Protein drug transporters play an important role in the absorption, distribution, metabolism and excretion of drugs. The study of protein-drug interaction is a significant issue for drug development. Drug transporters are multi-specific transmembrane proteins that facilitate the membrane passage of a large number of drugs. Drug transporters have a distinct expression pattern in the human body lining pharmacological barrier tissues, most importantly the small intestinal epithelium, the endothelial cells in the blood–brain barrier, the epithelium of the proximal tubule cells in the kidney, and hepatocytes in the liver. However, it is both expensive and time-consuming to perform physical experiments to determine whether a drug and a protein are interacting with each other, while these studies are really useful in medical sciences. Bovine milk β-lactoglobulin (b-LG) demonstrates significant resistance against both gastric and simulated duodenal digestions. Therefore, it seems a realistic protein candidate for safe delivery and protection of particularly pH sensitive drugs in stomach. Prednisolone is a synthetic adrenal corticosteroid which is used to achieve prompt suppression of inflammation in many inflammatory and allergic conditions and to treat blood cell cancers (leukemia (and lymph gland cancers (lymphomas). In this study the intermolecular interaction of prednisolone with b -LG was investigated using UV-Vis spectroscopy. Moreover, the effect of Prednisolone complexation on the secondary structures of b -LG was studied and the results showed that the secondary structure of b -LG was preserved upon interaction of this drug. Based on the achieved results, this protein might be useful for delivery of Prednisolone.