Abstract: Over a complete annual growth cycle, leaves at four different growth and development stages were collected each month from three Eucalyptus globulus Labill. sample plants grown under the same site conditions. Eucalyptus oil was extracted from the leaves by steam distillation and relative content of 1,8-cineole was measured and calculated by gas chromatography-mass spectrometry (GC-MS) to explore the accumulation of eucalyptus oil and 1,8-cineole in leaves with progressing growth and alternation of leaf harvesting season in the natural state, which may guide efficient cultivation and harvest management of economically valuable E. globulus forests. Results showed that oil and 1,8-cineole content differed significantly in leaves at different growth and development stages. Oil content in falcate lanceolate normal leaves (L) was significantly higher than that in leaves at the other three growth stages. The relative content of 1,8-cineole was significantly lower in broad and sessile young leaves (N) on the shoot tips and non-lignified stem segments than in leaves at the other three growth stages. The L stage leaves were a suitable leaf type for extracting eucalyptus oil and 1,8-cineole. Leaf oil content and relative 1,8-cineole content were significantly different between different sampling months. July to October was a suitable leaf harvesting period for the extraction of eucalyptus oil and 1,8-cineole. Oil content and relative 1,8-cineole content were significantly positively correlated with soil water content. Thus, soil water content may be a regulatory factor for efficient field cultivation and management and should be controlled at 16.1% – 26.1%.