Poor storability of recalcitrant seeds is due to their inability to tolerate low moisture content. Understanding the processes underlying their recalcitrance is a prerequisite to developing a maintenance strategy and prolonging their lifespan. Multiple studies have investigated the differences between orthodox (desiccation-tolerant) and recalcitrant (desiccation-sensitive) seeds. Information on epigenetic regulation, however, is lacking and thus limits our understanding of the processes defining the physiology of seeds. In the present comparative study, changes in the global levels of 5-methylcytosine (m5C) in orthodox and recalcitrant seeds of Acer platanoides L. and Acer pseudoplatanus L. were characterized during progressive stages of severe drying. Concomitant with their differential sensitivity to desiccation stress, we demonstrate variation in the response of embryonic axes and cotyledons to water deficit at the level of DNA methylation. Results indicate that desiccation-induced changes in m5C are both tissue- and seed category-specific and are highly correlated with recalcitrant seed viability. Moreover, we demonstrate that m5C global changes in response to desiccation are not retained in DNA isolated from seedlings, except in seedlings that are derived from strongly desiccated orthodox seeds (moisture content of 3.5%). Finally, the potential utilization of m5C status as a universal seed viability marker is discussed.