Department of Pharmaceutics, College of Pharmacy, University of Minnesota, Minneapolis, Minnesota 55455, United States.
Pfizer Biotherapeutics, Pfizer Inc., Andover, Massachusetts 01810, United States.
Mol Pharm. 2023 Sep 4;20(9):4587-4596. doi: 10.1021/acs.molpharmaceut.3c00312. Epub 2023 Aug 3.
The phase behavior of poloxamer 188 (P188) in aqueous solutions, characterized by differential scanning calorimetry (DSC) and synchrotron X-ray diffractometry, revealed solute crystallization during both freezing and thawing. Sucrose and trehalose inhibited P188 crystallization during freeze-thawing (FT). While trehalose inhibited P188 crystallization only during cooling, sucrose completely suppressed P188 crystallization during both cooling and heating. Lactate dehydrogenase (LDH) served as a model protein to evaluate the stabilizing effect of P188. The ability of P188, over a concentration range of 0.003-0.800% w/v, to prevent LDH (10 μg/mL) destabilization was evaluated. After five FT cycles, the aggregation behavior (by dynamic light scattering) and activity recovery were evaluated. While LDH alone was sensitive to interfacial stress, P188 at concentrations of ≥0.100% w/v stabilized the protein. However, as the surfactant concentration decreased, protein aggregation after FT increased. The addition of sugar (1.0% w/v; sucrose or trehalose) improved the stabilizing function of P188 at lower concentrations (≤0.010% w/v), possibly due to the inhibition of surfactant crystallization. Based on a comparison with the stabilization effect of polysorbate (both 20 and 80), it was evident that P188 could be a promising alternative surfactant in frozen protein formulations. However, when the surfactant concentration is low, the potential for P188 crystallization and the consequent compromise in its functionality warrant careful consideration.
聚氧乙烯-188 （P188）在水溶液中的相行为，通过差示扫描量热法（DSC）和同步加速器 X 射线衍射法进行了表征，发现溶质在冷冻和解冻过程中结晶。蔗糖和海藻糖抑制了 P188 在冻融过程中的结晶（FT）。虽然海藻糖仅在冷却过程中抑制 P188 结晶，但蔗糖完全抑制了 P188 在冷却和加热过程中的结晶。乳酸脱氢酶（LDH）作为模型蛋白，用于评估 P188 的稳定作用。评估了 P188 在 0.003-0.800% w/v 浓度范围内防止 LDH（10 μg/mL）失稳的能力。经过五次 FT 循环后，评估了聚集行为（通过动态光散射）和活性恢复。虽然 LDH 本身对界面应力敏感，但浓度为≥0.100% w/v 的 P188 稳定了该蛋白。然而，随着表面活性剂浓度的降低，FT 后蛋白质聚集增加。添加糖（1.0% w/v；蔗糖或海藻糖）可提高 P188 在较低浓度（≤0.010% w/v）下的稳定功能，这可能是由于抑制了表面活性剂结晶。与聚山梨醇酯（20 和 80）的稳定效果进行比较后，显然 P188 可能是冷冻蛋白制剂中一种有前途的替代表面活性剂。然而，当表面活性剂浓度较低时，P188 结晶的可能性及其功能的潜在妥协需要仔细考虑。
