BACKGROUND: As one of the most abundant and destructive pests in agriculture, aphids cause significant damage to crops due to their sap-taking and as virus vectors. Chemical insecticides are the most effective method to control aphids, but they bring insecticide resistance problems and harm nontarget organisms, especially bees, therefore the search for novel eco-friendly aphid control agents with low bee toxicity is urgent. Insect kinins are a class of small neuropeptides that control important functions in insects. In our previous study, we found insect kinin analog IV-3 has good aphicidal activity and the location of the aromatic ring on the side chain of Phe 2 is the key to the formation of the ⊎-turn resulting in the biological activity of insect kinin analogs. However, there are few studies on insect kinin Phe 2 substitution and modification, and its structure-activity relationship is still unclear.RESULTS: In this project, 44 insect kinin analogs with the Phe 2 modification, replacing it with different natural or unnatural amino acids, were designed and synthesized based on the lead IV-3 to explore the role of the Phe 2 residues. Bioassays with soybean aphids of Aphis glycines indicated that nine analogs have better aphicidal activity than the lead IV-3. In particular, compound L 25 exhibits excellent aphicidal activity (LC 50 = 0.0047 mmol L −1 ) and has low toxicity to bees. Furthermore, a reliable three-dimensional quantitative structure-activity relationship (3D-QSAR) was established to produce a helpful clue that introducing hydrophobic groups away from the backbone chain is beneficial to improve aphicidal activity.CONCLUSION: The residue Phe 2 of insect kinin analogs is the key position and has a significant impact on the activity. L 25 has a high toxicity for aphids, while a low toxicity to bees, and therefore can be considered as a lead compound to develop new biosafe aphid control agents. Finally, we provide a useful 3D-QSAR model as theoretical guidance for further structural optimization.