Abstract: In order to partition water conservation areas in the upper Yangtze River, inner and external indexes in an ecosystem were used to constructed a delimitation index system, and the key water conservation areas were delimitated by the delimitation model in combination with the ArcGIS Spatial Analyst. Based on the actual situation of research areas, a multi-level and multi-structure index system for eco-security assessment was built with the PSR (pressure-state-response) model framework. Eco-security assessment of the key water conservation areas was carried out by a combined application of AHP (analytic hierarchy process), vector similarity and principal component projection method. Gray correlation was added in finding out the main influencing factors on regional eco-security. The results show that these main factors are rainfall, increase rate of GDP, effective utilization rate of water resources, vegetation coverage, and water environmental quality.