Metabolomics

, Volume 8, Issue 2, pp 335–346 | Cite as

1H NMR-based metabolic profiling and target analysis: a combined approach for the quality control of Thymus vulgaris

  • Valerio Pieri
  • Sonja Sturm
  • Christoph Seger
  • Chlodwig Franz
  • Hermann Stuppner
Original Article

Abstract

The characterization of T. vulgaris plant material for quality control purposes was performed by NMR-based methods. Direct extraction of 141 T. vulgaris samples with DMSO-d 6 enabled the obtainment of crude extracts with a representative composition in terms of both volatile and non-volatile constituents. The acquisition of 600 MHz 1H NMR spectra resulted in a dataset which was analyzed by a combination of metabolic profiling and target analysis approaches. Preliminary analysis of the 1H NMR spectra was performed by principal component analysis, which revealed sample discrimination on a chemotype basis (thymol, carvacrol and linalool chemotypes). Further minor discriminative constituents were identified as p-cymene, γ-terpinene, rosmarinic acid, and 3,4,3′,4′-tetrahydroxy-5,5′-diisopropyl-2,2′-dimethylbiphenyl. Metabolite identification was accomplished by 1D and 2D NMR techniques and supported by spiking experiments. Fast dereplication of constituents not available as reference compounds was performed by HPLC–SPE–NMR experiments. A targeted approach based on qHNMR was validated for quantification of the identified secondary metabolites. Validation was performed in terms of precision (intra-day RSD ≤ 4.51%, inter-day RSD ≤ 4.18%), repeatability (RSD ≤ 2.30%), accuracy (recovery rates within 93.4 and 103.4%), linearity (correlation coefficients ≥ 0.9990), robustness, and stability. The amount of the dominant monoterpene in thymol, carvacrol, and linalool chemotypes was respectively found to be within 0.4–2.6, 0.7–2.3, and 1.1–3.6% (w/w). Variable amounts of the precursors p-cymene and γ-terpinene were found in thymol and carvacrol chemotypes. The highest amount of rosmarinic acid and 3,4,3′,4′-tetrahydroxy-5,5′-diisopropyl-2,2′-dimethylbiphenyl in the analyzed samples was respectively 4.6 and 0.4% (w/w). Since quantification is performed on a weight basis, the essential oil content can be estimated based on the sum of the quantified monoterpenes. The NMR-based analysis of T. vulgaris represents a more comprehensive approach in comparison to traditional chromatographic methods such as GC and LC, respectively employed for the analysis of volatile and non-volatile constituents. Further advantages lie in the simple sample preparation, rapidity and reproducibility of the NMR analysis.

Keywords

Thymus vulgaris 1H NMR Metabolic profiling Target analysis Principal component analysis qHNMR 

Notes

Acknowledgment

This work was financially supported by Bionorica Research GmbH, 6020 Innsbruck, Austria.

Supplementary material

11306_2011_317_MOESM1_ESM.docx (4.8 mb)
Supplementary material 1 (DOCX 4888 kb)

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Valerio Pieri
    • 1
  • Sonja Sturm
    • 1
  • Christoph Seger
    • 1
  • Chlodwig Franz
    • 2
  • Hermann Stuppner
    • 1
  1. 1.Institute of Pharmacy/Pharmacognosy, Center for Molecular Biosciences InnsbruckUniversity of InnsbruckInnsbruckAustria
  2. 2.Institute for Applied Botany and PharmacognosyUniversity of Veterinary Medicine ViennaViennaAustria

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