Simultaneous quantitative analysis of nine constituents in six Chinese medicinal materials from Citrus genus by high‐performance liquid chromatography and high‐resolution mass spectrometry combined with chemometric methods

In this study, we aim to determine the chemical constituents of six Chinese medicinal materials from the Citrus genus using high‐performance liquid chromatography and high‐resolution mass spectrometry. Eight flavonoids and one coumarin were identified and further quantified as marker substances by high‐performance liquid chromatography method. The separation was performed on an Agilent TC‐C₁₈ column with 0.1% formic acid and acetonitrile as the mobile phase under gradient elution. The analytical method was fully validated in terms of linearity, sensitivity, intra‐ and inter‐day precision and repeatability, limit of detection, limit of quantitation, and recovery. It was subsequently applied to evaluate the quality of 103 batches of the Chinese medicinal materials from the Citrus genus. In addition, the principal constituent analysis was used to compare the samples of different species from the Citrus genus leading to successful classification of the samples in accordance with their origins. It was found that the contents of nine constituents varied greatly in different ripening stages and varieties of the samples from the Citrus genus. In addition, neoeriocitrin and 5,7‐dimethoxycoumarin were determined as two unique constituents of ‘Zhiqiao’ and ‘Foshou’, respectively. In conclusion, this study provides a chemical basis for quality control of Chinese medicinal materials from the Citrus genus.     

Synthesis of Natural Atisane‐Type Diterpenoids by retro‐Biomimetic Transformations

An efficient one step, retro‐biomimetic procedure for the synthesis of natural products having the atisane structure is described (Scheme 2), natural products which are components of medicinal plants and possess relevant biological activity. Their structures were confirmed by chemical transformations and spectral data. The starting materials were the known ent‐kaur‐16‐en‐19‐oic acid ( 1 ) and ent‐trachyloban‐19‐oic acid ( 2 ), diterpenoids readily available from the waste of sunflower.     

Simultaneous determination of three purines in Alysicarpus vaginalis (L.) DC. by hollow fiber‐based liquid‐phase microextraction combined with high‐performance liquid chromatography

In this paper, a three‐phase hollow fiber liquid‐phase microextraction (HF‐LPME) method combined with high‐performance liquid chromatography (HPLC) was developed for the determination of hypoxanthine (HX), xanthine (Xan) and adenine (A) and then for the first time successfully applied to the analysis of HX, Xan and A in Alysicarpus vaginalis (L.) DC. medicinal materials. Different factors affecting the HF‐LPME procedure were investigated and optimized. Under optimal extraction conditions (1‐octanol as organic solvent, pH of the donor and acceptor phase 10.0 and 3.5, respectively, extraction time 40 min, stirring rate 800 rpm and salt addition 10%, w/v), HX, Xan and A could be determined within the test ranges with a good correlation coefficient (r² > 0.9992). The limit of detection for HX, Xan and A was 153, 173 and 97 ng/mL, respectively, and the intra‐ and inter‐day relative standard deviations were no more than 9.8%. The content of HX, Xan and A in Alysicarpus vaginalis (L.) DC. medicinal materials was 120.40, 18.37 and 62.75 µg/g, respectively. This procedure afforded a convenient, sensitive, accurate and inexpensive method with a high extraction efficiency for determination of HX, Xan and A. Copyright © 2013 John Wiley & Sons, Ltd.     

Qualitative and quantitative analysis of the major constituents in Jin‐Mu‐Gan‐Mao tablet by high‐performance liquid chromatography with diode‐array detection and quadrupole time‐of‐flight tandem mass spectrometry

Jin‐Mu‐Gan‐Mao tablet is a well‐known traditional Chinese medicinal preparation, which has been used to treat the common cold in China. In this study, a systematic method was established for the qualitative and quantitative analysis of the major constituents in Jin‐Mu‐Gan‐Mao tablet. First, a method of high‐performance liquid chromatography with diode‐array detection and quadrupole time‐of‐flight mass spectrometry was developed for identification of the multi‐constituents. Thirty‐one compounds including ten phenolic acids, 18 flavonoids, and three iridoid glycosides were clearly identified by comparison with the reference standards, and 11 compounds were deduced by comparison with the literature data. Second, a new quantitative analysis method of Jin‐Mu‐Gan‐Mao tablet was established by high‐performance liquid chromatography with diode‐array detection. Twelve compounds, either with high contents or strong bioactivities, were chosen as marker components. This analytical method was validated through intra‐ and interday precision, repeatability, and stability, with respective relative standard deviations less than 1.74, 2.54, 2.44, and 2.48%. The limits of detection and quantification were less than 0.327 and 0.881 μg/mL, respectively. The overall recoveries ranged from 97.04–102.76% (relative standard deviation ≤ 2.91%). Then this validated method was applied to determine ten batches of Jin‐Mu‐Gan‐Mao tablet. The results indicated that these new approaches can be applicable for the qualitative and quantitative analysis of Jin‐Mu‐Gan‐Mao tablet.     

Piperazine‐functionalized nickel ferrite nanoparticles as efficient and reusable catalysts for the solvent‐free synthesis of 2‐amino‐4H‐chromenes

Piperazine‐functionalized nickel ferrite (NiFe₂O₄) nanoparticles were synthesized as recoverable heterogeneous base catalysts using a routine method. The synthesized materials were characterized using various spectroscopic techniques such as infrared, X‐ray diffraction, scanning electron microscopy, energy‐dispersive X‐ray, thermogravimetry analysis, and vibrating sample magnetometry. Catalytic efficiency was investigated in the synthesis of 2‐amino‐4H‐chromene derivatives via a one‐pot three component reaction of aldehyde and malononitrile with β or α‐naphthol/5‐methyle resorcinol under solvent‐free conditions with good to high yields. This method is operationally simple and has several advantages such as good to high yield, short reaction times, solvent‐free conditions, and easy synthesis. Moreover, the catalyst was recovered easily using an external magnet and reused three times without distinctive loss in catalytic activity.     

N‐Oxide 1,2,4,5‐Tetrazine‐Based High‐Performance Energetic Materials

One route to high density and high performance energetic materials based on 1,2,4,5‐tetrazine is the introduction of 2,4‐di‐N‐oxide functionalities. Based on several examples and through theoretical analysis, the strategy of regioselective introduction of these moieties into 1,2,4,5‐tetrazines has been developed. Using this methodology, various new tetrazine structures containing the N‐oxide functionality were synthesized and fully characterized using IR, NMR, and mass spectroscopy, elemental analysis, and single‐crystal X‐ray analysis. Hydrogen peroxide (50 %) was used very effectively in lieu of the usual 90 % peroxide in this system to generate N‐oxide tetrazine compounds successfully. Comparison of the experimental densities of N‐oxide 1,2,4,5‐tetrazine compounds with their 1,2,4,5‐tetrazine precursors shows that introducing the N‐oxide functionality is a highly effective and feasible method to enhance the density of these materials. The heats of formation for all compounds were calculated with Gaussian 03 (revision D.01) and these values were combined with measured densities to calculate detonation pressures (P) and velocities (ν_D) of these energetic materials (Explo 5.0 v. 6.01). The new oxygen‐containing tetrazines exhibit high density, good thermal stability, acceptable oxygen balance, positive heat of formation, and excellent detonation properties, which, in some cases, are superior to those of 1,3,5‐tritnitrotoluene (TNT), 1,3,5‐trinitrotriazacyclohexane (RDX), and octahydro‐1,3,5,7‐tetranitro‐1,3,5,7‐tetrazocine (HMX).     

Tandem One‐pot Synthesis of Polysubstituted 1,3‐Thiazine Derivatives

A tandem one‐pot synthesis of polysubstituted 1,3‐thiazines has been developed by reacting with cyanoacetamide and isothiocyanate derivatives to give rise to 2‐cyano‐3‐mercaptoacrylamides, which are trapped in situ by various aldehydes or diversely substituted ketones through intermolecular cyclization, providing polysubstituted 1,3‐thiazine derivatives in short reaction times with good to excellent yields. The salient features of this novel protocol are operational simplicity, accessing the desired products from the readily available starting materials and easy of product isolation and may find wide spread applications in medicinal chemistry.     

Identification of chemical constituents in traditional Chinese medicine formula using HPLC coupled with linear ion trap‐Orbitrap MS from high doses of medicinal materials to equivalent doses of formula: Study on Xiang‐Sha‐Liu‐Jun‐Zi‐Jia‐Jian granules

High‐resolution mass spectrometry has been a powerful tool for the research of chemical constituents in traditional Chinese medicine (TCM) formulas. However, the chromatographic peaks were difficult to discriminate clearly in data collection or analysis because of the complexity and the greatly different content of the constituents in TCM formula, which increased the difficulty of identification. In this study, a high‐performance liquid chromatography coupled with linear ion trap‐Orbitrap mass spectrometry based strategy focused on the comprehensive identification of TCM formula constituents was developed. Identification was carried out from a high dose of medicinal materials to equivalent dose of formula. Meanwhile, combined with mass spectrometry data, chromatographic behaviors, reference standards and previous reports, the identification of constituents in Xiang‐Sha‐Liu‐Jun‐Zi‐Jia‐Jian granules was described. 169 compounds were unambiguously or tentatively characterized, mainly including flavonoids, alkaloids, triterpenic acids, triterpene saponins, lactones, sesquiterpenoids and some other compounds. Among them, 11 compounds were unambiguously confirmed by comparing with reference standards. These results demonstrated that the method was effective and reliable for comprehensive identification of constituents of Xiang‐Sha‐Liu‐Jun‐Zi‐Jia‐Jian granules extracts and reveal the material basis of its therapeutic effects. This strategy might propose a research idea for the characterization of multi‐constituents in TCM formula.     

Analysis of variation in the ovine ultra‐high sulphur keratin‐associated protein KAP5‐4 gene using PCR‐SSCP technique

Keratin‐associated proteins (KAPs) are one of the main structural components of the wool fibre. Variation in the KAP genes (KRTAPs) may affect the structure of KAPs and hence wool characteristics. In this study, we used PCR‐SSCP to analyse ovine KRTAP5‐4, a gene encoding a member of the KAP5 family. Five different PCR‐SSCP patterns were detected in the 250 sheep that were analysed. Either one or a combination of two patterns was observed for each sheep, which was consistent with these sheep being either homozygous or heterozygous at this locus. DNA sequencing revealed that these patterns represent five different DNA sequences. One of the sequences was identical to a published ovine KRTAP5‐4 sequence. The remaining four were unique, but shared a high homology with the published ovine KRTAP5‐4 sequence, suggesting that these sequences represent allelic variants of KRTAP5‐4. There were a total of six SNPs and one length polymorphism in the sequences. Of the five SNPs found in the coding region, four were non‐synonymous SNPs and would result in amino acid changes. The length polymorphism would affect the cysteine content of the putative peptide and this along with the SNPs may have an impact on the structure of KAP5‐4, and hence affect wool traits.     

Characterization of tropane and cinnamamide alkaloids from Scopolia tangutica by high‐performance liquid chromatography with quadrupole time‐of‐flight tandem mass spectrometry

Scopolia tangutica is a traditional Chinese medicine used for antispasmodic, anesthesia, analgesia, and sedation. Its medicinal activity is associated to alkaloid constituents, including tropane and cinnamamide types. Low content of alkaloids in plant makes them difficult to be isolated and identified. The present work developed an effective method to quickly characterize alkaloids from Scopolia tangutica by high‐performance liquid chromatography coupled with quadrupole time‐of‐flight tandem mass spectrometry. Thirteen reference compounds were studied for their fragmentation pathways, including five tropane alkaloids and eight cinnamamide ones. Alkaloid constituent was analyzed by an optimized high‐performance liquid chromatography method and mass spectrometry analysis to achieve systematic characterization of alkaloids from Scopolia tangutica. As a result, 53 compounds were identified, including 21 tropane alkaloids (eight new ones), 18 caffeoyl ones (ten new ones) and 14 dicaffeoyl ones (seven new ones). It was important to provide rich information in phytochemical study and structure‐guided isolation of important compounds from this plant.     

Validation of green‐solvent extraction combined with chromatographic chemical fingerprint to evaluate quality of Stevia rebaudiana Bertoni

An approach that combined green‐solvent methods of extraction with chromatographic chemical fingerprint and pattern recognition tools such as principal component analysis (PCA) was used to evaluate the quality of medicinal plants. Pressurized hot water extraction (PHWE) and microwave‐assisted extraction (MAE) were used and their extraction efficiencies to extract two bioactive compounds, namely stevioside (SV) and rebaudioside A (RA), from Stevia rebaudiana Bertoni (SB) under different cultivation conditions were compared. The proposed methods showed that SV and RA could be extracted from SB using pure water under optimized conditions. The extraction efficiency of the methods was observed to be higher or comparable to heating under reflux with water. The method precision (RSD, n = 6) was found to vary from 1.91 to 2.86% for the two different methods on different days. Compared to PHWE, MAE has higher extraction efficiency with shorter extraction time. MAE was also found to extract more chemical constituents and provide distinctive chemical fingerprints for quality control purposes. Thus, a combination of MAE with chromatographic chemical fingerprints and PCA provided a simple and rapid approach for the comparison and classification of medicinal plants from different growth conditions. Hence, the current work highlighted the importance of extraction method in chemical fingerprinting for the classification of medicinal plants from different cultivation conditions with the aid of pattern recognition tools used.     

Electrostatic field–induced tip‐electrospray ionization mass spectrometry for direct analysis of raw food materials

Rapid characterization of metabolites and risk compounds such as chemical residues and natural toxins in raw food materials such as vegetables, meats, and edible living plants and animals plays an important part in ensuing food quality and safety. To rapidly characterize the analytes in raw food materials, it is essential to develop in situ method for directly analyzing raw food materials. In this work, raw food materials including biological tissues and living samples were placed between an electrode and mass spectrometric (MS) inlet under a strong electrostatic field; analytes were rapidly induced to generate electrospray ionization (ESI) from the sample tip by adding a drop of solvent onto the sample. Therefore, the electrostatic field–induced tip‐ESI‐MS allows raw samples to avoid contacting high voltage, and thus this method has the advantage for in vivo analysis of food living plants and animals. Metabolite profiling, residues of pesticides and veterinary drugs, and natural toxins from raw food materials have been successfully detected. The analytical performances, including the linear ranges, sensitivity, and reproducibility, were investigated for direct sample analysis. The ionization mechanism of electrostatic field–induced tip‐ESI was also discussed in this work.     

TMSCl Promoted Direct sp3 C‐H Alkenylation to Construct (E)‐2‐Styryl‐tetrahydrobenzo[d]thiazoles

A high‐efficient and stereo‐specific approach for the preparation of biologically important (E)‐2‐styryl‐tetrahydrobenzo[d]thiazoles has been developed via TMSCl promoted direct sp³ C‐H alkenylation of 2‐methyl‐5,6‐dihydrobenzo[d]thiazol‐7(4H)‐one under metal‐free conditions. Seventeen target compounds were synthesized in excellent yields of 82% –98% under the optimal conditions of 300 mol% TMSCl at 110°C for 2 h, and their chemical structures were elucidated by IR, NMR, ESI‐MS, elemental analyses and X‐ray crystallography analysis. A plausible mechanism was also proposed, and this method provided a good functional group conversion for the sp³ C‐H substrates.     

Study on the spectrum‐effect relationship of the xanthine oxidase inhibitory activity of Ligustrum lucidum

To evaluate the xanthine oxidase inhibitory activity of the chemical constituents of Ligustrum lucidum in vitro, the spectrum‐effect relationship was investigated. The high‐performance liquid chromatography fingerprint was established by ultraviolet spectrophotometry, and the xanthine oxidase inhibitory activity was tested in vitro by a high‐throughput screening method. Cluster analysis, principal component analysis, gray correlation analysis, and partial least squares regression were used to explore the spectrum‐effect relationships. Sixty batches of Ligustrum lucidum were collected from 16 provinces for testing. The results revealed differences among the batches of medicinal materials, and the similarity score was between 0.635 and 0.968. Thirty‐three characteristic peaks (1–33) were calibrated by fingerprint evaluation software for traditional Chinese medicine. The spectrum‐effect relationship study further revealed that the contents of peaks 1, 2, 4, 5, 6, 7, 14, 17, 25, 28, 31, and 33, which are potentially critical ingredients for quality control of Ligustrum lucidum fruit, were highly correlated with the inhibition of xanthine oxidase activity.     

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.     

Rapid screening and identification of lycodine‐type alkaloids in Lycopodiaceae and Huperziaceae plants by ultra‐performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry

Lycodine‐type alkaloids have gained significant interest owing to their unique skeletal characteristics and acetylcholinesterase activity. This study established a rapid and reliable method using ultra‐performance liquid chromatography coupled with electrospray ionization quadrupole time‐of‐flight tandem mass spectrometry (UPLC‐ESI‐Q/TOF‐MS/MS) for comprehensive characterization of lycodine‐type alkaloids for the first time. The lycodine‐type alkaloids were detected successfully from Lycopodiastrum casuarinoides, Huperzia serrata and Phlegmarirus carinatus in seven plants of the Lycopodiaceae and Huperziaceae families, based on the established characteristic MS fragmentation of five known alkaloids. Furthermore, a total of 13 lycodine‐type alkaloids were identified, of which three pairs of isomers were structurally characterized and differentiated. This study further improves mass analysis of lycodine‐type alkaloids and demonstrates the superiority of UPLC with a high‐resolution mass spectrometer for the rapid and sensitive structural elucidation of other trace active compounds. Copyright © 2016 John Wiley & Sons, Ltd.     

Relationship between the UPLC‐Q‐TOF‐MS fingerprinted constituents from Daphne genkwa and their anti‐inflammatory,anti‐oxidant activities

Daphne genkwa Sieb.et Zucc. is a well‐known medicinal plant. This study was designed to apply the ultra‐high performance liquid chromatography system to establish a quality control method for D. genkwa. Data revealed that there were 15 common peaks in 10 batches of D. genkwa Sieb. Et Zucc. (Thymelaeaceae) from different provinces of China. On this basis, the fingerprint chromatogram was established to provide references for quality control. Afterwards, the chemical constitutions of these common peaks were analyzed using the UPLC‐Q‐TOF‐MS system and nine of them were identified. In addition, LPS‐stimulated RAW264.7 murine macrophages and DPPH assay were used to study the anti‐inflammatory and anti‐oxidation effects of D. genkwa . Then the fingerprint–efficacy relationships between UPLC fingerprints and pharmacodynamic data were studied with canonical correlation analysis. Analysis results indicated that the anti‐inflammatory and anti‐oxidation effects differed among the 10 D. genkwa samples owing to their inherent differences of chemical compositions. Taken together, this research established a fingerprint–efficacy relationship model of D. genkwa plant by combining the UPLC analytic technique and pharmacological research, which provided references for the detection of the principal components of traditional Chinese medicine on bioactivity.     

Method development for the analysis of resinous materials with MALDI‐FT‐ICR‐MS: novel internal standards and a new matrix material for negative ion mode

Matrix‐assisted laser desorption/ionization (MALDI) is a mass spectrometry (MS) ionization technique suitable for a wide variety of sample types including highly complex ones such as natural resinous materials. Coupled with Fourier transform ion cyclotron resonance (FT‐ICR) mass analyser, which provides mass spectra with high resolution and accuracy, the method gives a wealth of information about the composition of the sample. One of the key aspects in MALDI‐MS is the right choice of matrix compound. We have previously demonstrated that 2,5‐dihydroxybenzoic acid is suitable for the positive ion mode analysis of resinous samples. However, 2,5‐dihydroxybenzoic acid was found to be unsuitable for the analysis of these samples in the negative ion mode. The second problem addressed was the limited choice of calibration standards offering a flexible selection of m/z values under m/z 1000. This study presents a modified MALDI‐FT‐ICR‐MS method for the analysis of resinous materials, which incorporates a novel matrix compound, 2‐aminoacridine for the negative ion mode analysis and extends the selection of internal standards with m/z <1000 for both positive (15 different phosphazenium cations) and negative (anions of four fluorine‐rich sulpho‐compounds) ion mode. The novel internal calibration compounds and matrix material were tested for the analysis of various natural resins and real‐life varnish samples taken from cultural heritage objects. Copyright © 2017 John Wiley & Sons, Ltd.     

The e‐tongue‐based classification and authentication of mineral water samples using cross‐correlation‐based PCA and Sammon's nonlinear mapping

The present paper elaborates on the design of classifiers based on cross‐correlation‐based principal component analysis (PCA) and Sammon's nonlinear mapping (NLM) using current signals obtained from electronic tongue (e‐tongue) with commercial mineral water samples available in the Indian market. The pulse‐voltammetric method is used to capture the electroanalytical/electrochemical characteristics of the sampled mineral waters by considering a real model for the liquid–electrode interface in a given e‐tongue apparatus. Then the cross‐correlation coefficients between the output and input signals are determined. Both PCA and Sammon's NLM create a subspace from high‐dimensional mineral water data by considering the principal eigenvectors and minimising the stress function, respectively. The proposed cross‐correlation‐based PCA and Sammon's classifiers establish the highest separation distance among the investigated water brands and carries out the authentication of more than one unknown sample of the same brand with a certain degree of variability with respect to their sources. Copyright © 2013 John Wiley & Sons, Ltd.