Effects of mixed surfactant system on chemical and physical stability of human growth hormone

Sammanfattning: In protein formulation, addition of surfactants is commonly executed to enhance product stability and shelf-life. The reason is that surfactants can protect proteins against surface-induced aggregation. However, the interaction between surfactant and protein depends on the type of surfactant used. Anionic surfactants can unfold proteins and form complexes in equilibrium manner. The complex structure depends on the ratio of anionic surfactant-to-protein. On the other hand, nonionic surfactants can weaken interactions between anionic surfactants and proteins. Here, human growth hormone (hGH) is used. Sodium dodecyl sulfate (SDS) in the SDS-hGH complex was slowly extracted to form mixed micelles with n-dodecyl-β-D-maltopyranoside (DDM). This gentle process allows the protein to refold back to its native form with less aggregation. From previous studies, the DDM/SDS ratio is pivotal for the protein refolding procedure using a nonionic surfactant, regardless of mixing order. This study is aimed at assessing the stability of refolded SDS-unfolded hGH after incubating at 37°C for 42 days. Regarding hGH properties, deamidation seems to be dominant in chemical degradation. One of the formulations was designed for less deamidation, at pH 5.4, to eliminate other destabilizing effects compared to pH 7. In the physical stability aspect, the SDS-hGH formulation at pH 5.4 had the lowest stability of refolded hGH, as seen in the results from FlowCAM, DLS, and Probe Drum. However, the refolded hGH structure in this formulation was the most chemically stable as seen from HPLC analysis. Nuclear magnetic resonance is needed to assess the complex conformation in this formulation.