Aconitum heterophyllum is a high altitude medicinal plant that has become endangered due to overexploitation for their aconitins. The most effective, conventional propagation method for any plant species is by seed. However, in Aconitum seed germination is erratic, and seedling survival is low. In the present study results have been discussed on the possible implication of ethanol treatment on removal of barriers on radical emergence in terms of protein changes. Eighty seven percent of seed germination was achieved in Aconitum with ethanol treatment. Comparative 2-DE analysis of ethanol treated and untreated seed protein profiles in Phase II of germination revealed 40 differentially expressed proteins. Twenty-seven out of 40 proteins were induced, 5 were increased and 8 were repressed. Mass spectrometry and subsequent identification confirmed that these proteins were involved in metabolism, DNA regulation, stress tolerance and plasmamembrane/cell wall biosynthesis/extension processes. These protein changes might be responsible for physiological and physical changes, respectively, resulted in increase in germination percentage. Further, characterization of these proteins will be of great help in understanding the molecular mechanism lying behind enhanced germination in response to ethanol treatment.