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Microheterogeneity in frozen protein solutions
Alan Twomey1, Kosaku Kurata2, Yutaka Nagare2
1Biostabilization Laboratory, Mechanical Engineering Department, College of Science and Engineering, University of Minnesota, Minneapolis, MN 55455, United States.
International Journal of Pharmaceutics
|April 19, 2015
Summary
Physical separation, or microheterogeneity, of biologics from cryo-protectants during freezing hinders stabilization. This study introduces a novel method to optimize freezing temperatures for uniform frozen formulations and improved biological stability.
Area of Science:
- Biophysics
- Formulation Science
- Analytical Chemistry
Background:
- Effective stabilization of biologics (proteins, vesicles) in frozen or lyophilized systems requires co-localization with cryo-/lyo-protectants.
- Physical separation, termed microheterogeneity (MH), during freezing can compromise stabilization efficiency by separating biologics from protectants.
Purpose of the Study:
- To develop and validate a novel technique for evaluating microheterogeneity (MH) in frozen formulations across a range of freezing temperatures.
- To determine the impact of freezing temperatures on the distribution of albumin and trehalose in freeze-concentrated liquid (FCL).
Main Methods:
- Utilized confocal Raman microspectroscopy combined with counter-gradient freezing to assess MH.
- Investigated a model system comprising albumin and trehalose across various freezing temperatures (-20°C to 0°C).
Main Results:
- Microheterogeneity (MH) was observed in all tested solutions, with albumin accumulating near the ice interface, separate from trehalose.
- Low MH was found in 10wt% trehalose solutions across all tested freezing temperatures.
- Optimal albumin-to-trehalose ratios in the FCL required specific freezing ranges: -16°C to -10°C for 20wt% trehalose and -12°C to -10°C for 30wt% trehalose.
Conclusions:
- The developed method effectively quantifies MH in frozen formulations, aiding in the optimization of freezing protocols.
- Controlling freezing temperature is critical for achieving homogeneous FCL and ensuring the stability of biological formulations.
- This technique facilitates the development of uniformly frozen and stable formulations for biologics.
