HSS HPLC Columns: An Additional Option for Transferring Methods between HPLC and UPLC Technology
The Application Notebook
As UPLC users convert or replace their existing HPLC systems with UPLC systems there is a transition period where a method must be run on both platforms. Thus, having the same particle substrate and bonded phases available in HPLC and UPLC particle sizes can significantly ease the burden of method development and transfer from one platform to another. In addition to the ethylene bridged hybrid (BEH) particle, three new high strength silica (HSS) stationary phases for HPLC applications are introduced. Scalability between both column diameter and particle size is demonstrated on both UPLC and HPLC instrumentation.
Zhe Yin, Kenneth J. Fountain, Doug McCabe and Diane M. Diehl, Waters Corporation, Milford, Massachusetts, USA.
Introduction
As UPLC users convert or replace their existing HPLC systems with UPLC systems there is a transition period where a method must be run on both platforms. Thus, having the same particle substrate and bonded phases available in HPLC and UPLC particle sizes can significantly ease the burden of method development and transfer from one platform to another. In addition to the ethylene bridged hybrid (BEH) particle, three new high strength silica (HSS) stationary phases for HPLC applications are introduced. Scalability between both column diameter and particle size is demonstrated on both UPLC and HPLC instrumentation.
Experimental
Systems: UPLC — ACQUITY UPLC and PDA HPLC — Alliance 2695 and 2998 PDA
Columns (Figure 1): ACQUITY UPLC HSS C18 SB
Columns (Figure 2): ACQUITY UPLC HSS C18, 3.5 µm,
Mobile phase A: 0.1% TFA in H2O
Mobile phase B: 0.1% TFA in MeCN
Results and Discussion
Figure 1 shows the scalability between UPLC and HPLC columns for the separation of paroxetine-related compounds on HSS C18 SB. Gradient, flow-rate and injection volume were properly scaled between different particle sizes and column dimensions. The system volume was properly compensated for when transferring method between UPLC and HPLC instrumentations. Chromatographic resolution and selectivity were maintained across all separation platforms.
Figure 1
Figure 2 demonstrates the scalability between columns with different internal diameters for the separation of a forced degradation sample of glimepiride.
Figure 2
Conclusions
A successful method transfer requires careful consideration of key parameters including system volume, column dimension and particle size, injection volume and gradient profile. Methods can be easily transferred between different systems (UPLC → HPLC → prep), particle sizes and column diameters on the new HSS columns.
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