Combined NMR and LC-MS Analysis Reveals the Metabonomic Changes in Salvia miltiorrhiza Bunge Induced by Water Depletion

Abstract

Plant metabonomic analysis is essential for understanding plant systems responses to osmotic stresses. To understand the comprehensive metabolic responses of Salvia miltiorrhiza Bunge (SMB) to continuous and exhaustive water depletion, we characterized the SMB metabonomic variations induced by three different drying processes using the combined NMR and LC-DAD-MS method. NMR results showed that SMB extracts were dominated by 29 primary metabolites such as sugars, carboxylic acids and amino acids, which were comprehensively reported for the first time, and 8 secondary metabolites including polyphenolic acids and diterpenoids. LC-DAD-MS methods detected 44 secondary metabolites, among which 5 polyphenolic acids together with genipin, umbelliferone and tormentic acid were found for the first time in this plant. We found that aqueous methanol was efficient in extracting both primary metabolites and polyphenolic acids, whereas chloroform-methanol was effective in selectively extracting diterpenoids. We further found that air- and sun-drying markedly affected both primary and secondary metabolisms of SMB by enhancing tanshinone and glutamate-mediated proline biosynthesis and altering carbohydrate and amino acid metabolisms. The shikimate-mediated biosynthesis of polyphenolic acids was promoted by air-drying but suppressed by sun-drying. These findings fill the gap of our understandings to the metabolic responses of S. miltiorrhiza Bunge to water depletion and demonstrated effectiveness of the combined NMR and LC-DAD-MS methods in plant metabonomic analysis.

Plant Metabolomics: A Characterisation of Plant Responses to Abiotic Stresses

“…However, under water deficit conditions, the concentration of these compounds was found to be significantly higher in pCOR15a-Myb4 transgenic lines than in wt and to be comparable to that found in pCaMV35S-Myb4 transgenic plants (Vannini et al, 2007). Dai et al (2010b) have performed a systematic characterisation of the metabolic changes induced by water depletion in the roots of the medicinal plant, Salvia miltiorrhiza Bunge, comparing the results obtained using two metabolite analysis techniques ( 1 H-NMR and LC-DAD-MS) and four extraction methods based on different solvents (see also section 2.2). As phytomedicines are usually either air-dried or sun-dried for the purposes of transportation, storage or pharmacological requirements, the effect of these two different drying processes on the metabolite composition was also investigated and compared with a freeze-drying process, taken as the control.…”

“…Different effects of airand sun-drying processes were also highlighted on secondary metabolism by the LC-DAD-MS method, with air-drying enhancing the biosynthesis of oligomeric caffeic acids and tanshinones and sun-drying promoting the biosynthesis of tanshinones but inhibiting that of polyphenolic acids. The differences in metabolite content variations between the two sample groups was suggested to be attributable to the different drying speed of the two methods and to the concomitant occurrence of light and thermal stresses in sun-, but not in air-dried roots (Dai et al, 2010b). This study has the merit of showing the effectiveness of the combination of two different analytical approaches ( 1 H-NMR and LC-DAD-MS) and of highlighting the importance to carefully consider and optimise the extraction method when metabolomic analyses are performed.…”

“…A comparison between MS and NMR techniques have been reviewed by Krishnan et al (2005). More recently Dai et al (2010aDai et al ( , 2010b successfully applied and combined NMR and LC-DAD-MS analysis to investigate the metabolic variations of three cultivars of Salvia milthiorrhiza Bunge (SMB), as well as changes induced by water depletion. The combination of these two analytical techniques has allowed the detection of both the primary and secondary metabolites content.…”

Tanshinone IIA Exhibits Anticonvulsant Activity in Zebrafish and Mouse Seizure Models

“…One-and two-dimensional NMR analyses of the isolated peaks showed that they are structurally related to each other as tanshinones (Table 1), with the chemical shifts matching those listed in the literature. 16 Active peaks were identified as 15,16-dihydrotanshinone I (1), cryptotanshinone (2), tanshinone IIA (3), and miltirone (4) (Figure 3), with average yields of 2.8% for miltirone to 13.1% for cryptotanshinone (Table 1).…”

NMR metabolite comparison of local pigmented rice in Yogyakarta

“…The TSP was used as a reference at δ 0.00. The metabolites were identified based on the 1 H-NMR spectra and comparisons with the NMR spectra of the reference compounds and published literatures (Fan 1996;Dai et al 2010;Kim et al 2010;Jones et al 2011;Nuringtyas et al 2012;Nam et al 2015;Uawisetwathana et al 2015;Liu et al 2017;Pramai et al 2017).…”