, Volume 8, Issue 1, pp 99–108 | Cite as

Profiling of the charged metabolites of traditional herbal medicines using capillary electrophoresis time-of-flight mass spectrometry

  • Keiko Iino
  • Masahiro Sugimoto
  • Tomoyoshi Soga
  • Masaru Tomita
Original Article


The quantification of a small number of bioactive components in herbal medicines is often inadequate when attempting to elucidate a medicine’s biological effects. Despite rapid advances in analytical technologies, obtaining comprehensive metabolomic profiles of herbal medicines remains difficult, due to the complexity of natural product mixtures. Toki-Shakuyaku-San is a Chinese medicine used widely to treat gynecological and obstetric disorders, such as infertility, dysmenorrhea, toxemia during pregnancy and neural dysfunction. It consists of Angelica acutiloba Radix (Toki), Cnidium officinale Rhizoma (Senkyu), Paeonia lactiflora Radix (Shakuyaku), Atractylodes lancea Rhizoma (Sojutsu), Alisma orientale Rhizoma (Takusha) and Poria cocos Hoelen (Bukuryo). To elucidate the composition of these herbal medicines individually, we conducted non-targeted profiling analyses of extracts of these herbs using capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS), which allows the simultaneous quantification of hundreds of charged metabolites. In total, 737 ± 183.1 (average ± SD) metabolite-derived features were observed, and of these, 119 metabolites were identified. Score plots of principal component analysis (PCA) showed a clear cluster including Shakuyaku, Bukuryo, and Sojutsu, while the other three herbs were distributed over PCA spaces. Loading plots revealed that amino acids and shikimate-derived alkaloids were the predominant metabolite constituents. Hierarchical clustering analysis revealed that few clusters overlapped in the herbal medicines tested. This report is the first demonstration of the characterization of a herbal medicine using large-scale metabolomic analysis, which is complementary to traditional quality control methods.


Capillary electrophoresis time-of-flight mass spectrometry Herbal medicine Charged metabolite Metabolomic profiling 



This work was supported by research funds from the Yamagata Prefectural Government and the city of Tsuruoka. We thank Dr. Kazuko Otomo for technical assistance, and Wanjun Kong and Guo Jing for fruitful discussions.

Supplementary material

11306_2011_290_MOESM1_ESM.doc (1.2 mb)
Supplementary material 1 (DOC 1,259 kb)


  1. Akase, T., Onodera, S., Matsushita, R., & Tashiro, S. (2004). A comparative study of laboratory parameters and symptoms effected by Toki-Shakuyaku-San and an iron preparation in rats with iron-deficiency anemia. Biological and Pharmaceutical Bulletin, 27, 871–878.PubMedCrossRefGoogle Scholar
  2. Bohrmann, H., Stahl, E., & Mitsuhashi, H. (1967). Studies of the constituents of umbelliferae plants. 8. Chromatographic studies on the constituents of Cnidium officinale Makino. Chemical and Pharmaceutical Bulletin, 15, 1606–1608.CrossRefGoogle Scholar
  3. Brown, M., Dunn, W. B., Dobson, P., et al. (2009). Mass spectrometry tools and metabolite-specific databases for molecular identification in metabolomics. Analyst, 134, 1322–1332.PubMedCrossRefGoogle Scholar
  4. Calixto, J. B. (2000). Efficacy, safety, quality control, marketing and regulatory guidelines for herbal medicines (phytotherapeutic agents). Brazilian Journal of Medical and Biological Research, 33, 179–189.PubMedCrossRefGoogle Scholar
  5. Chan, K. (2003). Some aspects of toxic contaminants in herbal medicines. Chemosphere, 52, 1361–1371.PubMedCrossRefGoogle Scholar
  6. Chan, E. C., Yap, S. L., Lau, A. J., Leow, P. C., Toh, D. F., & Koh, H. L. (2007). Ultra-performance liquid chromatography/time-of-flight mass spectrometry based metabolomics of raw and steamed Panax notoginseng. Rapid Communications in Mass Spectrometry, 21, 519–528.PubMedCrossRefGoogle Scholar
  7. Cheever, K. L., Richards, D. E., & Plotnick, H. B. (1982). The acute oral toxicity of isomeric monobutylamines in the adult male and female rat. Toxicology and Applied Pharmacology, 63, 150–152.PubMedCrossRefGoogle Scholar
  8. Chen, L., Qi, J., Chang, Y. X., Zhu, D., & Yu, B. (2009). Identification and determination of the major constituents in Traditional Chinese Medicinal formula Danggui-Shaoyao-San by HPLC-DAD-ESI-MS/MS. Journal of Pharmaceutical and Biomedical Analysis, 50, 127–137.PubMedCrossRefGoogle Scholar
  9. Daly, J. W. (2007). Caffeine analogs: Biomedical impact. Cellular and Molecular Life Sciences, 64, 2153–2169.PubMedCrossRefGoogle Scholar
  10. Ganzera, M. (2008). Quality control of herbal medicines by capillary electrophoresis: Potential, requirements and applications. Electrophoresis, 29, 3489–3503.PubMedCrossRefGoogle Scholar
  11. Gong, F., Liang, Y. Z., Cui, H., Chau, F. T., & Chan, B. T. (2001). Determination of volatile components in peptic powder by gas chromatography-mass spectrometry and chemometric resolution. Journal of Chromatography A, 909, 237–247.PubMedCrossRefGoogle Scholar
  12. Hatip-Al-Khatib, I., Egashira, N., Mishima, K., et al. (2004). Determination of the effectiveness of components of the herbal medicine Toki-Shakuyaku-San and fractions of Angelica acutiloba in improving the scopolamine-induced impairment of rat’s spatial cognition in eight-armed radial maze test. Journal of Pharmacological Sciences, 96, 33–41.PubMedCrossRefGoogle Scholar
  13. He, C. N., Peng, Y., Xu, L. J., et al. (2010). Three new oligostilbenes from the seeds of Paeonia suffruticosa. Chemical and Pharmaceutical Bulletin, 58, 843–847.CrossRefGoogle Scholar
  14. Hurtado-Fernandez, E., Gomez-Romero, M., Carrasco-Pancorbo, A., & Fernandez-Gutierrez, A. (2010). Application and potential of capillary electroseparation methods to determine antioxidant phenolic compounds from plant food material. Journal of Pharmaceutical and Biomedical Analysis, 53, 1130–1160.PubMedCrossRefGoogle Scholar
  15. Kanehisa, M., Goto, S., Furumichi, M., Tanabe, M., & Hirakawa, M. (2010). KEGG for representation and analysis of molecular networks involving diseases and drugs. Nucleic Acids Research, 38, D355–D360.PubMedCrossRefGoogle Scholar
  16. Kim, M. R., Abd El-Aty, A. M., Choi, J. H., Lee, K. B., & Shim, J. H. (2006). Identification of volatile components in Angelica species using supercritical-CO2 fluid extraction and solid phase microextraction coupled to gas chromatography-mass spectrometry. Biomedical Chromatography, 20, 1267–1273.PubMedCrossRefGoogle Scholar
  17. Kitajima, J., Kamoshita, A., Ishikawa, T., et al. (2003). Glycosides of Atractylodes lancea. Chemical and Pharmaceutical Bulletin, 51, 673–678.CrossRefGoogle Scholar
  18. Kono, N., Arakawa, K., Ogawa, R., et al. (2009). Pathway projector: Web-based zoomable pathway browser using KEGG atlas and Google Maps API. PLoS One, 4, e7710.PubMedCrossRefGoogle Scholar
  19. Lao, Y. M., Jiang, J. G., & Yan, L. (2009). Application of metabonomic analytical techniques in the modernization and toxicology research of traditional Chinese medicine. British Journal of Pharmacology, 157, 1128–1141.PubMedCrossRefGoogle Scholar
  20. Li, X. N., Cui, H., Song, Y. Q., Liang, Y. Z., & Chau, F. T. (2003). Analysis of volatile fractions of Schisandra chinensis (Turcz.) Baill. using GC–MS and chemometric resolution. Phytochemical Analysis, 14, 23–33.PubMedCrossRefGoogle Scholar
  21. Liang, X. M., Jin, Y., Wang, Y. P., Jin, G. W., Fu, Q., & Xiao, Y. S. (2009). Qualitative and quantitative analysis in quality control of traditional Chinese medicines. Journal of Chromatography A, 1216, 2033–2044.PubMedCrossRefGoogle Scholar
  22. Liao, J. F., Jan, Y. M., Huang, S. Y., Wang, H. H., Yu, L. L., & Chen, C. F. (1995). Evaluation with receptor binding assay on the water extracts of ten CNS-active Chinese herbal drugs. Proceedings of the National Science Council, Republic of China. Part B, Life Sciences, 19, 151–158.PubMedGoogle Scholar
  23. Liu, S., Yi, L. Z., & Liang, Y. Z. (2008). Traditional Chinese medicine and separation science. Journal of Separation Science, 31, 2113–2137.PubMedCrossRefGoogle Scholar
  24. Lu, G. H., Chan, K., Liang, Y. Z., et al. (2005). Development of high-performance liquid chromatographic fingerprints for distinguishing Chinese Angelica from related umbelliferae herbs. Journal of Chromatography A, 1073, 383–392.PubMedCrossRefGoogle Scholar
  25. Ma, C. M., Winsor, L., & Daneshtalab, M. (2007). Quantification of spiroether isomers and herniarin of different parts of Matricaria matricarioides and flowers of Chamaemelum nobile. Phytochemical Analysis, 18, 42–49.PubMedCrossRefGoogle Scholar
  26. Miller, F. G., Emanuel, E. J., Rosenstein, D. L., & Straus, S. E. (2004). Ethical issues concerning research in complementary and alternative medicine. JAMA, 291, 599–604.PubMedCrossRefGoogle Scholar
  27. Monteith, D. K., Emmerling, M. R., Garvin, J., & Theiss, J. C. (1996). Cytotoxicity study of tacrine, structurally and pharmacologically related compounds using rat hepatocytes. Drug and Chemical Toxicology, 19, 71–84.PubMedCrossRefGoogle Scholar
  28. Monton, M. R., & Soga, T. (2007). Metabolome analysis by capillary electrophoresis–mass spectrometry. Journal of Chromatography A, 1168, 237–246. discussion 236.PubMedCrossRefGoogle Scholar
  29. Murray, R. H., & Rubel, A. J. (1992). Physicians and healers—Unwitting partners in health care. New England Journal of Medicine, 326, 61–64.PubMedCrossRefGoogle Scholar
  30. Namba, T., & Tsuda, Y. (1998). Shoyakugakugairon (3rd ed.). Tokyo: Nankodo.Google Scholar
  31. Ohta, H., Ni, J. W., Matsumoto, K., Watanabe, H., & Shimizu, M. (1993). Peony and its major constituent, paeoniflorin, improve radial maze performance impaired by scopolamine in rats. Pharmacology, Biochemistry and Behavior, 45, 719–723.CrossRefGoogle Scholar
  32. Ren, M. T., Chen, J., Song, Y., Sheng, L. S., Li, P., & Qi, L. W. (2008). Identification and quantification of 32 bioactive compounds in Lonicera species by high performance liquid chromatography coupled with time-of-flight mass spectrometry. Journal of Pharmaceutical and Biomedical Analysis, 48, 1351–1360.PubMedCrossRefGoogle Scholar
  33. Saeed, A. I., Bhagabati, N. K., Braisted, J. C., et al. (2006). TM4 microarray software suite. Methods in Enzymology, 411, 134–193.PubMedCrossRefGoogle Scholar
  34. Sato, S., Soga, T., Nishioka, T., & Tomita, M. (2004). Simultaneous determination of the main metabolites in rice leaves using capillary electrophoresis mass spectrometry and capillary electrophoresis diode array detection. The Plant Journal, 40, 151–163.PubMedCrossRefGoogle Scholar
  35. Smyth, D. D., & Penner, S. B. (1995). Renal I1-imidazoline receptor-selective compounds mediate natriuresis in the rat. Journal of Cardiovascular Pharmacology, 26(Suppl. 2), S63–S67.PubMedGoogle Scholar
  36. Soga, T., Baran, R., Suematsu, M., et al. (2006). Differential metabolomics reveals ophthalmic acid as an oxidative stress biomarker indicating hepatic glutathione consumption. The Journal of Biological Chemistry, 281, 16768–16776.PubMedCrossRefGoogle Scholar
  37. Soga, T., Ohashi, Y., Ueno, Y., Naraoka, H., Tomita, M., & Nishioka, T. (2003). Quantitative metabolome analysis using capillary electrophoresis mass spectrometry. Journal of Proteome Research, 2, 488–494.PubMedCrossRefGoogle Scholar
  38. Sugimoto, M., Koseki, T., Hirayama, A., et al. (2010a). Correlation between sensory evaluation scores of Japanese sake and metabolome profiles. Journal of Agriculture and Food Chemistry, 58, 374–383.CrossRefGoogle Scholar
  39. Sugimoto, M., Wong, D. T., Hirayama, A., Soga, T., & Tomita, M. (2010b). Capillary electrophoresis mass spectrometry-based saliva metabolomics identified oral, breast and pancreatic cancer-specific profiles. Metabolomics, 6, 78–95.PubMedCrossRefGoogle Scholar
  40. Wambach, G., & Casals-Stenzel, J. (1983). Structure-activity relationship of new steroidal aldosterone antagonists. Comparison of the affinity for mineralocorticoid receptors in vitro and the antialdosterone activity in vivo. Biochemical Pharmacology, 32, 1479–1485.PubMedCrossRefGoogle Scholar
  41. Wang, H. X., Liu, C. M., Liu, Q., & Gao, K. (2008). Three types of sesquiterpenes from rhizomes of Atractylodes lancea. Phytochemistry, 69, 2088–2094.PubMedCrossRefGoogle Scholar
  42. Wang, Y., Liu, H., Mckenzie, G., et al. (2010). Kynurenine is an endothelium-derived relaxing factor produced during inflammation. Nature Medicine, 16, 279–285.PubMedCrossRefGoogle Scholar
  43. Wang, Z. G., & Ren, J. (2002). Current status and future direction of Chinese herbal medicine. Trends in Pharmacological Sciences, 23, 347–348.PubMedCrossRefGoogle Scholar
  44. Wang, J., Van Der Heijden, R., Spruit, S., et al. (2009). Quality and safety of Chinese herbal medicines guided by a systems biology perspective. Journal of Ethnopharmacology, 126, 31–41.PubMedCrossRefGoogle Scholar
  45. Wang, Y., Zhang, M., Ruan, D., et al. (2004). Chemical components and molecular mass of six polysaccharides isolated from the sclerotium of Poria cocos. Carbohydrate Research, 339, 327–334.PubMedCrossRefGoogle Scholar
  46. Weng, Q., & Jin, W. (2002). Carbon fiber bundle–Au–Hg dual-electrode detection for capillary electrophoresis. Journal of Chromatography A, 971, 217–223.PubMedCrossRefGoogle Scholar
  47. Xie, G., Plumb, R., Su, M., et al. (2008). Ultra-performance LC/TOF MS analysis of medicinal Panax herbs for metabolomic research. Journal of Separation Science, 31, 1015–1026.PubMedCrossRefGoogle Scholar
  48. Yi, T., Leung, K. S., Lu, G. H., & Zhang, H. (2007). Comparative analysis of Ligusticum chuanxiong and related umbelliferous medicinal plants by high performance liquid chromatography–electrospray ionization mass spectrometry. Planta Medica, 73, 392–398.PubMedCrossRefGoogle Scholar
  49. Zhang, A., Sun, H., Wang, Z., Sun, W., Wang, P., & Wang, X. (2010). Metabolomics: Towards understanding traditional Chinese medicine. Planta Medica, 76, 2026–2035.PubMedCrossRefGoogle Scholar
  50. Zhao, M., Xu, L. J., & Che, C. T. (2008). Alisolide, alisols O and P from the rhizome of Alisma orientale. Phytochemistry, 69, 527–532.PubMedCrossRefGoogle Scholar
  51. Zhu, Y. Y., Zhu-Ge, Z. B., Wu, D. C., et al. (2007). Carnosine inhibits pentylenetetrazol-induced seizures by histaminergic mechanisms in histidine decarboxylase knock-out mice. Neuroscience Letters, 416, 211–216.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Keiko Iino
    • 1
    • 2
  • Masahiro Sugimoto
    • 1
    • 3
  • Tomoyoshi Soga
    • 1
    • 2
    • 3
  • Masaru Tomita
    • 1
    • 2
    • 3
  1. 1.Institute for Advanced BiosciencesKeio UniversityTsuruokaJapan
  2. 2.Department of Environment and Information StudiesKeio UniversityFujisawaJapan
  3. 3.Systems Biology Program, Graduate School of Media and GovernanceKeio UniversityFujisawaJapan

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