Metabonomics Study on the Effects of the Ginsenoside Rg3 in a -Cyclodextrin-Based Formulation on Tumor-Bearing Rats by a Fully Automatic Hydrophilic Interaction/Reversed-Phase Column-Switching HPLC-ESI-MS Approach Yuan Wang, † Jiangshan Wang, † Ming Yao, ‡ Xinjie Zhao, † Jens Fritsche, § Philippe Schmitt-Kopplin, | Zongwei Cai, ⊥ Dafang Wan, ‡ Xin Lu, † Shengli Yang, †,‡,# Jianren Gu, ‡ Hans Ulrich Ha ¨ ring, ¶ Erwin D. Schleicher, ¶ Rainer Lehmann,* ,¶ and Guowang Xu* ,† National Chromatography Research and Analysis Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 16023 Dalian, China, State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Shanghai Jiao Tong University, 200032 Shanghai, China, Immatics Biotechnologies GmbH, 72076 Tuebingen, Germany, Helmholtz-Zentrum MuenchensGerman Research Center for Environmental Health, Institute for Ecological Chemistry, Ingoldstaedter Landstrasse 1 D-85764 Neuherberg, Germany, Department of Chemistry, Hong Kong Baptist University, Kowloon, Hong Kong SAR, China, Shanghai Research Center of Biotechnology, Chinese Academy of Sciences, Shanghai 200032, China, and Department of Internal Medicine 4, Division of Clinical Chemistry and Pathobiochemistry (Central Laboratory), University Hospital Tuebingen, D-72076 Tuebingen, Germany The goal of this study was the application of a novel, fully automatic column-switching approach in a metabonomics study combining the orthogonal selectivities of hydrophilic interaction chromatography (HILIC) and reversed-phase chromatography. The temporal, pharmacodynamic effects of the ginsenoside Rg3 on the metabonome in urine of healthy and liver-tumor-bearing rats have been investi- gated. Within a total analysis time of 52 min we detected 5686 polar, and on the second column an additional 1808 apolar, urinary metabolite ions. The administration of a single, high dose of Rg3 in a -cyclodextrin-based formulation led to a considerable change of the metabolic pattern in cancer rats during 3 days studied. Seventeen biomarker candidates including three apolar metabolites, which were not retained on the HILIC column, were detected. Overall, the results suggest that the developed liquid chromatography-mass spectrometry strategy is a promising tool in metabonomics studies for global analy- sis of highly complex biosamples. It may not only increase the number of discovered biomarkers but consequently improve the comprehensive information on metabolic changes in a fully automatic manner. Over the past decade, the application of metabonomics analyses of body fluids increased considerably in various research fields like pharmaceutical discovery and development, 1,2 nutrition and epidemiology investigations, 3 research projects for the evalu- ation of drug toxicity, 4 studies for the diagnosis and treatment of diseases, 5 and discovery of novel biomarkers. 6–10 For the analysis of body fluids urine is the most favorable sample material, because it can be collected noninvasively. Furthermore, urine contains a great number of metabolites indicating the metabolic state of the organism 5 and reflecting specific traits to perturbations. 11–17 On the basis of the aqueous character of urine all excreted metabolites * Corresponding authors. Fax: +86-411-84379559 (G.X.), +49-7071-5348 (R.L.). 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