The impact of silicone oil on the formation of free fatty acids particles.

Silicone oil is a ubiquitous presence throughout the life cycle of biopharmaceutical products, originating from manufacturing processes and primary packaging like pre-filled syringes. While its role in protein aggregation is well-studied, its impact on degradants like free fatty acids (FFAs) remains poorly understood. The only prior study on this topic concluded that silicone oil did not delay FFA particle formation, despite confirming FFA partitioning into the oil phase. Here, we demonstrate for the first time that the presence of silicone oil (viscosity 50-10,000 cSt) delays the growth kinetics and suppresses the accumulation of FFA particles over a 10-week study period. This effect was observed even at silicone oil concentrations as low as 2.7 µg/mL. Using a multi-modal complementary analytical suite-comprising total holographic characterization (THC), backgrounded membrane imaging (BMI), and flow imaging (FI)-we tracked FFA particles from the early-stage sub-micron range (500 nm) to the visible range. Furthermore, machine learning-assisted morphological analysis revealed that silicone oil fundamentally reconfigures the physical state of the FFA precipitate, transitioning from a characteristic dendritic structure to a semi-transparent film-like morphology.