Department of Pharmaceutical Chemistry, University of Pavia, Pavia, Italy.
J Sep Sci. 2010 Mar;33(6-7):803-16. doi: 10.1002/jssc.200900672.
The separation of 12 model compounds chosen among purine and pyrimidine bases and nucleosides was studied by using hydrophilic interaction chromatography (HILIC). The compounds investigated were small molecules with relevant properties for biomedical and pharmaceutical studies. The mixture of pyrimidines and purines was applied on a ZIC-HILIC 150 x 2.1 mm, 5 microm, and two TSKgel Amide-80 150 x 2.0 mm, 5 microm and 3 microm particle size columns. The retention of the analytes was studied by varying ACN%, ammonium formate concentration, pH, and column temperature. The results obtained confirmed the elution order of nucleobases, nucleosides, and nucleotides based on their hydrophobicity. The retention mechanism of the columns was studied considering the models used for describing partitioning and surface adsorption. The influence on retention of chromatographic conditions (ACN%, salt concentration, pH, and temperature) was described and discussed for both columns. The optimization of the conditions studied allowed to assess a gradient method for the separation of the 12 analytes. The developed method is a valuable alternative to existing methods for the separation of the compounds concerned.
选用嘌呤和嘧啶碱基及核苷中的 12 种模型化合物，通过亲水作用色谱（HILIC）对其进行分离。所研究的化合物为具有生物医学和药物研究相关特性的小分子。将嘧啶和嘌呤混合物应用于 ZIC-HILIC 150×2.1mm，5μm 和 TSKgel Amide-80 150×2.0mm，5μm 和 3μm 粒径的两根色谱柱上。通过改变 ACN%、甲酸铵浓度、pH 值和柱温来研究分析物的保留情况。结果证实了基于疏水性的碱基、核苷和核苷酸的洗脱顺序。考虑到用于描述分配和表面吸附的模型，研究了柱的保留机制。描述并讨论了色谱条件（ACN%、盐浓度、pH 值和温度）对保留的影响。对研究条件的优化允许评估用于分离 12 种分析物的梯度方法。所开发的方法是现有分离相关化合物方法的一种有价值的替代方法。
