Attraction of virgin female Anastrepha serpentine Wiedemann (Diptera: Tephritidae) to the odor of calling males was demonstrated. This sex pheromone-mediated attraction occurred during the latter half of a 13-h photophase but not during the first half of the day. Two major components of emissions of calling males, 2,5-dimethylpyrazine (DMP) and 3,6-dihydro-2,5-dimethylpyrazine (DHDMP), and trimethylpyrazine (TMP), a minor component, were tested for pheromonal activity. DMP and TMP elicited antennal responses using electroantennogram assays, but no response was observed for DHDMP. DMP and combinations of DMP and TMP were not attractive in laboratory bioassays. Bioassays of DHDMP were complicated by its instability making purification of the compound unfeasible. Bioassays of DHDMP in unpurified form were further complicated by the presence of reaction byproducts and unreacted reagent in crude preparations of the DHDMP. However, statistical analysis indicated that combinations of 100–500 ng of DHDMP in mixtures with DMP as the major component elicited attraction in laboratory bioassays with sexually active female flies. Taking into account all of the data, we could not conclude that any of the chemicals emitted by males are pheromones, but based on female fly responses to mixtures of DHDMP and DMP, this is the most likely scenario.