Metabolomics Strategy Using High Resolution Mass Spectrometry Reveals Novel Biomarkers and Pain-Relief Effect of Traditional Chinese Medicine Prescription Wu-Zhu-Yu Decoction Acting on Headache Modelling Rats

Abstract

Headache is a common episodic or chronic neurologic disorder. Treatment options and diagnosis are restricted by an incomplete understanding of disease pathology and the lack of diagnostic markers. Wu-Zhu-Yu decoction (WZYD), a traditional Chinese medicine (TCM) formula containing four TCM herbs, is commonly used in the treatment of headache in China. To deeply understand more about headache and investigate the pain-relief mechanism of WZYD, a comprehensive metabolomics study combined with multivariate data processing strategy was carried out. An LC-high resolution mass spectrometry-based metabolomics approach was applied to characterize metabolic biomarker candidates. Multiple pattern recognition including principal component analysis-discriminant analysis, partial least squares-discriminant analysis and hierarchical cluster analysis were used to determine groups and confirm important variables. A total of 17 potential biomarkers were characterized and related metabolic pathways were identified. The study demonstrated that the established metabolomics strategy is a powerful approach for investigating the mechanism of headache attack and WZYD. In addition, the approach may highlight biomarkers and metabolic pathways and can capture subtle metabolite changes from headache, which may lead to an improved mechanism understanding of central nervous system diseases and TCM treatment.

Keywords: Wu-Zhu-Yu decoction; headache; high resolution mass spectrometry; metabolomics; traditional Chinese medicine.

Figure 1

The principal component analysis-discriminant analysis (PCA-DA) recognition based on the brain homogenates and plasma metabolomic profiling. (A) Brain metabolomics score plot of control group (green), model group (red), Wu-Zhu-Yu decoction (WZYD) group (blue) and flunarizine group (purple); (B) Loading plot, each dot represents a compound which contributed in group separation; (C) Plasma metabolomics score plot; (D) Plasma metabolomics loading plot.

Figure 2

(A) Fragmentation pathway and product ion spectrum of oxidized glutathione; (B) MS/MS spectrum of oxidized glutathione by mzCloud and (C) the comparison spectrum between the experimental and reference MS/MS spectrum by METLIN.

Figure 3

The hierarchical clustering heatmap of the potential biomarkers.

Figure 4

The metabolic pathways related to headache, as analyzed by MetaboAnalyst. The map was generated using the 379 reference map by KEGG. (A) Glutathione metabolism; (B) Nicotinate and nicotinamide metabolism; (C) Arginine and proline metabolism; (D) Purine metabolism; (E) Glycerophospholipid metabolism; (F) Tryptophan metabolism.

Figure 5

Part of the nitric oxide signaling pathway.

Figure 6

Tryptophan metabolism pathway.