Free fatty acid particles in protein formulations, part 1: microspectroscopic identification.

We report, for the first time, the identification of fatty acid particles in formulations containing the surfactant polysorbate 20. These fatty acid particles were observed in multiple mAb formulations during their expected shelf life under recommended storage conditions. The fatty acid particles were granular or sand-like in morphology and were several microns in size. They could be identified by distinct IR bands, with additional confirmation from energy-dispersive X-ray spectroscopy analysis. The particles were readily distinguishable from protein particles by these methods. In addition, particles containing a mixture of protein and fatty acids were also identified, suggesting that the particulation pathways for the two particle types may not be distinct. The techniques and observations described will be useful for the correct identification of proteinaceous versus nonproteinaceous particles in pharmaceutical products.

[1]  Z. Wen,et al.  Identification of a Mixed Microparticle by Combined Microspectroscopic Techniques: A Real Forensic Case Study in the Biopharmaceutical Industry , 2010, Applied spectroscopy.

[2]  Jennifer R Litowski,et al.  Free fatty acid particles in protein formulations, part 2: contribution of polysorbate raw material. , 2015, Journal of pharmaceutical sciences.

[3]  Wei Wang,et al.  Immunogenicity of protein aggregates--concerns and realities. , 2012, International journal of pharmaceutics.

[4]  E. Maggio,et al.  Polysorbates, peroxides, protein aggregation, and immuno- genicity - a growing concern. , 2012 .

[5]  W. S. Singleton,et al.  The infrared spectra of saturated fatty acids with even number of carbon atoms from caproic, C6 (Hexanoic), to stearic, C18 (Octadecanoic), and of their methyl and ethyl esters , 1951 .

[6]  Hanns-Christian Mahler,et al.  The Degradation of Polysorbates 20 and 80 and its Potential Impact on the Stability of Biotherapeutics , 2011, Pharmaceutical Research.

[7]  Wei Wang,et al.  Protein aggregation--pathways and influencing factors. , 2010, International journal of pharmaceutics.

[8]  Y. Kao,et al.  Analysis Methods of Polysorbate 20: A New Method to Assess the Stability of Polysorbate 20 and Established Methods That May Overlook Degraded Polysorbate 20 , 2002, Pharmaceutical Research.

[9]  B. Kerwin.  Polysorbates 20 and 80 used in the formulation of protein biotherapeutics: structure and degradation pathways. , 2008, Journal of pharmaceutical sciences.

[10]  N. Dixit,et al.  The effect of Tween(®) 20 on silicone oil-fusion protein interactions. , 2012, International journal of pharmaceutics.

[11]  R. Jones,et al.  The Infrared Absorption Spectra of Saturated Fatty Acids and Esters1 , 1952 .

[12]  Z. Wen,et al.  Identification and Root Cause Analysis of Cell Culture Media Precipitates in the Viral Deactivation Treatment with High-Temperature/Short-Time Method , 2013, PDA Journal of Pharmaceutical Science and Technology.

[13]  B. Kerwin,et al.  Effects of Tween 80 and sucrose on acute short-term stability and long-term storage at -20 degrees C of a recombinant hemoglobin. , 1998, Journal of pharmaceutical sciences.

[14]  A. P. Gunning,et al.  SURFACTANT-PROTEIN INTERACTIONS AT AIR-WATER AND OIL-WATER INTERFACES OBSERVED BY ATOMIC FORCE MICROSCOPY , 2000 .

[15]  R. Jones.  THE EFFECTS OF CHAIN LENGTH ON THE INFRARED SPECTRA OF FATTY ACIDS AND METHYL ESTERS , 1962 .

[16]  R. Jones,et al.  Band Progressions in the Infrared Spectra of Fatty Acids and Related Compounds1 , 1952 .

[17]  Wei Wang,et al.  Antibody structure, instability, and formulation. , 2007, Journal of pharmaceutical sciences.

[18]  B. Kerwin,et al.  Effects of Tween 80 and Sucrose on Acute Short-Term Stability and Long-Term Storage at − 20 C of a Recombinant Hemoglobin , 1998 .

[19]  Theodore W Randolph,et al.  Surfactant-protein interactions. , 2002, Pharmaceutical biotechnology.

[20]  A. Pappenberger,et al.  Degradation of polysorbates 20 and 80: studies on thermal autoxidation and hydrolysis. , 2011, Journal of pharmaceutical sciences.

[21]  R. Jones,et al.  The Infrared Absorption Spectra of Unsaturated Fatty Acids and Esters1 , 1952 .