Pheromone communication is an essential component of reproductive isolation in animals. Spodoptera littoralis has evolved its own repertoire of olfactory receptors (Ors) to satisfy its ecology, essentially through duplication events. Duplicates vary significantly in sequence identity and tuning breadths. However, how the duplicates diverged and acquired new functions is not known. Previous functional studies showed that SlitOR5 is specifically tuned to (Z,E)-9,11-14:OAc (Bastin-Héline et al., 2019), while SlitOR75, a recent duplicate of SlitOR5, can detect many compounds. Here, we used ancestral gene reconstruction using the codeml of the PAML suite and heterologous expression in drosophila neurons to resurrect AncOR5_75, their common ancestor, as well as AncOR5. Then, the 3D structure and binding pockets were predicted by AlphaFold2 and Deepsite, respectively. At last, ancestors were functionally characterized using Xenopus oocyte heterologous expression system, and site-directed mutagenesis experiments were performed on AncOR5_75 to recreate the evolutionary trajectory that led to SlitOR5 (Li & Capoduro et al., 2023).