In recent years, insecticide resistance has reached alarming levels, requiring the development of alternatives for vector control. This study investigated the phytochemical composition by high-performance liquid chromatography with ultraviolet detection (HPLC/UV), total polyphenolic content (TPC), antioxidant activity (evaluated using 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) assays), and larvicidal activity of leaf extracts from Lavandula stoechas (A), Urtica pilulifera (B), and Cupressus sempervirens (C), tested individually and in ternary combinations (D, E, F) against fourth-instar larvae of Culiseta longiareolata (Diptera: Culicidae) through contact exposure bioassays. The results showed that the highest TPC was recorded in extract C, reaching 85.92 ± 0.26 mg GAE g−1. Furthermore, mixture F exhibited the greatest polyphenol content (52.30 ± 0.184 mg GAE g−1). In antioxidant assays, the findings verified that extract C demonstrated the strongest radical scavenging activity, with half inhibitory concentration (IC50)of 45.00 ± 0.01 mg ml−1 for the DPPH assay and 28.20 ± 0.21 mg ml−1 for the ABTS assay. Similarly, mixture F was the most effective among the combinations, with IC50values of 97.00 ± 0.08 mg ml−1 and 31.00 ± 0.05 mg ml−1 for the DPPH and ABTS assays, respectively. Moreover, HPLC profiling revealed the presence of key flavonoids, and prominent phenolic acid compounds, where orcinol hydrate, 4‑hydroxybenzoic acid, and caffeic acid were detected exclusively in mixtures D and E, but were absent or only trace amounts in the single extracts. Larvicidal activity, assessed by the median lethal concentration (LC50) and the median lethal time(LT50), showed that mixtures D, E, and F exhibited strong larvicidal effects with low LC50 values of 2.210, 1.889, and 2.705 mg ml−1 at 72 h, respectively, and LT50 values of 29.74, 17.08, and 39.86 h, respectively, confirming their higher toxicity. Moreover, synergistic effects occurred in all mixtures (16 to 72 h) except for mixture F at 16 h. The docking analysis revealed favorable binding affinities and stable interactions between the tested compounds and the active site of acetylcholinesterase (AChE), highlighting their potential as effective larvicidal agents. These findings imply that the synergistic interactions were more effective on tested insects than the individual plants. Consequently, such formulations hold significant potential as alternative agents for mosquito vector control.

Phytochemical and Synergistic Insecticidal Potential of Three Medicinal Plants Against Culiseta longiareolata: In Vivo/In Silico Investigations

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发表日期：

2026年3月6日

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Amel Hammoudi,
Karim Arab,
Ouahiba Bouchenak,
Widad Hadjab,
Razika Laoufi,
Khalid Rezig

摘要

In recent years, insecticide resistance has reached alarming levels, requiring the development of alternatives for vector control. This study investigated the phytochemical composition by high-performance liquid chromatography with ultraviolet detection (HPLC/UV), total polyphenolic content (TPC), antioxidant activity (evaluated using 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) assays), and larvicidal activity of leaf extracts from Lavandula stoechas (A), Urtica pilulifera (B), and Cupressus sempervirens (C), tested individually and in ternary combinations (D, E, F) against fourth-instar larvae of Culiseta longiareolata (Diptera: Culicidae) through contact exposure bioassays. The results showed that the highest TPC was recorded in extract C, reaching 85.92 ± 0.26 mg GAE g⁻¹. Furthermore, mixture F exhibited the greatest polyphenol content (52.30 ± 0.184 mg GAE g⁻¹). In antioxidant assays, the findings verified that extract C demonstrated the strongest radical scavenging activity, with half inhibitory concentration (IC₅₀)of 45.00 ± 0.01 mg ml⁻¹ for the DPPH assay and 28.20 ± 0.21 mg ml⁻¹ for the ABTS assay. Similarly, mixture F was the most effective among the combinations, with IC₅₀values of 97.00 ± 0.08 mg ml⁻¹ and 31.00 ± 0.05 mg ml⁻¹ for the DPPH and ABTS assays, respectively. Moreover, HPLC profiling revealed the presence of key flavonoids, and prominent phenolic acid compounds, where orcinol hydrate, 4‑hydroxybenzoic acid, and caffeic acid were detected exclusively in mixtures D and E, but were absent or only trace amounts in the single extracts. Larvicidal activity, assessed by the median lethal concentration (LC₅₀) and the median lethal time(LT₅₀)_, showed that mixtures D, E, and F exhibited strong larvicidal effects with low LC₅₀ values of 2.210, 1.889, and 2.705 mg ml⁻¹ at 72 h, respectively, and LT₅₀ values of 29.74, 17.08, and 39.86 h, respectively, confirming their higher toxicity. Moreover, synergistic effects occurred in all mixtures (16 to 72 h) except for mixture F at 16 h. The docking analysis revealed favorable binding affinities and stable interactions between the tested compounds and the active site of acetylcholinesterase (AChE), highlighting their potential as effective larvicidal agents. These findings imply that the synergistic interactions were more effective on tested insects than the individual plants. Consequently, such formulations hold significant potential as alternative agents for mosquito vector control.