Profiles of Semi-Polar Metabolites from Leaves of In Vitro- Derived Plants of Indonesian Pepper Varieties (Capsicum annuum) after Gamma Irradiation Treatments

Abstract

Gamma irradiation has been widely utilized for mutation breeding as it induces random mutations in plant cells. Due to limitations on other breeding approaches, including cross-breeding and transgenic plants, gamma irradiation-induced mutation breeding has regained its popularity among breeders and scientists. Untargeted metabolomics analysis can be used to profile any perturbation between untreated samples and treated sample groups without having prior knowledge of particular metabolites, including semi-polar metabolites, which cover phenolic acids, flavonoids, glycosylated steroids, alkaloids, and other glycosylated species. This study aimed to profile semi-polar metabolite modification in leaves of plants derived from in vitro cultures after gamma irradiation treatments in Indonesian chili pepper varieties. Seeds of Laris and Kopay were exposed to gamma-ray doses (0, 100, 200, 300, 400, and 500 Gy), after which they were germinated and cultivated in vitro. Plantlets were acclimated in the greenhouse and leaves were collected for metabolite analysis at 80 days after the germination of irradiated seeds. Semi-polar metabolites from leaves were extracted using methanol and the extracts were subjected to LC-MS analysis. Results showed that the composition and levels of semi-polar metabolites of Laris 100, 200, 300, 400, and 500 Gy were similar to Laris 0 Gy (control; unirradiated seeds), although several abundancies of that of 200 Gy gamma irradiation dose were different than control. Except at dose 400 Gy, Kopay at 0 Gy, 200 Gy, and 300 Gy also showed similar metabolite profiles, indicating that gamma-ray doses did not induce mutation at genes regulating metabolite biosynthetic pathways. In contrast, Kopay 400 Gy showed low levels of terpenoids and flavonoids, indicating that a 400 Gy dose of gamma ray may affect the upstream part of the shikimate biosynthetic pathway, resulting in low levels of precursors at the upstream biosynthetic pathway of terpenoids and flavonoids. Hence, the accumulation of terpenoids and flavonoids was very low. These findings provide insights into the effect of gamma irradiation for mutation breeding that may be important for future pepper breeding programs.