HPLC Analysis of Basic Drugs and Acidic Counter-Ions by Mixed-Mode Chromatography

In pharmaceutical formulations, the majority of drugs are delivered in salt form, often involving combinations of organic or inorganic cations and anions. Because these counter-ions can vary in nature—such as inorganic cations paired with organic acids, inorganic anions with organic bases, or organic cations with organic anions—their analysis typically requires multiple HPLC methods due to differing chemical behaviors and stoichiometry.

Using mixed-mode chromatography on the Zodiac HST RP column enables simultaneous quantitation of both the active pharmaceutical ingredients (APIs) and their counter-ions within a single run. This is particularly useful for hydrophobic basic drugs such as dextromethorphan, verapamil, and trimipramine, along with their acidic counter-ions like chloride, chlorate, bromide, bromate, perchlorate, maleate, fumarate, tartrate, succinate, phosphate, citrate, benzenesulfonate, and toluenesulfonate.

In this method:

• Hydrophobic basic drugs are retained primarily through the reversed-phase mechanism.

• Acidic counter-ions are retained via a combination of anion-exchange and reversed-phase interactions.

• Highly polar counter-ions are retained mainly by the anion-exchange mechanism alone.

Retention time and selectivity can be finely tuned by adjusting the acetonitrile concentration and ionic strength of the mobile phase. Depending on the UV absorbance properties of the counter-ions, ELSD (Evaporative Light Scattering Detection) can be used—especially when the counter-ion forms a non-volatile salt with additives like ammonium formate.

This versatile HPLC approach is ideal for the simultaneous quantification of a wide range of basic drugs and their counter-ions, offering independent control of retention and even the ability to alter elution order when needed—thanks to the dual retention mechanisms of the Zodiac HST RP column.