Plant regeneration and biochemical accumulation of hydroxybenzoic and hydroxycinnamic acid derivatives in Hypoxis hemerocallidea organ and callus cultures

Micropropagation of Hypoxis hemerocallidea Fisch. and C.A. Mey was used as a model system to study the influence of cytokinins (CKs) on plant regeneration and biochemical accumulation of hydroxybenzoic and hydroxycinnamic acid derivatives in organ and callus cultures and their antioxidant activity. Fourteen free phenolic acids were detected using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) while antioxidant activity was evaluated using oxygen radical absorbance capacity (ORAC) and 2,2-diphenyl-1-picryl hydrazyl (DPPH) radical scavenging activity. Cytokinins had a significant effect on the biochemical accumulation of hydroxybenzoic and hydroxycinnamic acid derivatives in H. hemerocallidea organ cultures. In particular, meta-topolin-treated organ cultures produced high concentrations of gallic, protocatechuic, gentisic, p-hydroxybenzoic, m-hydroxybenzoic, salicylic, chlorogenic and trans-cinnamic acids. The isoprenoid CK, N6-(2-isopentenyl)-adenine significantly increased the accumulation of hydroxycinnamic acid derivatives, namely, caffeic, p-coumaric, sinapic and ferulic acids. Cytokinin-treated organ cultures exhibited a significant increase in antioxidant activity, particularly in the ORAC model. In callus cultures, CKs decreased the concentrations of hydroxycinnamic acid derivatives and antioxidant activity when compared to the control. Overall, both CK type and concentration had a significant effect on plant regeneration, callus proliferation, biochemical accumulation of free phenolic acids and antioxidant activity of the resultant extracts.