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Classification of commercial cultivars of Humulus lupulus L. (hop) by chemometric pixel analysis of two dimensional nuclear magnetic resonance spectra


The development of fast and effective spectroscopic methods that can detect most compounds in an untargeted manner is of increasing interest in plant extracts fingerprinting or profiling projects. Metabolite fingerprinting by nuclear magnetic resonance (NMR) is a fast growing field which is increasingly applied for quality control of herbal products, mostly via 1D 1H NMR coupled to multivariate data analysis. Nevertheless, signal overlap is a common problem in 1H NMR profiles that hinders metabolites identification and results in incomplete data interpretation. Herein, we introduce a novel approach in coupling 2D NMR datasets with principal component analysis (PCA) exemplified for hop resin classification. Heteronuclear multiple bond correlation (HMBC) profile maps of hop resins (Humulus lupulus) were generated for a comparative study of 13 hop cultivars. The method described herein combines reproducible metabolite fingerprints with a minimal sample preparation effort and an experimental time of ca. 28 min per sample, comparable to that of a standard HPLC run. Moreover, HMBC spectra provide not only unequivocal assignment of hop major secondary metabolites, but also allow to identify several isomerization and degradation products of hop bitter acids including the sedative principal of hop (2-methylbut-3-en-2-ol). We do believe that combining 2D NMR datasets to chemometrics, i.e. PCA, has great potential for application in other plant metabolome projects of (commercially relevant) nutraceuticals and or herbal drugs.

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One dimensional proton NMR


Two dimensional NMR


Heteronuclear multiple bond correlation


Heteronuclear single quantum correlation


Principal component analysis


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Dr. M. A. A. Farag, Dr. A. Porzel and Prof. Dr. L. A. Wessjohann thank the Alexander von Humboldt-foundation, Germany for financial support. We are grateful to Hopsteiner Inc. (and here especially Dr. Martin Biendl, Hallertau, and Harald Schwarz) and Orgentis GmbH, Gatersleben, Germany, for providing hop resins, hop bitter acid and hop flavonoid reference samples.

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Correspondence to Mohamed A. Farag or Ludger Wessjohann.

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Farag, M.A., Mahrous, E.A., Lübken, T. et al. Classification of commercial cultivars of Humulus lupulus L. (hop) by chemometric pixel analysis of two dimensional nuclear magnetic resonance spectra. Metabolomics 10, 21–32 (2014).

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  • Humulus lupulus L. hop
  • 2D NMR metabolomics
  • Bitter acids
  • Principal component analysis
  • Quality control
  • HMBC
  • Pixel analysis