Arginine as a protein stabilizer and destabilizer in liquid formulations

作者：Kim Nam Ah; Hada Sharvron; Thapa Ritu; Jeong Seong Hoon^*

来源：International Journal of Pharmaceutics, 2016, 513(1-2): 26-37.

DOI：10.1016/j.ijpharm.2016.09.003

• Arginine oxidation
• Protein denaturant
• Protein aggregation
• DSC
• Etanercept
• NOx

Even though arginine monohydrochloride (ArgHCl) is a useful additive for protein stabilization, its mechanism is not yet fully elucidated. Moreover, there is a concern that ArgHCl may be a protein denaturant since it decreases transition melting temperature (T-m) of certain proteins. It contains a guanidinium group, a critical structure for denaturating activity of guanidine hydrochloride (GndHCl). Effects of ArgHCl, GndHCl, and creatinine on a model protein, etanercept, were examined by biophysical analyses including DLS, DSC, FT-IR, microviscometer, and SEC. Accelerated storage stability of the protein was examined in the absence and presence of H2O2 at different incubation temperatures with pH monitoring. ArgHCl reduced protein aggregation and retained monomer, but increased fragmentation at high temperature. T(m)1 and T(m)2 of the protein increased with ArgHCl, but slight decrease (>1 degrees C) in T(m)3 was observed. GndHCl and creatinine decreased all three T(m)s. In the presence of heat and H2O2, the effect of ArgHCl was significantly decreased compared to GndHCl and creatinine. In addition, it accelerated the loss of monomer and increased fragmentation with decreasing pH. ArgHCl differed from GndHCl in the mode of physical interaction with the protein, due to its unique balance of three steric functional groups (guanidinium, carboxylic acid, and carbon aliphatic straight chain). In contrast, ArgHCl acted as a protein denaturant at high temperature since NOx generated from the amine group at the 3-carbon aliphatic straight chain and it is supported by GndHCl which did not change the pH nor accelerated the monomer loss after oxidation by H2O2 at high temperature.

• 出版日期2016-11-20