Department of Chemistry and Polymer Science, University of Stellenbosch, Private Bag X1, Stellenbosch, 7602, South Africa.
L'Oréal Research and Innovation, 93600, Aulnay-sous-Bois, France.
Anal Bioanal Chem. 2019 Jun;411(15):3321-3330. doi: 10.1007/s00216-019-01799-x. Epub 2019 Apr 15.
Hyaluronic acid and its acrylate derivatives are important intermediates for various pharmaceutical, biomedical, and cosmetic applications due to their biocompatibility and viscoelasticity properties. However, these polymers are inherently difficult to characterize due to their significant heterogeneity regarding molar mass and chemical composition (degree of substitution, DS). The present study describes the development of a comprehensive online two-dimensional liquid chromatography (2D-LC) approach to characterize hyaluronic acid and its acrylate derivatives (DS ranging from 0.4 to 3.1) in terms of molar mass and degree of substitution. In the first dimension of the 2D-LC method, separation according to chemical composition/DS was achieved by using a stepwise solvent gradient and a reversed phase C8 column. Fractions from the first dimension were automatically transferred to the second dimension comprising size exclusion chromatographic separation of the fractions according to molar mass. It was found that the hyaluronic acid derivatives were broadly distributed with regard to both chemical composition and molar mass. Fractions with different degrees of substitution were identified, and their molar mass distributions were determined. The study proved that comprehensive 2D-LC is a powerful approach to reveal the complex nature of hyaluronic acid and its derivatives. Graphical abstract.
透明质酸及其丙烯酰胺衍生物由于其生物相容性和粘弹性而成为各种药物、生物医学和化妆品应用的重要中间体。然而，由于这些聚合物在摩尔质量和化学组成（取代度，DS）方面具有显著的异质性，因此它们本质上难以进行表征。本研究描述了一种综合的在线二维液相色谱（2D-LC）方法的开发，用于根据摩尔质量和取代度来表征透明质酸及其丙烯酰胺衍生物（DS 范围为 0.4 至 3.1）。在 2D-LC 方法的第一维中，通过使用逐步溶剂梯度和反相 C8 柱，根据化学组成/DS 进行分离。一维的馏分自动转移到二维，根据摩尔质量对馏分进行尺寸排阻色谱分离。结果发现，透明质酸衍生物在化学组成和摩尔质量方面都具有广泛的分布。鉴定了具有不同取代度的馏分，并确定了它们的摩尔质量分布。该研究证明，全面的 2D-LC 是一种揭示透明质酸及其衍生物复杂性质的强大方法。
