Induction of bioactive constituents and antioxidant enzyme activities in Achillea fragrantissima (Forskal) callus cultures using ZnO nanoparticles

Abstract

The medicinally effective plant Achillea fragrantissima exhibits a magnitude of pharmacological activities. In this study, the effects of different ZnONP concentrations on antioxidant enzymes, bioactive secondary metabolites, redox potential, and molecular changes in A. fragrantissima callus cultures were investigated. First, the concentrations of the growth regulators 2,4-D and BA were optimized using Murashige and Skoog (MS) medium. The MS medium was then administered with 2,4-D and BA at its optimal dosage (1.0 mg.L−1); afterward, different ZnONP supplements (0.0, 5.0, 10.0, 15.0, and 20.0 mg.L−1) were added. ZnONPs resulted in many physiological and molecular responses. ZnONPs significantly increased POD, APX, and SOD activities. While 10.0 mg.L−1 ZnONPs significantly increased POD and APX activities, 15.0 mg.L−1 ZnONPs significantly increased SOD. However, CAT activity gradually decreased with ZnONPs. Metabolically, ZnONPs increased phenolics, flavonoids, alkaloids, and saponin levels. Phenolic levels peaked at 20.0 mg.L−1, flavonoids at 15.0 mg.L−1, and alkaloids and saponins at 10.0 mg.L−1. Terpenoids were more prevalent at lower levels of ZnONPs. With 15.0 and 10.0 mg.L−1 giving the maximum activity, ZnONPs enhanced the DPPH activity and TAC of the callus culture extracts, respectively. RAPD and ISSR fingerprinting were applied using 12 random and ISSR primers to evaluate the genetic stability of ZnONP-induced callus cultures. Six RAPD primers showed 83% polymorphism while the seven ISSR primers achieved 30% polymorphism. Consequently, DNA mutations may have been induced by ZnONPs and caused DNA fragments to either appear or disappear in RAPD and ISSR callus profiles. The dendrogram based on RAPD and ISSR combined data showed that by increasing ZnONP concentration the genetic differentiation among callus cultures was elevated. In conclusion, higher accumulation of secondary metabolites and redox activity were increased in A. fragrantissima callus cultures using low ZnONPs (10.0 mg.L−1) concentration.