School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Boulevard d'Yvoy 20, 1211 Geneva 4, Switzerland.
J Chromatogr A. 2013 Nov 1;1314:288-97. doi: 10.1016/j.chroma.2013.09.039. Epub 2013 Sep 14.
Recently, there has been a renewed interest in supercritical fluid chromatography (SFC), due to the introduction of state-of-the-art instruments and dedicated columns packed with small particles. However, the achievable kinetic performance and practical possibilities of such modern SFC instruments and columns has not been described in details until now. The goal of the present contribution was to provide some information about the optimal column dimensions (i.e. length, diameter and particle size) suitable for such state-of the-art systems, with respect to extra-column band broadening and system upper pressure limit. In addition, the reliability of the kinetic plot methodology, successfully applied in RPLC, was also evaluated under SFC conditions. Taking into account the system variance, measured at ∼85μL(2), on modern SFC instruments, a column of 3mm I.D. was ideally suited for the current technology, as the loss in efficiency remained reasonable (i.e. less than 10% decrease for k>6). Conversely, these systems struggle with 2.1mm I.D. columns (55% loss in N for k=5). Regarding particle size, columns packed with 5μm particles provided unexpectedly high minimum reduced plate height values (hmin=3.0-3.4), while the 3.5 and 1.7μm packing provided lower reduced plate heights hmin=2.2-2.4 and hmin=2.7-3.2, respectively. Considering the system upper pressure limit, it appears that columns packed with 1.7μm particles give the lowest analysis time for efficiencies up to 40,000-60,000 plates, if the mobile phase composition is in the range of 2-19% MeOH. The 3.5μm particles were attractive for higher efficiencies, particularly when the modifier percentage was above 20%, while 5μm was never kinetically relevant with modern SFC instruments, due to an obvious limitation in terms of upper flow rate value. The present work also confirms that the kinetic plot methodology could be successfully applied to SFC, without the need for isopycnic measurements, as the difference in plate count between predicted and experimental values obtained by coupling several columns in series (up to 400mm) was on average equal to 3-6% and with a maximum of 13%.
最近，由于引入了最先进的仪器和专门装填小颗粒的柱，人们对超临界流体色谱（SFC）重新产生了兴趣。然而，直到现在，这种现代 SFC 仪器和柱所能达到的动力学性能和实际可能性还没有详细描述。本研究的目的是提供一些有关最适合这种最先进系统的柱尺寸（即长度、直径和粒径）的信息，同时考虑柱外展宽和系统上限压力的影响。此外，还评估了在 SFC 条件下成功应用于 RPLC 的动力学图谱方法的可靠性。考虑到现代 SFC 仪器上测量的系统方差约为 85μL(2)，对于当前技术，3mm ID 的柱是理想的，因为效率的损失仍然合理（即 k>6 时效率降低不超过 10%）。相反，这些系统与 2.1mm ID 柱（k=5 时效率降低 55%）难以兼容。关于粒径，装填 5μm 颗粒的柱提供了出人意料的高最小板高值（hmin=3.0-3.4），而 3.5μm 和 1.7μm 装填的柱则提供了较低的最小板高值 hmin=2.2-2.4 和 hmin=2.7-3.2。考虑到系统上限压力，似乎装填 1.7μm 颗粒的柱在流动相组成在 2-19%甲醇范围内时，可以为效率高达 40,000-60,000 板的分析提供最低的分析时间。当改性剂百分比高于 20%时，3.5μm 颗粒对于更高的效率具有吸引力，而 5μm 颗粒在动力学上与现代 SFC 仪器始终不相关，因为其上限流速值存在明显的限制。本工作还证实，动力学图谱方法可以成功地应用于 SFC，而无需进行等密度测量，因为通过串联连接几个柱（最多 400mm）获得的预测值和实验值之间的板数差异平均等于 3-6%，最大为 13%。
