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Two-dimensional reversed-phase liquid chromatography using two monolithic silica C18 columns and different mobile
Tohru Ikegami1, Takeshi Hara, Hiroshi Kimura
1Department of Polymer Science and Engineering, Kyoto Institute of Technology, Matsugasaki, Kyoto 606-8585, Japan. ikegami@kit.ac.jp
Journal of Chromatography. A
|December 14, 2005
Summary
Comprehensive two-dimensional (2D) High-Performance Liquid Chromatography (HPLC) using C18 silica monolith columns achieved high peak capacity for aromatic compounds. This advanced separation technique offers improved efficiency and shorter run times, especially with gradient elution.
Area of Science:
- Analytical Chemistry
- Chromatography
- Separation Science
Background:
- Two-dimensional High-Performance Liquid Chromatography (2D-HPLC) enhances separation power.
- Reversed-phase liquid chromatography (RPLC) is a common separation mode.
- Silica monolith columns offer advantages in flow properties and efficiency.
Purpose of the Study:
- To develop and demonstrate a comprehensive 2D-HPLC system for enhanced separation of aromatic compounds.
- To evaluate the performance of C18 silica monolith columns in a 2D-RPLC setup.
- To compare isocratic and gradient elution modes for 2D-HPLC separation efficiency and speed.
Main Methods:
- Utilized a 2D-HPLC system with C18 silica monolith columns (10 cm x 4.6mm I.D. for 1st-D, 5 cm x 4.6mm I.D. for 2nd-D).
- Employed water/tetrahydrofuran (THF) mobile phase for the first dimension (1st-D) and water/methanol (CH3OH) for the second dimension (2nd-D).
- Fractionated sample from 1st-D directly to the 2nd-D injection loop, with fractionation/separation times of 45 or 60s.
- Investigated both isocratic and gradient elution modes for separation.
Main Results:
- Achieved a total peak capacity up to 900 in approximately 60 minutes using the isocratic mode for aromatic compounds.
- Gradient elution in both dimensions resulted in shorter separation times and superior separation efficiencies compared to the isocratic mode.
- Demonstrated that 2D-HPLC systems with C18 stationary phases and different mobile phase organic modifiers yield high peak capacity.
- Observed different selectivities due to variations in polar interactions between solutes and mobile phase modifiers.
Conclusions:
- Comprehensive 2D-HPLC using C18 silica monolith columns is a powerful technique for achieving high peak capacity.
- The system effectively separates aromatic compounds, with gradient elution offering significant advantages in speed and efficiency.
- Employing different organic modifiers in each dimension provides orthogonal selectivity, enhancing overall separation performance.
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