Is acute low-dose gamma irradiation an effective elicitor for secondary metabolism in Leontopodium alpinum (Cass.) callus culture?

Leontopodium alpinum Cass. (Edelweiss) is a protected species in some European countries and has become of growing interest for exploitation by the food supplements and cosmetics industry, due to its valuable composition in bioactive and biocompatible compounds. Small scale field cultivation and in vitro biotechnological approaches are economically viable alternatives to the production via chemical synthesis. In this study, we aimed to compare two performant L. alpinum callus culture lines and to investigate the effect of acute low dose gamma irradiation on biomass accumulation (Gi), total phenolic content (TPC), total flavonoid content (TFC), antioxidant activity (DPPH) and secondary metabolites’ profile. A purple, pigmented line (PC) was obtained under lightning conditions and in the absence of light, an unpigmented greenish line (GC) was established. Both lines displayed impressive proliferative capacity after 14 days of culture, with an 18.31 ( ± 1.83) fold mass increase for PC and a 13.27 ( ± 2.42) for GC. In respect to Gi, TPC, TFC and DPPH, the purple callus line significantly outperformed the unpigmented line. According to the UPLC-HRMS analysis of the methanolic extracts, both callus lines displayed similar metabolic profiles, having similar quantities of 3,5-dicaffeoylquinic acid, leontopodic acids A and B, but 60% higher concentration of chlorogenic acid in the GC line. In the methanolic callus extracts, 9 economically important phenylpropanoid derivatives were identified. The two callus lines responded differently to low dose gamma irradiation. Although the control values measured for Gi, TPC, TFC and DPPH were significantly higher in the PC line, this line was generally negatively impacted by the gamma treatments. On the contrary, higher, but not significant, values were registered in some GC variants post irradiation with 25, 30 and 40 Gy. UPLC-HRMS investigations emphasized the overproduction of 7 compounds, post irradiation with all doses, for the PC line, while for the unpigmented line only one compound was stimulated by all irradiation treatments. The highest stimulative effect was obtained for the 35 and 40 Gy experimental variants, in the GC line, where dihydroxybenzoic acid glucoside accumulation was increased by 160%. The results of this study showed, for the first time, the different responses of two distinct L. alpinum callus lines using gamma irradiation elicitation method, which can be successfully applied to stimulate the production of certain industrially important compounds as chlorogenic acid, dicaffeoylquinic acid isomers or characteristic glucaric acids.