Median lethal concentrations (LC₅₀) on Ae. albopictus were 102.97, 85.40, and 91.38 ppm, while concentrations able to kill 90% of the exposed larvae (LC₉₀) were 122.4, 113.65, and 122.22 ppm for the EO, hexyl butyrate, and octyl acetate, respectively (Table 2).

Both hexyl butyrate and octyl acetate were more effective against this species than the EO (GLMM post hoc Bonferroni corrected—hexyl butyrate: OR = 0.101, SE = 0.037, z = −6.265, p < 0.0001; octyl acetate: OR = 0.161, SE = 0.059, z = −4.935, p < 0.0001) (Figure 1). Even if no significant difference between the two compounds was detected (OR = 1.588, SE = 0.300, z = 2.448, p = 0.129), the LC₅₀ and LC₉₀ were lower for hexyl butyrate if compared with octyl acetate (Figure 2).

LC₅₀ on Ae. aegypti was 59.09, 53.59, and 47.05 ppm, while the LC₉₀ was 101.62, 99.49, and 84.30 ppm for the EO, hexyl butyrate and octyl acetate, respectively (Table 2). Also in this case, hexyl butyrate and octyl acetate were more toxic than the EO (hexyl butyrate: OR = 0.357, SE = 0.128, z = −2.874, p = 0.036; octyl acetate: OR = 0.191, SE = 0.191, z = −4.555, p < 0.0001) (Figure 1). As above, no difference was detected between the two compounds for the mortality (OR = 0.534, SE = 0.251, z = −1.333, p = 1.000), but octyl acetate had lower LC₅₀ and LC₉₀ values with respect to hexyl butyrate (Figure 2).

LC₅₀ on An. gambiae was 97.91, 70.97, and 60.71 ppm, while the LC₉₀ was 116.02, 116.48, and 125.45 ppm for the EO, hexyl butyrate, and octyl acetate, respectively (Table 2). Again, both hexyl butyrate and octyl acetate were more effective against this species than the EO (hexyl butyrate: OR = 0.063, SE = 0.021, z = −8.116, p < 0.0001; octyl acetate: OR = 0.045, SE = 0.016, z = −8.449, p < 0.0001) (Figure 1), even if no significant difference was detected between these two compounds (OR = 0.709, SE = 0.151, z = −1.604, p = 0.978). Nevertheless, although the two compounds were found to be more effective than the EO, their efficacy is slower; even a slight increase in EO concentration leads to a higher mortality rate, whereas this is not as true for hexyl butyrate and octyl acetate. Thus, their LC₅₀ and LC₉₀ trends are significantly different from the EO ones (EO vs. hexyl butyrate: OR = 0.072, SE = 0.016, z = 4.372, p = 0.0001; EO vs. octyl acetate: OR = 0.092, SE = 0.016, z = 5.731, p < 0.0001) (Figure 2).

Usually, botanical products with a LC₅₀ lower than 100 ppm can be considered interesting for their application as insecticidal agents [45,46]. The EO herein investigated achieved LC₅₀ values lower than 100 ppm against An. gambiae and Ae. aegypti and slightly higher against Ae. albopictus. Interestingly, hexyl butyrate and octyl acetate showed significantly higher efficacy towards larvae of all mosquito species. Indeed, they could be mainly responsible for the effect of the EO. Considering the few reports regarding the mosquitocidal activity of EOs of the Heracleum genus, the presented results are of interest. Indeed, they could be useful to better assess the insecticidal potential of this Iranian plant. This study reports for the first time the larvicidal effects of H. persicum EO on Ae. aegypti, An. gambiae and Ae. albopictus. Previously, only Sedaghat et al. [23] assayed the EO from seeds of the same species on fourth instar larvae of An. stephensi, yielding comparable results to those reported in this study for mosquitoes of the Anopheles genus. The product showed moderate toxicity, with LC₅₀ and LC₉₀ values of 104.80 and 174.22 ppm, respectively, after 24 h of exposure. Unfortunately, the chemical composition of the EO has not been reported in that study. The insecticidal activity of the EO against agricultural pests has also been shown. Specifically, it showed toxicity against adults of C. maculatus and the sub-lethal doses of this EO reduced the longevity and fecundity of the beetle, also reducing female fertility by 21.2%, with a significant effect on the oviposition behavior [47]. Moreover, Manzoomi et al. [24] reported that the fumigant toxicity of this EO increased at increasing concentrations and exposure time (LC₅₀ of 337.58 μL/L) against the same species. Furthermore, H. persicum EO exhibited strong repellent effects also against T. castaneum. Other species of the genus Heracleum showed toxicity against mosquitoes. For instance, Baranová et al. [26] recently tested H. mantegazzianum EO against Ae. japonicus Theobald larvae. Its chemical composition was dominated by octyl acetate (58.65% of the total identified), which was also more efficient than the EO (LC₅₀ values of 67 mg/L and 52 mg/L for the EO and octyl acetate, respectively). Moreover, Govindarajan et al. [48] showed high toxicity of the Heracleum sprengelianum Wight & Arn. EO against larvae of Anopheles subpictus Grassi, a malaria vector, Ae. albopictus, and the Japanese encephalitis vector Culex tritaeniorhynchus Giles, obtaining LC₅₀ of 33.4, 37.5, and 40.9 mg/mL, respectively. The main EO components were lavandulyl acetate and bicyclogermacrene, which exhibited even lower larval toxicity. The LC₅₀ values ranged from 4.17 to 10.3 mg/mL against An. subpictus, 4.60 to 11.1 mg/mL against Ae. albopictus, and 5.11 to 12.5 mg/mL against C. tritaeniorhynchus. Finally, the acute toxicity of the EO obtained from H. sphondylium subsp. sphondylium and H. sphondylium subsp. ternatum also showed efficacy on third instar larvae of the filariasis vector Culex quinquefasciatus Say, reaching LC₅₀ values of 73.8 and 64.98 μL/L, respectively [44]. Based on the results herein obtained and those reported in the literature, it is worth continuing to investigate the EOs of the Heracleum genus for their exploitation as eco-friendly insecticides. Particularly, the mechanism(s) of action [18] and the non-target toxicity [6] of these botanical products should be assessed. Aliphatic esters already demonstrated to have a toxic behavior on the moth Depressaria radiella (Duponchel, 1838) which produces specific esterases involved in the detoxification system [49]. However, no data are available on the larvicidal mode of action of hexyl butyrate and octyl acetate. Nonetheless, the low LC₅₀ values obtained, especially on Ae. aegypti, highlight the possible role of these products in mosquito management. Additionally, H. persicum EO could be well-suited for industrial applications due to its widespread presence in Iran and Northern Europe, as well as its low price on the market (2–3 USD/kg).