The drastic changes in water regime are of great importance for maintaining water security in lakes with violent seasonal fluctuations. Based on hydrological data of Dongting Lake, the second largest freshwater lake in China, during 1980–2014, a set of water safety assessment methods suitable for seasonal lakes was constructed. Results demonstrated the following. (1) The year 2003 was a major time point for changes in water regime in Dongting Lake between 1980 and 2014. (2) The water regime changes in different parts of Dongting Lake exhibited certain similarities in periodicity and overall change trends. Among them, South Dongting Lake presented the best water security status, followed by East Dongting Lake and West Dongting Lake. (3) Compared with 1980–2002, the complexity of the water safety level change in East Dongting Lake increased in 2003–2014, as manifested in the tendency of the distribution of water safety levels to change sharply. (4) The influence of dry years on the water safety of Dongting Lake was greater than that of rainy years. The multi-time, multi-scale and multi-target lake water safety assessment method based on water regimes provides new ideas and methods for global research on seasonal lake water safety.