A Small-Scale Process for Predicting Donnan and Volume Exclusion Effects During Ultrafiltration/Diafiltration Process Development

Abstract

Achieving the desired final protein formulation using ultrafiltration/diafiltration (UF/DF) operations is an essential component of many protein purification processes. It is well documented that differences in the excipient and buffer concentrations exist between the DF and retentate solutions when they have achieved equilibrium. Several publications have proposed ways to calculate these differences. However, the accuracy of these methods has been limited by the use of an estimated protein charge value. In this article, a small-scale system is described, which can accurately determine the protein charge by making buffer and excipient concentration measurements and applying the determined values to the Donnan and volume exclusion equations. This information can be utilized to generate a standard curve, which in turn can be applied to at-scale UF/DF operations. For 2 different antibodies, the standard curve generated by the small-scale system yielded buffer concentrations and pH values that agreed well with those generated after UF/DF operations, whereas using the theoretical protein charge caused a departure from the measured results. This model also provides good estimates as to how the final formulation pH and buffer concentration vary as a function of the pH and buffer concentration in the DF buffer. This information is of important utility for the accurate formulation of high-concentration protein solutions (>100 mg/mL) where the coconcentration of buffers and the volume exclusion of certain excipients are amplified. The low material requirements of the small-scale system are a major benefit for early phase formulation and process development when sufficient time and material may not be available, in particular to ensure successful UF/DF operations for the development of high protein concentration formulations.