Integrative Effects of Stress- and Stress Tolerance-Inducing Elicitors on In Vitro Bioactive Compounds of Ajowan (Trachyspermum Ammi (L.) Sprague) Medicinal Plant

In the present study, the integrative effects of two sets of stress tolerance-inducing and stress-inducing elicitors, including polyethylene glycol–salicylic acid and NaCl–methyl jasmonate, were evaluated on regeneration efficiency, antioxidants activity and phytochemical profile of in vitro shoot cultures of ajowan. Combinations of salicylic acid (SA) (0, 10, 20, 40, 80 µM)–polyethylene glycol (PEG 6000) (0, 1, 2, 5%) and/or methyl jasmonate (MeJA) (0, 100, 200, 300 mg/L)–NaCl (0, 10, 15, 20 mM) were added to the shoot regeneration Murashige and Skoog medium containing KIN (1.50 mg/L) and NAA (0.25 mg/L) plant growth regulators. The number of regenerated shoots and the in vitro rooting decreased significantly with increasing PEG and NaCl levels. SA (40 µM) reduced the adverse effect of PEG on the number of regenerated shoots and in vitro rooting. The activities of catalase, superoxide dismutase and peroxidase enzymatic antioxidants were significantly increased in SA (80 µM)–PEG (5%) and MeJA (300 mg/L)–NaCl (20 mM) treated plants. The gas chromatography–mass spectrometry (GC–MS)-profiling revealed quantitative and qualitative phytochemical differences between control and SA–PEG and MeJA–NaCl treated plants. The greatest means of p-cymene and thymol bioactive compounds were obtained from in vitro shoots treated with 5% PEG + 40 µM SA. The interaction of PEG–SA was better than NaCl–MeJA elicitors in terms of the content of valuable γ-terpinene, p-cymene and thymol compounds. The inter-simple sequence repeats (ISSR) markers proved the genetic stability of in vitro regenerated plants. The presented protocol is useful for large-scale sustainable production of secondary metabolites (SMs) of medicinal plants. Secondary metabolites are important plants defense molecules. Stress induction and/or defense machine stimulation are two practical strategies to increase the accumulation of secondary metabolites. The combined effects of stress tolerance-inducing salicylic acid and stress-inducing polyethylene glycol were evaluated in indirect shoot culture of ajowan. Levels of p-cymene and thymol increased in PEG–SA treated ajowan plants.