University of Leuven (KU Leuven), Department for Pharmaceutical and Pharmacological Sciences, Pharmaceutical Analysis, Herestraat 49, Leuven, Belgium.
University of Leuven (KU Leuven), Department for Pharmaceutical and Pharmacological Sciences, Pharmaceutical Analysis, Herestraat 49, Leuven, Belgium.
J Chromatogr A. 2021 Jan 25;1637:461852. doi: 10.1016/j.chroma.2020.461852. Epub 2020 Dec 27.
In an ongoing effort to better understand the underlying mechanisms of band broadening in particle-packed reversed-phase liquid chromatography columns, new models for intra-particle diffusion, representing an adsorption- and partition-type retention behavior, are proposed. These models assume the mesoporous zone inside the particles is subdivided in four distinct regions: a fraction f filled with bulk mobile phase, a fraction f enriched in pure organic modifier extending outside the stationary phase layer, a fraction f comprising the liquid surrounding the alkyl chains and a fraction f consisting of the stationary phase alkyl chains. Intra-particle diffusion is calculated as a residence time weighted average of the diffusion in these different regions. Experimental procedures and models are proposed to determine the volumes of these four regions and applied to three reversed-phase liquid chromatography columns with different pore sizes (80 Å versus 300 Å) and different stationary phase types (C versus C). The newly proposed models are then applied to predict the intra-particle diffusion of butyrophenone across a wide range of retention factors (1 ≤ k" ≤ 40) in each of these columns. These predictions are compared to experimental data that are extracted from the effective diffusion coefficients of butyrophenone obtained via peak parking experiments. It is demonstrated that both adsorption- and partition-type models for intra-particle diffusion model the actual behavior of the test compound well, and require the determination of only one (partition) or two (adsorption) fitting factors: the obstruction to free movement the analytes experience from the alkyl chains in the retained state (partition and adsorption) and in the unretained state (adsorption). Finally, it is demonstrated that the major contributor to the intra-particle diffusion of retained compounds (k" > 2) is the diffusion these analytes undergo when retained in the organic-modifier enriched zone surrounding the alkyl chains (partition model) or when adsorbed onto the alkyl chains (adsorption model), confirming that surface diffusion plays an important role in the mass transfer of retained compounds in reversed-phase liquid chromatography columns.
为了更好地理解颗粒填充反相液相色谱柱中展宽的基本机制，提出了新的颗粒内扩散模型，这些模型代表了吸附和分配型保留行为。这些模型假设颗粒内部的介孔区域分为四个不同的区域：一部分 f 充满了体相流动相，一部分 f 富含纯有机改性剂，延伸到固定相层之外，一部分 f 由围绕烷基链的液体组成，一部分 f 由固定相烷基链组成。颗粒内扩散被计算为这些不同区域扩散的停留时间加权平均值。提出了实验程序和模型来确定这四个区域的体积，并将其应用于三种具有不同孔径（80Å 与 300Å）和不同固定相类型（C 与 C）的反相液相色谱柱。然后，将新提出的模型应用于预测丁酰苯在这些柱子中的广泛保留因子（1≤k"≤40）范围内的颗粒内扩散。将这些预测与通过峰停车实验获得的丁酰苯有效扩散系数的实验数据进行比较。结果表明，颗粒内扩散的吸附和分配型模型都很好地模拟了测试化合物的实际行为，并且只需要确定一个（分配）或两个（吸附）拟合因子：保留状态下（分配和吸附）和未保留状态下（吸附）分析物从烷基链自由移动所受到的阻碍。最后，结果表明，保留化合物（k"＞2）颗粒内扩散的主要贡献是这些分析物在围绕烷基链的富含有机改性剂的区域中保留时（分配模型）或吸附到烷基链上时（吸附模型）所经历的扩散，这证实了表面扩散在反相液相色谱柱中保留化合物的传质中起着重要作用。
