The modification of human insulin by the covalent attachment of monosaccharide moieties to insulin amino group(s) alters the aggregation and self-association behavior, improving both the pharmaceutical stability and biological response. The synthesis of p-succinamidophenyl glucopyranoside-insulin conjugate(s) (SAPG-insulin) has resulted in seven possible glucosylated insulin derivatives (three monosubstituted, three disubstituted, and one trisubstituted). These derivatives were isolated and purified using ion exchange chromatography. Characterization of the derivatives includes determining the site and number of sugar groups attached for each individual derivative and an evaluation of biological activity. Nearly all the derivatives retained in vivo biological activity comparable to insulin. In addition, extensive physicochemical characterization of the glucosylated insulin derivatives was undertaken to determine association/aggregation properties using GPC, dynamic light scattering, UV/Vis, and CD spectroscopy. Protein self-association was most suppressed with the disubstituted derivatives, especially those substituted on PheB1, and the trisubstituted derivative. The same general pattern was observed for physical stability of glucosylated insulin derivatives. As the number of glucosyl moieties attached to insulin increased, solution physical stability dramatically improved. Yet, the most significant impact to stability was glycosylation at the PheB1 site.