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Effect of anionic additive type on ion pair formation constants of basic pharmaceuticals
Jun Dai1, Shaun D Mendonsa, Michael T Bowser
1Department of Chemistry, University of Minnesota, Minneapolis, MN 55455-0431, USA.
Journal of Chromatography. A
|April 16, 2005
Summary
Anionic additives in reversed-phase chromatography (RPLC) and LC/MS form ion pairs with basic drugs, influencing separation. This study confirms ion pair formation and quantifies its extent, revealing PF6- > ClO4- > CF3COO- > Cl- as the order of effectiveness.
Area of Science:
- Analytical Chemistry
- Separation Science
- Chromatography
Background:
- Mobile phase anionic additives like formate, chloride, and trifluoroacetate are increasingly used in RPLC and LC/MS for improved chromatography.
- Perchlorate is a common ion-pairing agent for peptide separation in RPLC.
- While ion pair formation between anions and cationic analytes is hypothesized to explain their effects, direct evidence has been lacking.
Purpose of the Study:
- To independently verify ion pair formation between basic drugs and anionic additives under RPLC conditions.
- To quantify the extent of ion pairing and understand its influence on chromatography and electrospray ionization.
- To establish a quantitative order of ion pair formation for common anionic additives.
Main Methods:
- Studied ion pair formation using capillary electrophoresis (CE) by measuring the effect of anionic additives on the electrophoretic mobility of probe drugs.
- Investigated ion pair formation under buffered acetonitrile-water solvent conditions typical for RPLC.
- Measured ion pair formation constants (Kip) for various anionic additives including hexafluorophosphate (PF6-), perchlorate (ClO4-), trifluoroacetate (CF3COO-), and chloride (Cl-).
Main Results:
- Confirmed ion pair formation between basic drugs and anionic additives under RPLC-relevant conditions for the first time.
- Established a quantitative order for ion pair formation constants: PF6- > ClO4- > CF3COO- > Cl-.
- Demonstrated that the extent of ion pairing is generally not large, with percentages of analytes as ion pairs at 20mM additive concentration being approximately 15% (PF6-), 6% (ClO4-), and 3% (CF3COO-), and even smaller for Cl-.
Conclusions:
- Anionic additives in RPLC do form ion pairs with basic drugs, confirming the long-standing hypothesis.
- The extent of ion pairing varies significantly among different anions, with PF6- being the most effective and Cl- the least.
- While ion pairing occurs, its contribution to the overall analyte retention and ionization may be limited at typical concentrations, necessitating further investigation into other influencing mechanisms.
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