Standardization of zygotic embryo culture from Nerium oleander L. and comparative analysis of biosynthesized cardiac glycosides within in vitro and acclimatized plants

The primary result of our experiment revealed that the germination percentage of N. oleander mature seeds is only 30%. From this observation, the concept of protocol standardization for zygotic embryo culture of this plant was originated. Zygotic embryo culture was proved an efficient in vitro multiplication system of N. oleander. The maximum germination percentage (96%) of zygotic embryos was observed on ¼ MS medium with 15 gm/L sucrose, whereas the best growth medium was optimized as ½ B5 with same sucrose concentration. The second part of this study was aimed to find out the cardiac glycoside accumulation pattern in both in vitro and acclimatized plants. For this purpose, one-month-old in vitro plantlets and acclimatized plants were subjected to LC-MS analysis and 09 cardiac glycosides were detected and quantified in both the systems. Most of the cardiac glycosides including odoroside A (32.71 mg/gm DW), odoroside H (4.69 mg/gm DW) and oleandrin (0.52 mg/gm DW) were found to be accumulated at maximum level within in vitro plantlets. CG 840b (1.89 mg/gm DW) is the only cardiac glycoside, which was maximally accumulated in acclimatized plants. From this study, it can be concluded that, zygotic embryo culture is a better choice for in vitro multiplication of N. oleander when compared to matured seeds and in vitro grown plantlets of this species favor cardiac glycosides biosynthesis in comparison to acclimatized plants. Therefore, all future research on the enrichment of cardiac glycosides from this plant may be conducted on zygotic embryos derived in vitro grown plantlets or cultures.