TY - JOUR
T1 - Microheterogeneity in frozen protein solutions
AU - Twomey, Alan
AU - Kurata, Kosaku
AU - Nagare, Yutaka
AU - Takamatsu, Hiroshi
AU - Aksan, Alptekin
N1 - Publisher Copyright:
© 2015 Elsevier B.V.
PY - 2015/6/20
Y1 - 2015/6/20
N2 - In frozen and lyophilized systems, the biological to be stabilized (e.g. therapeutic protein, biomarker, drug-delivery vesicle) and the cryo-/lyo-protectant should be co-localized for successful stabilization. During freezing and drying, many factors cause physical separation of the biological from the cryo-/lyo-protectant, called microheterogeneity (MH), which may result in poor stabilization efficiency. We have developed a novel technique that utilized confocal Raman microspectroscopy in combination with counter-gradient freezing to evaluate the effect of a wide range of freezing temperatures (-20 < TF < 0 °C) on the MH generated within a frozen formulation in only a few experiments. The freezing experiments conducted with a model system (albumin and trehalose) showed the presence of different degrees of MH in the freeze-concentrated liquid (FCL) in all solutions tested. Mainly, albumin tended to accumulate near the ice interface, where it was physically separated from the cryoprotectant. In frozen 10 wt% trehalose solutions, heterogeneity in FCL was relatively low at any TF. In frozen 20 wt% trehalose solutions, the optimum albumin to trehalose ratio in the FCL can only be ensured if the solution was frozen within a narrow range of temperatures (-16 < TF < -10 °C). In the 30 wt% trehalose solutions, freezing within a much more narrow range (-12 < TF < -10 °C) was needed to ensure a fairly homogeneous FCL. The method developed here will be helpful for the development of uniformly frozen and stable formulations and freezing protocols for biological as MH is presumed to directly impact stability.
AB - In frozen and lyophilized systems, the biological to be stabilized (e.g. therapeutic protein, biomarker, drug-delivery vesicle) and the cryo-/lyo-protectant should be co-localized for successful stabilization. During freezing and drying, many factors cause physical separation of the biological from the cryo-/lyo-protectant, called microheterogeneity (MH), which may result in poor stabilization efficiency. We have developed a novel technique that utilized confocal Raman microspectroscopy in combination with counter-gradient freezing to evaluate the effect of a wide range of freezing temperatures (-20 < TF < 0 °C) on the MH generated within a frozen formulation in only a few experiments. The freezing experiments conducted with a model system (albumin and trehalose) showed the presence of different degrees of MH in the freeze-concentrated liquid (FCL) in all solutions tested. Mainly, albumin tended to accumulate near the ice interface, where it was physically separated from the cryoprotectant. In frozen 10 wt% trehalose solutions, heterogeneity in FCL was relatively low at any TF. In frozen 20 wt% trehalose solutions, the optimum albumin to trehalose ratio in the FCL can only be ensured if the solution was frozen within a narrow range of temperatures (-16 < TF < -10 °C). In the 30 wt% trehalose solutions, freezing within a much more narrow range (-12 < TF < -10 °C) was needed to ensure a fairly homogeneous FCL. The method developed here will be helpful for the development of uniformly frozen and stable formulations and freezing protocols for biological as MH is presumed to directly impact stability.
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U2 - 10.1016/j.ijpharm.2015.04.032
DO - 10.1016/j.ijpharm.2015.04.032
M3 - Article
C2 - 25888798
AN - SCOPUS:84927728698
SN - 0378-5173
VL - 487
SP - 91
EP - 100
JO - International journal of pharmaceutics
JF - International journal of pharmaceutics
IS - 1-2
ER -