Lithium is a critical element for the modern society due to its uses in various industrial sectors. Despite its unequal distribution in the environment, Li occurrence in Romania was scarcely studied. In this study a versatile measurement method using ICP-MS technique was optimized for the determination of Li from various matrixes. Water, soil, and plant samples were collected from two important karst areas in the Dobrogea and Banat regions, Romania. The Li content was analyzed together with other macro- and microelement contents to find the relationship between the concentration of elements and their effect on the plants’ Li uptake. In Dobrogea region, half of the studied waters had high Li concentration, ranging between 3.00 and 12.2 μg/L in the case of water and between 0.88 and 11.1 mg/kg DW in the case of plants, while the Li content in the soil samples were slightly comparable (from 9.85 to 11.3 mg/kg DW). In the Banat region, the concentration of Li was lower than in Dobrogea (1.40–1.46 μg/L in water, 6.50–9.12 mg/kg DW in soil, and 0.19–0.45 mg/kg DW in plants). Despite the high Li contents in soil, the Li was mostly unavailable for plants uptake and bioaccumulation.