Design and elucidation of an insecticide from natural compounds targeting mitochondrial proteins of Aedes aegypti

Developing new pesticides poses a significant challenge in designing next-generation natural insecticides that selectively target specific pharmacological sites while ensuring environmental friendliness. In this study, we aimed to address this challenge by formulating novel natural pesticides derived from secondary plant metabolites, which exhibited potent insecticide activity. Additionally, we tested their effect on mitochondrial enzyme activity and the proteomic profile of Ae. aegypti, a mosquito species responsible for transmitting diseases. Initially, 110 key compounds from essential oils were selected that have been reported with insecticidal properties; then, to ensure safety for mammals were performed in silico analyses for toxicity properties, identifying non-toxic candidates for further investigation. Subsequently, in vivo tests were conducted using these non-toxic compounds, focusing on the mosquito's larval stage. Based on the lethal concentration (LC), the most promising compounds as insecticidal were identified as S-limonene (LC₅₀ = 6.4 ppm, LC₉₅ = 17.2 ppm), R-limonene (LC₅₀ = 9.86 ppm, LC₉₅ = 27.7 ppm), citronellal (LC₅₀ = 40.5 ppm, LC₉₅ = 68.6 ppm), R-carvone (LC₅₀ = 61.4 ppm, LC₉₅ = 121 ppm), and S-carvone (LC₅₀ = 62.5 ppm, LC₉₅ = 114 ppm). Furthermore, we formulated a mixture of R-limonene, S-carvone, and citronellal with equal proportions of each compound based on their LC₅₀. This mixture specifically targeted mitochondrial proteins and demonstrated a higher effect that showed by each compound separately, enhancing the insecticidal activity of each compound. Besides, the proteomic profile revealed the alteration in proteins involved in proliferation processes and detoxification mechanisms in Ae. aegypti. In summary, our study presents a formulation strategy for developing next-generation natural insecticides using secondary plant metabolites with the potential for reducing the adverse effects on humans and the development of chemical resistance in insects. Our findings also highlight the proteomic alteration induced by the formulated insecticide, showing insight into the mechanisms of action and potential targets for further exploration in vector control strategies.