Although polymer-based ATPS represents an eco-compatible alternative to conventional organic/aqueous extraction techniques, it has the disadvantage of the difficulty of separating the target compounds from the phase-forming polymers and recovering the polymers after the extraction. The use of a polymer phase immobilized to a solid support or a cross-linked polymer gel is one of the most effective methods to overcome these problems because the polymer gels or the immobilized polymer matrices are isolated from the aqueous solution phase. This chapter discloses the separation mechanism exerted by some hydrophilic polymer gels conventionally used as column packing materials for aqueous liquid chromatography (dextran, polyacrylamide, polyethylene glycol, and methacrylate) in relation to their hydration structures. Liquid chromatography and differential scanning calorimetry studies indicate that the separation of solute compounds in an aqueous solution with the hydrated cross-linked polymer gels is caused by the partition of the solutes into hydrated polymer gel phase similar to the polymer-based ATPS. Water in the polymer gel phase has a different affinity to solutes from that of the bulk water and functions as a component of the separation media. The stationary phase water shows the melting behavior different from that of the bulk water.