Abstract  Tissue cultured plants’ vessel headspace is subject to changes during subculture, and the analysis of its variation offers a non-destructive approach for monitoring plant physiology. Among the volatile organic compounds (VOCs) that can be released by plants and be potentially recovered in the airspace of plant tissue cultures, terpenes are very important since they can offer a snapshot of the physiological status of the plant under in vitro cultivation. Terpenes are synthesized from carbon directly shunted from the photosynthetic carbon fixation cycle and their emission is under genetic and environmental control. The experiments described in this paper propose the evaluation of the plant terpene profile in the culture’s headspace as an early indicator of plant stress through the characterization of plant terpene production. Monitoring of terpene emission as a plant response to mechanical stress such as plant wounding showed an increased isoprene and monoterpene emission rates in the first hour after cutting. The comparison of headspace composition of cultures of two fruit rootstocks, Colt and GF677, showed the former having higher emissions of isoprene, α-pinene and limonene than the latter. A decreasing emission trend was observed during subculture, apparently as a result of culture aging. The in vitro headspace analysis of different myrtle (Myrtus communis L.) clones showed a specific and distinctive terpene emission profile. VOC monitoring of in vitro culture headspace is discussed as a non-destructive approach useful for its relation with major physiological activities of culture and for the determination of the potential production of terpenes.