Background: The destructive insect pest Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae) is a polyphagous species targeting many economically important plants. The extensive and arbitrary use of insecticides has resulted in the build-up of insecticide resistance and pesticide residues accumulating in food. Therefore, it is becoming evident that alternative pest management tools are needed to reduce risks to humans, the environment, and non-target organisms, and at the same time, they should be used in field application at the lowest cost. Methods: In view of this objective, the present study demonstrates the toxicity of lemongrass (Cymbopogon citratus (DC.) Stapf) essential oil (EO), against the black cutworm A. ipsilon under controlled laboratory conditions in terms of measuring the activity of peroxidase and detoxification enzymes. The chemical components of the EO were analyzed using GC–MS. Results: The results show that after 96 h post treatment, the LC₁₅ and LC₅₀ values were 427.67 and 2623.06 mg/L, respectively, of C. citratus EO on second-instar larvae of A. ipsilon. A slight significance in elongation of the larval duration with LC₁₅ and LC₅₀ value was found with control. By GC–MS analysis, the main compounds identified in the EO were α-citral and β-citral with percentages of 35.91%, and 35%, respectively. The oxidative stress indicates a significant increase in CAT and lipid peroxidase enzyme activity after 96 h post treatment at the LC₁₅ and LC₅₀. Conversely, the detoxification enzyme activity shows an inhibition of CarE and GST enzymes of larvae exposed to LC₁₅ and LC₅₀ values in response to C. citratus EO. Conclusions: The present data show that lemongrass EO has insecticidal activity against the black cutworm, A. ipsilon.