Engineering phytochemical co-production through interspecific co-cultivation of Quillaja saponaria and Aristotelia chilensis in temporary immersion bioreactors (TIBs)

Sustainable production of high-value plant secondary metabolites requires bioprocesses that combine controlled biomass production with flexible modulation of phytochemical outputs. This study established an interspecific co-cultivation strategy for the simultaneous production of saponin-related metabolites (SRM) and phenylpropanoid-related metabolites (PRM) from two native Chilean woody species, Quillaja saponaria and Aristotelia chilensis , using temporary immersion bioreactors (TIBs). Biomass production was first optimized using 6-benzylaminopurine (BAP; 0–2.5 mg L⁻¹), followed by co-cultivation at three species ratios (1QS:1AC, 1QS:2AC, and 2QS:1AC) and elicitation with abscisic acid (ABA; 0, 1, and 2 mg L⁻¹) for 30 days. Biomass growth, SRM and PRM levels, and antioxidant capacity, assessed using the oxygen radical absorbance capacity (ORAC) assay, were evaluated in biomass and culture medium (SRM and PRM) and in combined biomass–culture medium samples (ORAC). The highest biomass accumulation was obtained at 1 mg L⁻¹ BAP, while co-culture biomass accumulation remained within the range observed for the individual cultures; however, species-specific growth within the co-cultures was not assessed. Species ratio influenced phytochemical output, with 1QS:2AC favoring PRM accumulation and antioxidant capacity, whereas 2QS:1AC favored SRM accumulation. ABA further modulated metabolite levels, with 1 mg L⁻¹ favoring PRM accumulation and 2 mg L⁻¹ accelerating SRM accumulation at intermediate sampling times, with comparable final levels at both concentrations. Notably, the 1QS:2AC treatment exhibited a progressive increase in ORAC activity from 10.2 to 15.9 µmol Trolox equivalents g⁻¹ fresh weight over 30 days, contrasting with the decline observed in the A. chilensis control. Overall, these findings support interspecific co-cultivation in TIBs as a tunable proof-of-concept platform for the simultaneous production of complementary plant-derived metabolite classes. They also suggest that species composition is associated with changes in phytochemical output and the temporal pattern of antioxidant capacity during cultivation.