A method for screening active components from Chinese herbs by cell membrane chromatography-offline-high performance liquid chromatography/mass spectrometry and an online statistical tool for data processing.

Cell membrane chromatography (CMC) has been successfully applied to screen bioactive compounds from Chinese herbs for many years, and some offline and online two-dimensional (2D) CMC-high performance liquid chromatography (HPLC) hyphenated systems have been established to perform screening assays. However, the requirement of sample preparation steps for the second-dimensional analysis in offline systems and the need for an interface device and technical expertise in the online system limit their extensive use. In the present study, an offline 2D CMC-HPLC analysis combined with the XCMS (various forms of chromatography coupled to mass spectrometry) Online statistical tool for data processing was established. First, our previously reported online 2D screening system was used to analyze three Chinese herbs that were reported to have potential anti-inflammatory effects, and two binding components were identified. By contrast, the proposed offline 2D screening method with XCMS Online analysis was applied, and three more ingredients were discovered in addition to the two compounds revealed by the online system. Then, cross-validation of the three compounds was performed, and they were confirmed to be included in the online data as well, but were not identified there because of their low concentrations and lack of credible statistical approaches. Last, pharmacological experiments showed that these five ingredients could inhibit IL-6 release and IL-6 gene expression on LPS-induced RAW cells in a dose-dependent manner. Compared with previous 2D CMC screening systems, this newly developed offline 2D method needs no sample preparation steps for the second-dimensional analysis, and it is sensitive, efficient, and convenient. It will be applicable in identifying active components from Chinese herbs and practical in discovery of lead compounds derived from herbs.

Development of a QuEChERS method for simultaneous analysis of antibiotics in carcasses for supplementary feeding of endangered vultures.

Antibiotics have been beneficial for human and animal health. However, an excessive use in livestock and a deficient management of the carcasses can lead to adverse effects in the scavengers that ingest them, especially in "supplementary feeding sites" (SFS). The aim of this study was to assess the potential risk of exposure to antibiotics for an endangered population of Cinereous vultures (Aegypius monachus) from southeastern Portugal. Hence, a multi-residue method based on QuEChERs was adapted and validated to analyse, in small volumes of tissues, the most frequent antibiotics used in livestock. The method was applied to 87 samples of liver, muscle and kidney from 7 goats and 25 sheep disposed in SFS. According to questionnaires to farmers, the animals had not been treated with antibiotics, but analyses showed residues in 29% of the samples. Antibiotics were more frequent in goats (42.9%) than in sheep (24.2%), and oxytetracycline and trimethoprim were the most common (both 13.8%). Oxytetracycline, the most common antibiotic for livestock in Portugal, showed the highest concentration (1452.68 ng g-1). To our knowledge, this is the first study of presence of antibiotics in carrion from SFS. The concentrations of antibiotics in carrion do not seem to pose a risk of acute intoxication for adult Cinereous vultures. However, subtle and likely chronic exposure with unknown health consequences may occur, which requires more research. Moreover, the results of this first study can be used in future studies to assess the risk for avian scavengers.

Rapid analysis of components in Rhizoma Anemarrhenae by HPLC-DAD-MS and HPLC-DAD-TOFMS / 西安交通大学学报·英文版

A global quality control method based on high performance liquid chromatography (HPLC) coupled with diode array detection (DAD), single quadrupole mass spectrometry (MS) and time-of-flight mass spectrometry (TOFMS) was developed for simultaneous determination of seven major components (mangiferin, neomangiferin, timosaponin E1, timosaponin E, timosaponin BII, timosaponin BIII, and timosaponin A III) and identification of most components in extracts of Rhizoma Anemarrhenae (RA). HPLC analysis was performed on an Agilent SB-C18 column (4.6 mm X 150 mm, 5 μm) by gradient elution using acetonitrile and water-acetic acid(100:0.05, v/v) as the mobile phase. Seven major components in RA were successfully separated. This quantitative method was fully validated in respect of the following performance criteria, linearity, precision, repeatability, stability, accuracy, limits of detection (LOD) and quantification (LOQ). A formula database of known compounds in RA was established, against which, most of the reported components in this herbal extract were identified effectively based on the extract masses acquired by TOFMS. This qualitative and quantitative method was successfully used to analyze the components in 10 batches of RA samples collected from different regions in China. This global quality control method, which consisted of HPLC-DAD-MS assay of seven major components and unambiguous identification of nineteen components, is suitable for routine quantification and comprehensive quality control of RA.

HPLC-TOFMS in rapid separation and identification of chemical components in Oldenlandia diffusa and its injection preparations / 第二军医大学学报

Objective To rapidly separate and identify the chemical components in traditional Chinese herbal medicine of Oldenlandia diffusa and its injection preparations by high performance liquid chromatography-time of flight mass spectrometry (HPLC-TOFMS). Methods An Agilent Zorbax XDB-Q18 column (250 mm×4. 6 mm, 5 μm) was used for separation and identification of chemical components in Oldenlandia diffusa, with a mobile phase of 0. 3% acetic acid (A) and methanol (B) in gradient elution, 0-30 min, 30%-90%B. The flow rate was set at 1. 0 ml/min and the injection volume was 10 μl. The time- of-flight mass spectrometer was equipped with an EIS ion source. The scanning mass range was between m/z 100-1 000. Results The traditional Chinese medicine of Oldenlandia diffusa and its injection preparation were on-line separated and characterized by HPLC-TOFMS, and 11 chemical compounds were identified in Oldenlandia diffusa, 6 compounds in market injection preparation, and 2 compounds in the injection prepared by ourselves. Conclusion Chromatographic demonstration of chemical compounds in Oldenlandia diffusa in one run provides a foundation for the further studying the metabolism and mechanism of Oldenlandia diffusa and its injection preparations.

Rapid analysis of components in Rhizoma Anemarrhenae by HPLC-DAD-MS and HPLC-DAD-TOFMS / 药物分析学报

A global quality control method based on high performance liquid chromatography (HPLC) coupled with diode array detection (DAD), single quadrupole mass spectrometry (MS) and time-of-flight mass spectrometry (TOFMS) was developed for simultaneous determination of seven major components (mangiferin, neomangiferin, timosaponin E1, timosaponin E, timosaponin BⅡ, timosaponin BⅢ, and timosaponin AⅢ) and identification of most components in extracts of Rhizoma Anemarrhenae (RA). HPLC analysis was performed on an Agilent SB-C18 column (4.6 mm×150 mm, 5 μm) by gradient elution using acetonitrile and water-acetic acid(100∶0.05, v/v) as the mobile phase. Seven major components in RA were successfully separated. This quantitative method was fully validated in respect of the following performance criteria: linearity, precision, repeatability, stability, accuracy, limits of detection (LOD) and quantification (LOQ). A formula database of known compounds in RA was established, against which, most of the reported components in this herbal extract were identified effectively based on the extract masses acquired by TOFMS. This qualitative and quantitative method was successfully used to analyze the components in 10 batches of RA samples collected from different regions in China. This global quality control method, which consisted of HPLC-DAD-MS assay of seven major components and unambiguous identification of nineteen components, is suitable for routine quantification and comprehensive quality control of RA.