The greater wax moth Galleria mellonella L. is a serious destructive pest of honeybee colonies in the beekeeping industry all over the world. The larvae feed on almost all of the honeybee products and have the ability to completely destroy the bee colony, especially the weakened ones. The toxicity and biochemical effect of garden cress, Lepidium sativum seed methanolic extract as natural product against greater wax moth was evaluated under laboratory conditions. The defatted seeds (250g) of garden cress were refluxed in 500 ml methanol, filtered and then dry in a rotary evaporator. Results revealed that the toxic effect of the garden cress extract against 5th instar larvae of G. mellonella was 1.15%, (LC50) after 24h. which induced more histopathological alterations on cuticle layers, fat bodies and muscle. Moreover, it causes highly significant inhibition in α, β esterase and phenol oxidase activities as well as possesses a significant reduction in protein content. On other hand, it causes an elevation in total lipid content and acid phosphatase activity. Also, apoptotic activity with DNA fragmentation was occurred at concentrations of 3, 2 and 1%. GC-MS analysis illustrated the toxic effect of methanol extract of L. sativum seeds related to a high ratio of phenolic, flavonoid and fatty acids constituents.