Ion-exchange chromatography systems have previously been utilized in HPLC analysis of ionic samples. Recently, reversed phase partition chromatography using ion-pair reagents has been developed and utilized. The ionic samples form an ion-pair with ion-pair reagents in the mobile phase to become electrically neutral. The increase in hydrophobic character of the ion-pair results in a greater affinity for the reverse stationary phase and leads to sample resolution.
UV and fluorescence detectors are widely used. Therefore ion-pair reagents must lack UV absorption and fluorescence themselves to obtain highly sensitive detection of samples. The UV absorption of sodium alkanesulfonates and quaternary ammonium salts is minimal so that these reagents can be used for reliable HPLC analysis. On the other hand, when a sample lacks sufficient UV absorption or fluorescence, the use of sodium 9,10-dimethoxyanthracene-2-sulfonate allows for high-sensitivity detection as a fluorimetric ion-pair reagent.
Recently, use of LC/MS in which mass spectrometry is incorporated in HPLC as a detector has become widespread. Sodium alkanesulfonates, a general ion-pair reagent, being non-volatile crystals pose a problem in that they contaminate the interface. The IPC-PFFA series is made of highly volatile ion-pair reagents allowing for continuous LC/MS analysis without contaminating the interfaces.
• Ion-Pair Chromatography for Acidic Samples
¶Analysis is performed with pH adjusted to 7.5 with the addition of quaternary ammonium salts to the mobile phase.
¶Acidic samples form an electrically neutral ion-pair with the quaternary ammonium salt and are retained in the reverse phase systems.
—Examples—
1. When 0.5mol/L Tetrabutylammonium Phosphate is used:
The reagent (10ml) is diluted to 1L with an aqueous solvent such as methanol - water. (pH adjustment is not required because the reagent is already buffered.)
2. When Tetrabutylammonium Hydroxide is to be used:
1) The reagent (12.5ml) is diluted to 1L with an aqueous solvent such as methanol - water.
2) The pH is adjusted to 7.5 by the addition of an aqueous phosphoric acid (10%).
• Ion-Pair Chromatography for Basic Samples
¶Analysis is performed by the addition of sodium alkanesulfonate to the mobile phase.
¶The basic samples form an electrically neutral ion-pair with sodium alkanesulfonate and are retained in the reverse phase system.
¶In the case of sodium alkanesulfonate, the greater the number of carbons in the alkyl group, the greater the partition ratio.
—Examples—
1) Sodium 1-Heptanesulfonate 1.011g (0.005mol) is weighed out.
2) The reagent is dissolved in 1L of an aqueous solvent such as methanol - water.
3) The pH is adjusted to 3.5 by the addition of aqueous phosphoric acid (50%).
Acidic Ion-Pair Reagents for HPLC
Acidic Ion-Pair Reagents for LC/MS
Basic Ion-Pair Reagents for HPLC
Basic Ion-Pair Reagents for LC/MS
Fluorimetric Ion-Pair Reagent for HPLC
Acidic Ion-Pair Reagents for HPLC
R4-N+ X-
I0453
|
I0365
|
I0366
|
I0368
|
I0364
|
I0367
|
I0363
|
Acidic Ion-Pair Reagents for LC/MS
R2-N+H2 X-
A5704
|
A5702
|
A5705
|
A5703
|
Basic Ion-Pair Reagents for HPLC
R-SO-3 Na+
I0342
|
I0348
|
I0350
|
I0345
|
I0344
|
I0347
|
I0346
|
I0343
|
I0341
|
I0351
|
I0349
|
I0352
|
Basic Ion-Pair Reagents for LC/MS
Rf-COOH
A5713
|
A5714
|
A5717
|
A5712
|
A5716
|
A5721
|
A5711
|
A5715
|
A5722
|
Fluorimetric Ion-Pair Reagent for HPLC
A5701
|
| A5701 | Sodium 9,10-Dimethoxyanthracene-2-sulfonate [Fluorimetric Ion-Pair Reagent for Amines] |
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