Spodoptera litura (F.) (Lepidoptera: Noctuidae) is a serious pest of several crops in India and other countries. Prothoracicotropic hormone (PTTH) release from the brain of penultimate fifth instars of the noctuid was followed in the laboratory and a spiracle index developed to assess PTTH action. PTTH release from the brain occurred during the scotophase at 4.30 h of day 1 (AZT). Ecdysone secreted in the prothoracic gland was released from the 7th hour of day 1 (AZT), reaching a maximum in the haemolymph on the 9th hour of day 1 (AZT). The prothoracic gland showed three distinct phases, namely growth, synthesis and release. Injected PTTH of 5th instar brain extract induced larval moulting in head-ligated larvae. Partially purified PTTH from 6th-instar S. litura brain induced moulting in 6th instars. Abdomen-ligated penultimate instars administered β-ecdysone showed variable moulting response. The phenomenon of hormonal interaction with development of the penultimate instar of S. litura is discussed.