Abstract The Amazonian tree Aniba rosaeodora Ducke, which provides a valued essential oil for perfume industry, is at risk of extinction. An alternative source of this product would be the oil obtained by steam distillation of the leaves of the same plant—which does not involve sacrifice of the tree. However, there is still not a technical criterion to ensure that determined oil is result of a sustainable production process. One step towards defining to know the differences in the oil compositions of trees of different stages of growth to establish the age from which the tree can be commercially explored and to differentiate products obtained from cultivated young plants and from native trees. In this paper, the characterization an differentiation of the essential oil extracted from the leaves collected from trees with different ages (4, 10 and 20 years old) was performed by Comprehensive Two-Dimensional Gas Chromatography coupled with Quadrupole Mass Spectrometric detection (GC × GC–qMS). GC × GC–qMS allowed the identification of ca. three times more chemical compounds on these samples, when compared to conventional gas chromatography. Depending on the age of the tree used to produce the oil, few differences in minor constituents of the oil samples were found; the amounts of the major compounds are similar all samples. Reliable differentiation of the essential oils according to the age of the source was only possible by GC × GC–qMS.