(595f) Probing Molecular Interactions between Protein Drugs and Silicone Oil at Buried Liquid/Liquidinterfaces in Situ in Real Time

Protein drugs such as antibody drugs have been rapidly developed to cure a variety of diseases. Silicone oil is commonly used as lubricant coating material for devices used in the pharmaceutical industry to store and administrate protein drug formulations. Protein molecules may strongly interact with silicone oil, which lead to the protein adsorption, denaturation, and aggregation. This could negatively impact the efficacy of protein drugs. Therefore, it is important to understand the interactions between proteins and silicone oil at the liquid/liquid interface in situ at the molecular level. However, such interfaces are buried, thus it is difficult to probe them in situ due to the lack of appropriate techniques. In the recent years, we developed sum frequency generation (SFG) vibrational spectroscopy into a powerful tool to study molecular interactions and molecular structures of proteins at the buried solid/liquid and liquid/liquid interfaces. Using SFG, we successfully studied molecular interactions between silicone oil and a monoclonal antibody as well as a bispecific antibody at the protein solution/silicone oil interface in situ in real time at the molecular level. It was found that the adsorbed antibody on silicone oil denatured, while with 0.002 % surfactant in the protein solution, adsorbed protein on silicone oil could adopt a native structure. With 0.02 % surfactant in the solution, surfactant could prevent antibody from adsorption to the silicone oil. We also applied SFG to investigate molecular interactions of a fusion protein with silicone oil at the liquid/liquid interface in situ, and how surfactant could prevent adsorption of the fusion protein to silicone oil. In addition, with model proteins, we successfully demonstrated that SFG can be used to study protein aggregations on silicone oil by showing that SFG can distinguish monomer, dimer, and monomer-dimer mixtures adsorbed on silicone oil. The protein adsorption kinetics, time-dependent coverage and structural changes on silicone oil were also examined in detailed using SFG. Our research showed that SFG is a generally applicable analytical tool to elucidate molecular interactions of proteins at solid/liquid and liquid/liquid interfaces in situ in real time.