Gritti Fabrice, Alden Bonnie A, McLaughlin Justin, Walter Thomas H
Waters Corporation, Instrument/Core Research/Fundamentals, and Chemistry R & D 34 Maple Street, Milford, MA, 01757, USA.
Waters Corporation, Instrument/Core Research/Fundamentals, and Chemistry R & D 34 Maple Street, Milford, MA, 01757, USA.
J Chromatogr A. 2023 Mar 15;1692:463828. doi: 10.1016/j.chroma.2023.463828. Epub 2023 Feb 4.
This work investigates the link between the retentivity and the stationary phase to mobile phase mass transfer resistance of hydrophilic interaction liquid chromatography (HILIC) columns packed with the same base ethylene-bridged hybrid particles (BEH). The retention volumes, the plate heights, and the volume of the adsorbed water layer were measured for the ACQUITY UPLC BEH 130 Å HILIC Column (unbonded BEH), ACQUITY UPLC BEH 130 Å Amide Column (amide group attached), and Atlantis Premier BEH 95 Å Z-HILIC (zwitterionic group attached) Column. The method of Guo (toluene retention volumes in pure acetonitrile and in the HILIC eluent) was validated from the UNIFAC group-contribution method and applied to measure accurately the water layer volumes in these columns. A strong correlation was found between the retention volumes of most neutral polar analytes and the volume of the water layer adsorbed in the HILIC column. The fraction of the pore volume occupied by the water layer increases significantly from the BEH HILIC Column to the BEH Amide Column, and to the BEH Z-HILIC Column. This is explained by the water solvation of the attached ligands in the pore volume of the BEH Particles and to the smaller average mesopore size of the BEH Z-HILIC Particles. A second and strong correlation is also observed between the water content in the HILIC particle and the stationary phase to mobile phase mass transfer resistance of the HILIC columns at high mobile phase linear velocities. The measured intra-particle diffusivity normalized to the bulk diffusion coefficient decreased from 0.33 (BEH HILIC Column) to 0.10 (BEH Amide Column) and to only 0.03 (BEH Z-HILIC Column) for comparable retention of cytosine. These results are fully consistent with the higher viscosity of the internal eluent (higher water content) and higher internal obstruction for diffusion (smaller mesopores and internal porosity) in the BEH Z-HILIC Particles. Still, in gradient elution mode, the peak capacity was found to be 18% higher for the BEH Z-HILIC Column than that on the BEH Amide Column because the retention factors at elution were smaller when maintaining the same analysis time and starting eluent composition.
本研究探讨了填充相同基础乙烯桥连杂化颗粒（BEH）的亲水作用液相色谱（HILIC）柱的保留性与固定相到流动相传质阻力之间的联系。对ACQUITY UPLC BEH 130 Å HILIC柱（未键合BEH）、ACQUITY UPLC BEH 130 Å酰胺柱（连接酰胺基团）和Atlantis Premier BEH 95 Å Z-HILIC（连接两性离子基团）柱测量了保留体积、塔板高度和吸附水层体积。Guo的方法（纯乙腈和HILIC洗脱液中的甲苯保留体积）通过UNIFAC基团贡献法得到验证，并用于准确测量这些柱中的水层体积。发现大多数中性极性分析物的保留体积与HILIC柱中吸附的水层体积之间存在很强的相关性。从BEH HILIC柱到BEH酰胺柱，再到BEH Z-HILIC柱，水层占据的孔体积分数显著增加。这可以通过BEH颗粒孔体积中连接配体的水合作用以及BEH Z-HILIC颗粒较小的平均中孔尺寸来解释。在高流动相线速度下，还观察到HILIC颗粒中的含水量与HILIC柱的固定相到流动相传质阻力之间存在第二个强相关性。对于可比的胞嘧啶保留，测量得到的归一化到本体扩散系数的颗粒内扩散率从0.33（BEH HILIC柱）降至0.10（BEH酰胺柱），仅为0.03（BEH Z-HILIC柱）。这些结果与BEH Z-HILIC颗粒中内部洗脱液的较高粘度（较高含水量）和较高的内部扩散阻碍（较小的中孔和内部孔隙率）完全一致。然而，在梯度洗脱模式下，发现BEH Z-HILIC柱的峰容量比BEH酰胺柱高18%，因为在保持相同分析时间和起始洗脱液组成时，洗脱时的保留因子较小。
