Integrative metabolomic and transcriptomic analysis reveals CYP450-driven biosynthesis of pentacyclic triterpene acids in Cecropia angustifolia

Pentacyclic triterpene acids (PTAs) are specialized plant metabolites known for their anti-inflammatory and antidiabetic preclinical reports. Cecropia angustifolia, a prominent species in the Andean montane forests, produces a variety of PTAs in its roots, but little is known about enzymes responsible for their oxidative structural decorations. In this study, we explored the biosynthetic route of PTAs oxidation by cytochrome P450 (P450s) enzymes, comparing wild-collected and in-vitro cultivated root tissues. Chemical profiling was carried out using targeted metabolomics to assess PTAs in both types of extracts. Oxidation patterns differed between the samples, suggesting distinct enzyme activity depending on growth conditions. To support these findings, we assembled a de novo transcriptome based on publicly available Cecropia RNA-seq data and predicted over 26,000 open reading frames. Four P450 families (CYP716A, CYP716C, CYP72A, and CYP71D) are potential contributors to triterpene oxidation, which were identified by homology-based annotation. Gene expression was examined by RT-qPCR, revealing differential transcriptional activity between wild and in-vitro roots. Nevertheless, these differences link the gene expression profile of P450s involved in this pathway to the TPAs metabolic expression. Notably, despite changes in expression, the capacity to generate oxidized PTAs was retained under both conditions. Our results indicate that the oxidative metabolism of triterpenes in C. angustifolia is largely conserved and responsive to environmental cues. This study enhances the current understanding of PTA biosynthesis and supports the use of in-vitro systems as a practical strategy to obtain valuable metabolites while minimizing ecological impact.