosclerosis, possibly acting via the decreased expression of adhesion molecules.

The herbalome--an attempt to globalize Chinese herbal medicine.

The herbalome is a project with the objective of globalizing Chinese herbal medicine (CHM) by clarification of its composition, structure, and function; by establishing a standard resource library; and by interpreting the synergistic and complementary mechanisms of multi-components on multi-targets. In phase I, it focuses on the development of systematic separation methodology for resolving and analyzing the complex components in CHM and establishment of a comprehensive resource library. This review summarizes recent advances in the herbalome project with regard to innovative separation techniques and demonstration of a resource library.

Separation and identification of flavonoids from complex samples using off-line two-dimensional liquid chromatography tandem mass spectrometry.

In this study, a LC-based identification strategy was proposed and off-line two-dimensional liquid chromatography was developed for the separation and identification of flavonoids from complex samples. The highly orthogonal separation system was composed of a CD column, an OEG column and an XTerra C(18) column. The CD column was employed for the first dimensional separation and the OEG column and XTerra C(18) column were used for the second dimensional separation in parallel. A mixture of six traditional Chinese medicines was used as a complex sample in testing this method. Unknown peaks were identified by matching their retention times and accurate mass molecular weights with those of references. The identification efficacy of parallel unidimensional liquid chromatography and off-line two-dimensional liquid chromatography was compared, and the latter was demonstrated to be more efficient for the identification of target compounds-especially co-eluted compounds or minor compounds from complex samples.

Selective separation of flavonoid glycosides in Dalbergia odorifera by matrix solid-phase dispersion using titania.

Dalbergia odorifera contains high concentrations of flavonoid aglycones and trace flavonoid glycosides. In this study, trace flavonoid glycosides were separated from D. odorifera by titania with matrix solid-phase dispersion (MSPD). Before the MSPD experiment, four standards, including two isoflavone glycosides (genistin and formononetin-8-C-apiosyl (1-6)-glucoside) and their aglycones (genistein and formononetin), were used to compare their retention on a titania column. The effect of acetonitrile concentration and pH on their retention was investigated and a conclusion was drawn that high acetonitrile concentration and pH lead to the greatest difference in the retention of flavonoid as glycosides and aglycones. Besides hydrophilic interaction and ligand-exchange interaction may exist between sugar moiety of flavonoid glycoside and titania, so that flavonoid glycosides have stronger retention than that of aglycones. Based on the chromatographic rule of flavonoid as glycosides and aglycones on the titania column, the MSPD method was optimized to elute high concentration flavonoid aglycones first with 90% acetonitrile and 10% water containing 100 mM ammonium acetate buffer, and then to elute trace flavonoid glycosides with 20% acetonitrile and 80% water containing 1% trifluoroacetate (TFA). Isolated flavonoid glycosides were further analyzed by UPLC-MS/MS, and their fragmentation in MS(2) showed they are C-glycosyl flavonoids.

[Effects of Salvia miltiorrhiza and Carthamus tinctorius aqueous extracts and compatibility on rat myocardial ischemic reperfusion injury].

OBJECTIVE: To separate and characterize aqueous extracts of Salvia miltiorrhiza and Carthamus tinctorius to efficient, high-throughput and strong polar components, to observe effects of their aqueous effective components compatibility on rat myocardial ischemic reperfusion injury. METHOD: Myocardial ischemic reperfusion injury model were established on SD rats by 40 min ligation of the left anterior descending artery and 120 min reperfusion. The rats were injected experimental drugs intravenously from femoral vein after 10 min ischemia. Rats were randomly divided into sham group (the suture around the left anterior descending coronary artery was not tied), model group, Danhong injection group (content of protocatechualdehyde is 0.05 g x L(-1), injection dosage equivalent to 1.80 g x kg(-1)), aqueous effective component of S. miltiorrhiza group (content of salvianolic acid B is 49 g x L(-1), injection dosage equivalent to 30.68 g x kg(-1)), aqueous effective component of S. miltiorrhiza group (content of hydroxysafflor yellow A is 31.76 g x L(-1), injection dosage equivalent to 17.87 g x kg(-1)), aqueous effective components compatibility of S. miltiorrhizae and C. tinctorius group (injection dosage is respectively 24.28 g x kg(-1) and 48.55 g x kg(-1)), each group have ten rats. Drugs were diluted with an equal dose of normal saline. The rats of sham group and model group were injected equivalent dosage of saline. The myocardial infarction size and the contents of serum cTnT and CK-MB were detected. The level of TXB2, 6-keto-PGF(1alpha) and platelet aggregation in blood plasma were investigated. RESULT: Compared with sham group, serum cTnT and CK-MB contents in model group increased significantly (P < 0.01). Compared with model group, myocardial infarction size and serum cTnT and CK-MB contents in aqueous effective component of S. miltiorrhiza group, aqueous effective component of C. tinctorius group and aqueous effective components compatibility of S. miltiorrhiza and C. tinctorius groups decreased significantly. Aqueous effective component of S. miltiorrhiza increased the level of 6-ke-to-PGF(1alpha), as well as decreased content of TXB2 and inhibited platelet aggregation (P < 0.01). Aqueous effective component of C. tinctorius also decreased the content TXB2 (P < 0.01). Improved extent of some detected markers in aqueous effective components compatibility of S. miltiorrhiza and C. tinctorius groups were better than that of Danhong injection group. CONCLUSION: Effective components compatibility of aqueous extracts from S. miltiorrhiza and C. tinctorius may reduce myocardial infarct size and leakage of myocardial enzyme, and increase the level of 6-keto-PGF1alpha, so as to inhibit platelet aggregation and prevent thrombosis, the result of which is to reduce myocardial ischemic reperfusion injury.

A new method for chemical identification based on orthogonal parallel liquid chromatography separation and accurate molecular weight confirmation.

Recent advances in the theory and application of orthogonal LC separation have allowed for the establishment of a more effective method for the chemical identification of target compounds in complex samples, especially structurally similar compounds. In this study, a new chemical identification method based on orthogonal parallel separation and accurate molecular weight confirmation was developed. An orthogonal separation system consisting of an XTerra MS C(18) column, a home-made Click OEG column, and a Click CD column was established for separation and identification. In addition, 82 flavonoids were selected as references, to be used for the construction of a library. Retention times of each reference flavonoid on each column and accurate molecular weights were recorded and imported into a searchable library as "tags" for the unknown screening. For the method validation, two complex mixtures, fractions of Dalbergia odorifera T. Chen and Scutellaria baicalensis Georgi, specifically, were separated and identified. In total, nine compounds were unequivocally identified by retention time and confirmation of accurate molecular weight, demonstrating that this method is suitable and efficient for the chemical identification of complex samples.

Purification of compounds from Lignum Dalbergia Odorifera using two-dimensional preparative chromatography with Click oligo (ethylene glycol) and C18 column.

Purification of compounds from traditional Chinese medicines (TCMs) is an important task for understanding the chemical composition of TCMs. However, it is difficult to obtain compounds with high enough purity for identification by NMR due to the complexity of TCMs in chemical composition. In this study, a two-dimensional purification method based on a Click oligo (ethylene glycol) column and a C18 column was developed to realize an orthogonal separation in preparative level for purifying compounds efficiently. The first dimensional preparation was performed on a Click oligo (ethylene glycol) column to simplify the sample into the fractions with good separation repeatability. On the first dimension, 7.2 g sample was separated into 11 fractions with a recovery of 86% within 6 h. A C18 column was taken as the second dimension to realize the high-performance separation and rapid preparation from the fractions collected from the first dimension. Eight compounds in fraction 6 and 2 compounds in fraction 8 were isolated and identified after optimizing the separation and collection parameters. This method is a high-efficient and orthogonal preparation method to improve the separation of a complex sample and increase the purity of the compounds, which benefits from the application of novel materials in the preparation and purification.