3-O-trans-p-coumaroyl esterification enhances the anti-inflammatory effects of tormentic acid by targeting NF-κB signaling

Tormentic acid (TA), a plant-derived pentacyclic triterpene, exhibits antioxidant and anti-inflammatory potential, yet the pharmacological effects of its 3-O-trans-p-coumaroyl ester (trans-TACE) remain underexplored. This study investigates how hydroxycinnamoyl esterification influences the biological activity of pentacyclic triterpenes by comparing TA and trans-TACE in cellular and in vivo stress models.

We assessed their ability to mitigate oxidative stress by evaluating the inhibition of ROS and NO molecules. Pro-inflammatory cytokine production in LPS-stimulated THP-1 macrophages was analyzed through cytokine arrays and multiplex immunoassays, while NF-κB activation was examined in both TLR4-dependent and -independent models using HEK-Blue reporter cells. Uptake efficiencies into Caco-2 enterocytes were measured via LC-MS. The in vivo relevance of these findings was assessed using C. elegans as a model for oxidative and inflammatory stress response.

Results showed that trans-TACE significantly reduced cellular ROS and NO levels compared to TA. Protein analyses of LPS-stimulated THP-1 macrophages indicated that trans-TACE significantly decreased pro-inflammatory cytokines involved in NF-κB signaling (e.g., TNFα, IL-8, CCL2, CXCL5 and CXCL11). Trans-TACE also inhibited NF-κB activation in both TLR4-dependent and -independent models. In C. elegans, both TA and trans-TACE downregulated several stress-induced genes, with trans-TACE exhibiting broader effects by additionally targeting daf-16 and gst-4 gene expression. Moreover, we revealed key differences in bioactivities between the trans and cis isoform of TACE, underscoring the importance of considering the structural properties of geometric isomers in therapeutic assessments.

Overall, this study suggests that esterification significantly enhances the biological activity of pentacyclic triterpenes and points towards new possibilities for developing effective natural anti-inflammatory therapies.