Thymus moroderi Pau ex Martínez (Lamiaceae) is a species endemic to South-eastern Spain valuable for its use in traditional medicine and in the production of a liqueur called cantueso. We describe a rapid and reliable in vitro propagation protocol for this species, starting from wild adult vegetative explants. Sterilization and establishment of in vitro culture were shown to be extremely difficult because of the high presence of contamination and the inability of non-contaminated explants to restore growth. In just two cases we were able to obtain non-contaminated explants with ability to grow: combining a fungicide dip with the presence of kanamycin in culture medium, and when mother plants were maintained for a week in a growth chamber with occasional fungicide spray. Once established in vitro, explants showed rapid and vigorous growth with spontaneous rooting. The presence of cytokinin alone or in combination with auxin gave worse results than untreated controls, thus revealing Thymus moroderi as a cytokinin-sensitive species. Better multiplication rates were however obtained when double phase system was included in the protocol. Fully acclimatized micropropagated plants were phenotypically indistinguishable from their wild relatives. Essential oils of micropropagated and wild plants were analyzed revealing a slightly different overall terpene profile. Micropropagated plants were abundant in monoterpene and sesquiterpene hydrocarbons but less in oxygenated sesquiterpenes with respect to wild plants. Major compounds were in both cases 1,8-cineole and camphor, although at different relative abundances. Despite these dissimilarities micropropagated plants produced a clearly recognizable and characteristic thyme odor.

The polyamine (PA) content of in vitro-grown explants of Thymus moroderi Pau ex Martínez has been evaluated during minimal growth storage. The growth restriction was imposed by the combined action of osmotically-active compounds (15 g dm -3 sucrose and 15 g dm -3 mannitol) present in the Murashige and Skoog medium and the modification of the physical environment of the culture (4 ºC and darkness). In these conditions, cultures were maintained up to 29 weeks without subculture. During this storage period, we analyzed contents of free, perchloric acid (PCA)-soluble and PCA-insoluble conjugated PA. Minimal growth storage brought about an increase in free putrescine (Put) coinciding with a reduction in PCA-soluble conjugated Put occurring during the first weeks of storage. PCA-insoluble conjugated spermidine (Spd) also accumulated in response to storage.