This study investigates the mosquitocidal potential of crude petroleum ether extract fractions derived from Gloriosa superba against the dengue mosquito vector Aedes aegypti. Chemical profiling through gas chromatography identified four major metabolites, with nonadecane showing the highest peak area percentage (70.34%). A lethal dosage of fraction IV of G. superba (Gs‐F4), at a maximum concentration of 5.0 ppm, exhibited high mortality rates in second (78%), third (71%) and fourth (51%) instars of Ae. aegypti. A sublethal dosage of Gs‐F4 (3.0 ppm) significantly altered the larval midgut enzymes of cytochrome P450, esterases and glutathione S‐transferases. The repellent activity of Gs‐F4 was dosage dependent, with repellency percentages decreasing from 96% to 50% as the exposure time increased from 15 to 210 mins. The impact of Gs‐F4 on aquatic mosquito predators was marginally toxic (<50%) compared with the commercial compound temephos (1.0 ppm), which exhibited a higher toxicity (>50%). In silico toxicity screening using the BeeToxAI server indicated that three of the four compounds in Gs‐F4 were safe or non‐toxic to honeybees, except for α‐gurjunene (6.29 μg/bee). Although G. superba demonstrates effective larvicidal and repellent activity against Ae. aegypti, the non‐target toxicity against mosquito predators necessitates further toxicological screening on beneficial species under field conditions.