Chemical Sciences Division, National Institute of Standards and Technology (NIST), 100 Bureau Drive, MS 8392, Gaithersburg, MD, 20899-8392, USA.
Chemical Sciences Division, National Institute of Standards and Technology (NIST), 100 Bureau Drive, MS 8392, Gaithersburg, MD, 20899-8392, USA.
Anal Chim Acta. 2019 Apr 11;1053:186-195. doi: 10.1016/j.aca.2018.11.051. Epub 2018 Dec 5.
Size-exclusion chromatography (SEC) with on-line static light scattering, specifically multi-angle static light scattering (MALS), and differential refractometry (DRI) detection remains the premier method by which to determine absolute, calibrant-independent molar masses of polymers. The method is restricted to the use of either neat solvents or solvents with a small amount of additive. In mixed solvents, preferential solvation (i.e., the enrichment, within the solvated volume of the polymer in solution, of one solvent over the other as compared to the solvent ratio outside said volume) leads to errors in the areas of the MALS and DRI chromatograms, as the solvent baseline does not accurately represent the solvent contribution to these detectors' peaks. A seemingly trivial way by which to overcome this problem is through the use of an isorefractive solvent pair. This "trivial" solution is complicated by the fact that the solvents in the pair must be miscible with each other in all proportions; the individual solvents as well as the mix must be able to fully dissolve the analyte; the solvents must possess sufficient optical contrast with the solution so as to generate an adequate detector signal; the solvent mix must be compatible with the chromatographic stationary phase, such that enthalpic contributions to the separation are minimal and analyte recovery from the columns is quantitative; and the difference in the Rayleigh factors of the solvents can be ignored. Herein, we present the analysis of narrow dispersity polystyrene (PS) and poly(methyl methacrylate) (PMMA) samples, across a four-fold range in molar mass, using SEC/MALS/DRI in a mix of tetrahydrofuran (THF) and methyl isoamyl ketone (MIAK), solvents which are shown to be isorefractive with each other at the temperature and wavelength of the experiments. Molar mass averages and dispersities are demonstrated to be statistically independent of solvent composition and to correspond well to the values in neat THF. The experiments were augmented by the use of on- and off-line quasi-elastic light scattering and of off-line MALS and DRI, to study the effect of solvent composition on polymer size in solution and on dilute solution thermodynamics. Additionally, H nuclear magnetic resonance spectroscopy was used to study the effect of tacticity on the insolubility of PMMA100 in 100% MIAK. We believe this constitutes the first example of obtaining accurate molar masses of polymers by SEC/MALS/DRI employing mixed solvents. The value of these experiments to other forms of macromolecular liquid chromatographic separations is also noted.
体积排阻色谱（SEC）与在线静态光散射，特别是多角度静态光散射（MALS）和差示折射检测，仍然是确定聚合物绝对、无校准剂摩尔质量的首选方法。该方法仅限于使用纯溶剂或少量添加剂的溶剂。在混合溶剂中，优先溶解（即与溶剂体积外的溶剂比相比，聚合物在溶液中的溶剂体积内一种溶剂相对于另一种溶剂的富集）会导致 MALS 和 DRI 色谱图中的面积出现误差，因为溶剂基线不能准确代表这些检测器峰的溶剂贡献。一种看似简单的克服此问题的方法是使用等折射溶剂对。然而，由于溶剂对中的溶剂必须以任意比例互溶；各个溶剂以及混合物必须能够完全溶解分析物；溶剂必须与溶液具有足够的光学对比度，以产生足够的检测器信号；溶剂混合物必须与色谱固定相兼容，使得分离的焓贡献最小，并且从柱子中定量回收分析物；并且溶剂的瑞利因子差异可以忽略不计。在此，我们在实验温度和波长下，使用 SEC/MALS/DRI 分析了窄分散聚苯乙烯（PS）和聚甲基丙烯酸甲酯（PMMA）样品，分子量范围为四倍，证明了在四氢呋喃（THF）和甲基异戊基酮（MIAK）的混合物中，溶剂彼此等折射。摩尔质量平均值和分散度被证明与溶剂组成在统计学上无关，并且与纯 THF 中的值非常吻合。实验还通过使用在线和离线准弹性光散射以及离线 MALS 和 DRI 来研究溶剂组成对聚合物在溶液中的尺寸以及稀溶液热力学的影响。此外，使用 1 H 核磁共振波谱研究了立构规整度对 PMMA100 在 100% MIAK 中不溶性的影响。我们相信，这是首次通过 SEC/MALS/DRI 使用混合溶剂获得聚合物准确摩尔质量的实例。还注意到这些实验对其他形式的高分子液相色谱分离的价值。
