Ilex paraguariensis (mate) is a species native to South America and is widely consumed in countries such Argentina, Uruguay, Paraguay, and Brazil. Mate consumption is associated with several phytotherapeutic functions, in addition to its cultural and regional importance. However, the harvest period can affect the properties of the mate, due to variations in the constituent proportions, as a consequence of seasonal changes. In this work, we employed nuclear magnetic resonance and chemometrics to evaluate the chemical variations in leaf extracts of I. paraguariensis over the four seasons of the year. We found significant changes in the levels of glucose, myo-inositol, caffeine, theobromine, and fatty acids. These changes can be related to resource allocation for the flowering period, or to responses to environmental stresses, such as temperature.