Synthesis and Evaluation of the Insulin–Albumin Conjugate with Prolonged Glycemic Control

[Image: see text] Engineering therapeutic proteins to improve their half-life so as to sustain physiologically relevant extended activity is the need of the hour in biopharmaceutical research. In this study, insulin and bovine serum albumin (BSA) were independently functionalized rationally and were later conjugated to prolong the half-life of insulin. The thiol functionalization of BSA with 2-imminothiolane in the ratio 1:20 yielded an average of 6–8 thiols/BSA, which then reacted with maleimide-functionalized insulin to form an insulin–albumin conjugate. The bioconjugate was purified by size exclusion chromatography, and the increase in size was confirmed by sodium dodecyl-sulfate polyacrylamide gel electrophoresis. Bioconjugation resulted in a multi-fold increase in the hydrodynamic volume of the insulin–albumin conjugate as measured in DLS when compared to BSA. The glucose uptake assay with 3LT3-L1 cell lines was performed, and the mean fluorescence intensity (MFI) of 16.16 observed for the insulin–albumin conjugate was comparable to insulin (19.42). The blood glucose reducing capacity of the insulin–albumin conjugate in streptozotocin induced diabetic male Wistar rats was well maintained up to 72 h when compared to native insulin. Further, a three-fold increase in plasma insulin concentration was observed in bioconjugate treated animals as against insulin treated animals after 24 h of treatment using ELISA. The histological analysis of different organs of the bioconjugate treated rats indicated that it was non-toxic. This study has paved a way for further detailed studies on similar bioconjugates to develop next-generation biotherapeutics for treating diabetes.