Studying the mechanism underlying tooth eruption is an important and promising area due to the increased incidence of the diseases associated with tooth eruption abnormalities or difficulties that can result in purulent and septic complications. The study was aimed to explore variability and structural features of the hard tissue mineral component and organic matrix in mandibular third molars being at different stages of tooth eruption. Microscopic examination and biochemical testing of the enamel, dentin, and dentin–enamel junction of the study participants’ (n = 67; females aged 14–36) mandibular third molars were performed by scanning electron microscopy and Fourier transform infrared (FT-IR) spectroscopy. The association of the tooth eruption stage with the hard tissue structural features, such as the degree of mineralization and the size of dentinal tubules, orientation and size of the enamel prisms, was revealed. There were significant differences in the mandibular third molar hard tissue water content, which was demonstrated by metabolic processes and maturation rate (p < 0.05). According to the IR spectroscopy data, intensity of the collagen absorption bands in the enamel increases with age, while in dentin it decreases (1202, 1249, and 1342 cm^–1). Furthermore the combination of the reduced intensity of the 1202 cm^–1 band with the increase in the 1342 cm^–1 dentin–enamel junction band confirms the important role it plays as a link between the enamel and dentin due to its metabolic, shock-absorbing, protective, and nutritional functions. The findings demonstrate significant changes in the wrapping and orientation of the collagen fibrils and fibers in the hard tissue, which affect primary spatial orientation and mandibular dental topography.