Tan M M, Corley C A, Stevenson C L
Biopharmaceutical R & D, ALZA Corporation, Palo Alto, California 94303, USA.
Pharm Res. 1998 Sep;15(9):1442-8. doi: 10.1023/a:1011914007940.
The purpose of this study was to characterize the conformation, aggregation, and stability of leuprolide on gelation.
Infrared spectra (FTIR) of leuprolide solutions and gels were collected in water, propylene glycol (PG), dimethyl sulfoxide (DMSO), and trifluoroethanol (TFE). Leuprolide solution and gel stability data were obtained by SEC and RP-HPLC.
Leuprolide was induced to gel with increasing peptide concentration, introduction of salts, and gentle agitation. Leuprolide dissolved in water (400 mg/ml) demonstrated FTIR spectra consisting of two major bands of equal intensity at 1615 cm(-1) and 1630 cm(-1), similar to inter- and intra-molecular beta-sheet structure in proteins. When samples were gently agitated for 24 hours at 25 degrees C, the formulation was observed to change from a viscous liquid to an opaque gel with a concomitant shift in infrared spectra from the equal intensity bands to mostly 1630 cm(-1), indicating a shift to a preferred beta-sheet structure. Incubation of leuprolide with 20-200 mM salts at 25 degrees C and 37 degrees C also produced gels ranging from clear to cloudy and stringy white precipitates. The gel and precipitate were marked by a shift of the predominant beta-sheet band to 1630 cm(-1) and 1615 cm(-1), respectively. Leuprolide was also observed to gel and/or precipitate in mixtures of water, PG or TFE, but not in DMSO.
Birefringence was noted in many of the firmer gels. Both solutions and gels demonstrated minimal dimer or trimer formation, with no larger order aggregates detected. The chemical stability profile of gelled leuprolide was similar to that of the non-gelled water formulation by RP-HPLC.
本研究的目的是表征亮丙瑞林在凝胶化过程中的构象、聚集和稳定性。
在水、丙二醇（PG）、二甲基亚砜（DMSO）和三氟乙醇（TFE）中收集亮丙瑞林溶液和凝胶的红外光谱（FTIR）。通过尺寸排阻色谱（SEC）和反相高效液相色谱（RP-HPLC）获得亮丙瑞林溶液和凝胶的稳定性数据。
随着肽浓度的增加、盐的引入和轻轻搅拌，亮丙瑞林被诱导形成凝胶。溶解在水中（400 mg/ml）的亮丙瑞林的FTIR光谱由1615 cm⁻¹和1630 cm⁻¹处两个强度相等的主要谱带组成，类似于蛋白质中的分子间和分子内β-折叠结构。当样品在25℃下轻轻搅拌24小时时，观察到制剂从粘性液体变为不透明凝胶，同时红外光谱从强度相等的谱带转变为主要为1630 cm⁻¹，表明转变为优选的β-折叠结构。在25℃和37℃下，将亮丙瑞林与20 - 200 mM的盐孵育也会产生从澄清到浑浊以及丝状白色沉淀的凝胶。凝胶和沉淀分别以主要β-折叠带向1630 cm⁻¹和1615 cm⁻¹的移动为特征。还观察到亮丙瑞林在水、PG或TFE的混合物中形成凝胶和/或沉淀，但在DMSO中不形成。
在许多较硬的凝胶中观察到双折射现象。溶液和凝胶均显示出极少的二聚体或三聚体形成，未检测到更大的有序聚集体。通过RP-HPLC分析，凝胶化的亮丙瑞林的化学稳定性与非凝胶化的水性制剂相似。
