Comparative chemical profiling of callus and seeds of Theobroma cacao by precolumn derivatization GC-MS and LC-MS

Theobroma cacao L. is a vital tropical economic crop. The seeds of cacao serve as core raw material for chocolate industry. However, conventional breeding for cacao production was challenged by long-term constraints from pests, diseases, climate changes, and soil pollution. Plant tissue culture offers a viable approach for cacao variety improvement, resource conservation and metabolism regulation research. In the present study, cacao callus was successfully induced from hypocotyls of aseptically germinated cacao seeds. The obtained cacao callus was maintained on Murashige & Skoog medium supplemented with 0.1 mg/L 1-naphthaleneacetic acid and 2 mg/L 6-benzylaminopurine for rapid and stable cacao callus proliferation. Subsequently, precolumn derivatization GC-MS and LC-MS coupled with multivariate statistical analysis were employed to conduct metabolomic profiling of cacao callus and cacao seeds. A total of 182 metabolites were annotated, including amino acids, organic acids, sugars, flavonoids, alkaloids and others. All the annotated metabolites were detected in both cacao callus and cacao beans but with their concentrations varied. The established cacao tissue culture system could not only provide stable materials for investigating cacao metabolic regulation mechanisms but also facilitate the preservation, rejuvenation, and genetic transformation of elite germplasm which would be helpful for the healthy development of cacao industry.