ApoA-I, which constitutes 70% of the apolipoprotein content of high-density lipoproteins, acts as an acceptor for the transfer of phospholipids and free cholesterol from peripheral tissues and transports cholesterol in the liver and other tissue for excretion and steroidogenesis. In order to verify its possible structural alteration in pathological states, plasma samples from healthy, diabetic, and nephropathic subjects have been analyzed by two-dimensional gel electrophoresis. By this approach, clear differences among the three classes of subjects become evident and, in the case of diabetic and nephropathic patients, intense spots are present. The matrix-assisted laser desorption/ionization mass spectrometry analysis of their digestion products shows that an overexpression of unglycated and glycated ApoA-I is present in both pathological states, reasonably affecting the efficiency in cholesterol transport.