High-level production of antibacterial pentacyclic triterpenoids from hairy root cultures of Bixa orellana via abiotic elicitation

Bixa orellana L ., a South American plant valued for its medicinal and culinary uses, was studied using hairy root cultures to isolate inducible bioactive triterpenoids. Hairy root cultures were elicited for 192 h with a combination of methyl- β -cyclodextrin, hydrogen peroxide, magnesium chloride, and methyl jasmonate. This study is the first to report the presence of 2 α ,6 β -dihydroxybetulinic and 6 β -hydroxybetulinic acid in B. orellana along with five known pentacyclic triterpenoids: arjunolic acid, maslinic acid, oleanolic acid, betulinic acid, and alphitolic acid. Elicitation increased the yield of these triterpenoids to 71.5 mg/g root dry weight, with a 36 % further increase upon re-elicitation. Semi-synthetic acetylated derivatives of arjunolic and maslinic acids were also synthesized. Maslinic acid exhibited potent antibacterial activity against vancomycin-resistant Enterococcus faecalis and methicillin-resistant Staphylococcus aureus , with MIC values of 6.25 and 12.5 µg/mL, respectively. The acetylated derivative of arjunolic acid showed strong anti-inflammatory effects at 20 µM concentration. This demonstrates the potential of B. orellana hairy roots as a sustainable platform for producing bioactive triterpenoids. • First report of 2 α ,6 β -dihydroxybetulinic and 6 β -hydroxybetulinic acids in B. orellana . • Combination of abiotic elicitors gave highest triterpenoid yield (71.5 mg/g). • Re-elicitation increased total triterpenoid yield by an additional 36 %. • Maslinic acid showed strong antibacterial activity against MRSA and VRE strains. • Acetylated arjunolic and maslinic acids showed enhanced anti-inflammatory effects.