In this study, phenolic derivatives and antioxidant activities of fourteen Ziziphora clinopodioides populations, as well as LC-MS/MS analysis of three specific flavonoids were evaluated. Generally, high contents of phenolic derivatives were found in shoot extracts compared to roots. LC-MS/MS, a powerful analytical technique, was employed for the identification and quantify the individual flavonoids in Z. clinopodioides populations’ extracts, in a quantity order of quercetin > rutin > apigenin. Scavenging activity by DPPH and FRAP was performed, and accordingly, in the shoot, the highest values for the DDPH were 4.61 ± 0.4 and 7.59 ± 0.26 µg ml− 1 in populations 1 and 13, respectively, and for the FRAP were 328.61 ± 5.54 and 292.84 ± 2.85 mg g DW− 1, in populations 6 and 1 respectively. Multivariate analysis results of the principal component analysis indicated the amount of polyphenols to be useful indicators in differentiating the geographical localities which explain 92.7% of the total variance. According to the results of hierarchical cluster analysis, the studied populations could be separated into two groups in that the contents of phenolic derivatives and antioxidant activities of different plant parts. Both shoot and root samples were well discriminated with the orthogonal partial least squares discriminant analysis (R2X: 0.861; Q2: 0.47) model. The validity of the model was confirmed by using receiver operating characteristic curve analysis and permutation tests. Such data make an important addition to our current knowledge of Ziziphora chemistry and are decisive in the identification of germplasms with a homogeneous phytochemical profile, high chemical content and bioactivity. The present results could also be helpful for the potential application of Z. clinopodioides in different kinds of industries as natural antioxidants.

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