As the concentrated discharge outlet of an aquifer or groundwater system, a karst spring is partly independent from the aquifer, due to its formation of a pool or lake after outcropping onto the surface. Understanding how to evaluate the unique and sensitive environment of the karst spring is essential for water resource protection. Five karst springs in South China were investigated by analyzing their hydrodynamic conditions, variations in physicochemical parameters, and phytoplankton community structures. Dominated by regional or local groundwater flow, these springs had different catchment area characteristics and hydrogeological conditions. The results showed that, although they had similar water quality, their physicochemical parameters needed to be distinguished and evaluated in different ways in order to determine the cause of the observed degradation in spring water quality. Ca2+, HCO3-, and specific electrical conductivity were the major parameters reflecting the impact of regional flow from the aquifer; pH, dissolved oxygen, and water temperature indicated the local environment in and around the springs; while nitrogen and CODMn both related to the aquifer and local environment, depending on seasonal variation and human activities. The comparison of long-term nitrate data revealed that environmental pressure has increased over time. Water deterioration of Lingshui Spring was attributed to the strong interaction of surface water and groundwater. High nutrient concentrations did not correspond with the highest phytoplankton abundance or the most species. The phytoplankton community structures in the karst springs varied from place to place, depending on the hydrogeological conditions and the surrounding environment. The water environment status, as reflected by the combination of water quality indices and biological indicators, could more comprehensively represent overall water health.