The production of self‐pollinated plants could be important for improving medicinal plants secondary metabolites. In this study, 11 Thymus populations from eight species were evaluated to determine the effect of self and open pollination on agro‐morphological characteristics, total phenolic content (TPC), essential oil (EO) content, and EO components. Inbreeding led to some positive effects of above mentioned traits in most of the studied populations. Total phenolic content ranged from 7.07 to 52.69 mg tannic acid equivalents (TAE) g−1 dry weight (DW) in open pollinated derived populations, while it varied from 1.2 to 55.03 mg TAE g−1 DW in self‐pollinated ones. Under open and self‐pollination condition, the highest EO content was obtained in T. trautvetteri (3.37 %) and T. pubescens (1.96 %), respectively. Gas chromatography‐mass spectrometry (GC/MS) identified 42 compounds including thymol, carvacrol, linalool, p‐cymene, γ‐terpinene, terpinen‐4‐ol, and α‐terpineol as the main compounds. In most cases, selfed plants compared to open pollinated ones, revealed higher thymol content. T. daenensis‐1 showed a significant increase in thymol content (from 25.22 % to 74.3 %) due to self‐pollination. Moreover, self‐pollination led to emergence of some new compounds. Carvacrol methyl ether was the constituents of Thymus EO that are being reported in self‐pollinated populations. Finally, inbreeding in Thymus might be suggested as a useful tool to increase genetic homogeneity for the selection of superior plants for improving secondary metabolite.
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