Metabolite Profiling of Dayak Onion (Eleutherine bulbosa) Bulb and Callus Cultures under Picloram and Kinetin Treatments

Eleutherine bulbosa is a medicinal plant widely used in traditional medicine and known to contain diverse bioactive secondary metabolites with strong antioxidant activities. However, sustainable production of these compounds from natural sources is limited, and information on biosynthesis in in vitro systems remains poorly understood, particularly under different PGR conditions. Callus culture offers a controlled platform for metabolite production, but the chemical profile is strongly influenced by plant growth regulators (PGRs). Therefore, this study aimed to compare metabolite richness, assess the conservation of chemotypes between bulb and callus cultures of E. bulbosa, and evaluate the effects of different PGR combinations on metabolite composition and abundance. Bulb explants were surface-sterilized and cultured for 40 days on Murashige–Skoog (MS) medium supplemented with varying concentrations of picloram and kinetin. Dried callus (0.20 g) was extracted using acidified methanol, hydrolyzed with 2 N HCl, and reconstituted for analysis. Subsequently, metabolite profiling was performed using liquid chromatography–mass spectrometry (LC–MS). The results showed that a total of 120 secondary metabolites were detected in bulb extracts, while callus culture produced 89–105 metabolites depending on the PGR treatment. The highest metabolite richness was observed in culture treated with 3.00 ppm picloram and 0.25 ppm kinetin. Several metabolites identified in callus overlapped with those in the bulb, indicating partial conservation of key chemical signatures under in vitro conditions. These results demonstrate that appropriate PGR combinations can maintain broad metabolite diversity in callus culture and emphasize the potential of E. bulbosa callus as a sustainable alternative source of bioactive compounds.