Department of Chemistry, University of California, 501 Big Springs Rd., Riverside, CA 92521, USA.
J Chromatogr A. 2010 Jan 22;1217(4):479-88. doi: 10.1016/j.chroma.2009.11.064. Epub 2009 Dec 3.
Reverse-phase ion-pair high performance liquid chromatography (RPIP-HPLC) and ultra-performance liquid chromatography (RPIP-UPLC) are increasingly popular chromatographic techniques for the separation of organic compounds. However, the fine details of the RPIP separation mechanism are still being debated. Many factors including type and concentration of the ion-pairing reagent, mobile phase pH, organic modifier, ionic strength, and stationary phase all play a role in the overall efficiency and optimization of ion-pairing separations. This study investigates the role that competition between ion-pairing reagents with different steric bulk and hydrophobicity plays in the separation of structural isomers of heparin and heparan sulfate (HS) disaccharides. In addition to providing insights into the mechanism by which RPIP-HPLC can resolve closely related disaccharides, the use of competition between ion-pairing agents could lead to new methods for the separation of larger heparin and HS oligosaccharides. This approach should also be applicable to the analysis of other compound classes, and could lead to a general approach for the chromatographic resolution of mixtures of charged analytes having similar structures.
反相离子对高效液相色谱(RPIP-HPLC)和超高效液相色谱(RPIP-UPLC)是越来越受欢迎的用于分离有机化合物的色谱技术。然而,RPIP 分离机制的细节仍在争论中。许多因素,包括离子对试剂的类型和浓度、流动相 pH 值、有机溶剂、离子强度和固定相,都在整体效率和离子对分离的优化中发挥作用。本研究探讨了具有不同空间位阻和疏水性的离子对试剂之间的竞争在肝素和硫酸乙酰肝素(HS)二糖结构异构体分离中的作用。除了深入了解 RPIP-HPLC 可以解析密切相关的二糖的机制外,离子对试剂之间的竞争的使用可能会导致分离更大的肝素和 HS 寡糖的新方法。这种方法也应该适用于其他化合物类别的分析,并可能导致用于具有相似结构的带电分析物混合物的色谱分辨率的一般方法。