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Updated: Mar 25, 2026

A Microfluidic Approach for the Study of Ice and Clathrate Hydrate Crystallization
Published on: August 18, 2022
Mutual Influence of Mannitol and Trehalose on Crystallization Behavior in Frozen Solutions
Sampreeti Jena1, Raj Suryanarayanan2, Alptekin Aksan3,4
1Biostabilization Laboratory, Department of Mechanical Engineering, University of Minnesota, Minneapolis, Minnesota, 55455, USA.
Purpose:
Phase separation of trehalose during freeze-drying could render it ineffective as a lyoprotectant. The bulking agent, mannitol, on the other hand, should crystallize readily upon freezing. It is therefore imperative to understand the mutual interaction of these sugars during freezing to ensure preservation of the API during freeze-drying.
Methods:
We investigated the effect of mannitol to trehalose ratio (R) on the crystallization behavior of both solutes using Differential Scanning Calorimetry, X-Ray Crystallography and FTIR Spectrosopy during controlled freezing and annealing.
Results:
When R = 1, crystallization of both mannitol (as hemihydrate) and trehalose (as α-anhydrate) were observed. When R ≥ 1, extent of mannitol crystallization was directly proportional to the value of R. When R < 1, trehalose completely suppressed mannitol crystallization. At R > 1, the freeze concentrate was heterogeneous and characterized by two glass transitions - the lower temperature transition (Tg") attributed to systems containing "extra" unfrozen water. When heated above Tg", crystallization of mannitol and the associated unfrozen water resulted in Tg' (glass transition temperature of the freeze-concentrate).
Conclusions:
R and not the total solute concentration, dictates the composition of the freeze concentrate as well as the physical stability of the excipients.
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