The changes in total phenolic content (TPC), total flavonoid content (TFC), proline, malondialdehyde (MDA), H2O2, and antioxidant activity were assessed based on three model systems in three Achillea species (Achillea millefolium, A. nobilis, and A. filipendulina) growing under four irrigation regimes, including 100% FC (field capacity as normal irrigation) 75% FC (low stress), 50% FC (moderate stress), and 25% FC (severe stress) conditions. The highest TPC (47.13 mg tannic acid/g DW) and TFC (20.86 mg quercetin/g W) were obtained in A. filipendulina under moderate and severe stress conditions. In 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, the highest and the lowest antioxidant activity was obtained for A. millefolium (70.28%) and A. filipendulina (53.21%), respectively, while in the FTC model system A. nobilis revealed the highest antioxidant activity (1.934) in severe drought condition. In the linoleic model system, the highest antioxidant activity was observed under low drought stress condition in A. nobilis. MDA and H2O2 content were increased due to both low (75% FC) and moderate (50% FC) drought stress, but they were decreased under severe stress condition (25% FC). Furthermore, A. millefolium revealed the lowest H2O2 (4.96 nm/g FW) and MDA content (176.32 μmol/g). Investigation of the relationship among different metabolites showed a strong positive correlation with TPC and TFC. Finally, the moderate drought stress treatment (50% FC) was introduced as the optimum condition to obtain appreciable TPC and TFC,, while the highest antioxidant activity was obtained in severe stress condition (25%FC).

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Subgenus Perovskia of the extended genus of Salvia comprises several Central Asian medicinal and aromatic species, of which S. yangii and S. abrotanoides are the most widespread. These plants are cultivated in Europe as robust ornamentals, and several cultivars are available. However, their medicinal potential remains underutilized because of limited information about their phytochemical and genetic diversity. Thus, we combined an ultra-high performance liquid chromatography quadrupole time of flight mass spectrometry (UHPLC-QTOF-MS) based metabolomics with DNA barcoding approach based on trnH-psbA and ITS2 barcodes to clarify the relationships between these two taxa. Metabolomic analysis demonstrated that aerial parts are more similar than roots and none of the major compounds stand out as distinct. Sugiol in S. yangii leaves and carnosic acid quinone in S. abrotanoides were mostly responsible for their chemical differentiation, whereas in roots the distinction was supported by the presence of five norditerpenoids in S. yangii and two flavonoids and one norditerpenoid in S. abrotanoides. To verify the metabolomics-based differentiation, we performed DNA authentication that revealed S. yangii and S. abrotanoides to be very closely related but separate species. We demonstrated that DNA barcoding coupled with parallel LC-MS profiling constitutes a powerful tool in identification of taxonomically close Salvia species.