The development of new ligand systems for solvent extraction demands knowledge of the fundamental chemistry behind these processes. Thus, herein, we report the synthesis and structural characterization of a 6-hydro-xymethylpyridine functionalized calix[4]arene (L-2). Its complexation behavior towards the divalent uranyl cation as well as the trivalent europium cation was spectroscopically studied. In solution the formation of two uranyl complex species with L-2 was proven by spectrophotometric titration experiments, with stability constants of log beta(1:1) = 5.82 +/- 0.04 and log beta(2:1) = 5.57 +/- 0.05, and of one europium complex species (log beta(1:1) = 6.85 +/- 0.03). In addition, the concept of the desired proper planar binding pocket for the linear uranyl cation has been verified in solution. Moreover, comparative U-VI and Eu-III extraction studies with L-2 and a structure related 8-hydroxyquinoline modified calix[4]arene (L-1) were performed in which the effects of various conditions such as pH of the aqueous phase, presence of relevant anions (sulfate, phosphate, carbonate) and competitive extraction with ethylenediaminetetraacetic acid (EDTA) were investigated. It has been shown that both macrocyclic ligands offer a strong chelate effect, which enables them to extract even stable uranyl hydrolysis species. With these ligands, the selective separation of U-IV from Eu-III-containing solutions is possible under a variety of conditions.