Chromatography Forum

Hi Everyone,
I just want to understand why normally we select the pH values less than its PKa value during any organic acid analysis while we can also select the pH value more than its PKa value but we don't prefer.
Is there any specific reason of column only

Depends on stationary phase.
Acids at low pH are (normaly) in their protonated form, therfore uncharged and show better retention on nonpolar surface.
If charged -> ionchromatography (ion exchange or ion exclusion)

And from 'older' days, silica based particles were not stable above pH ~8, so low pH was normaly used anyway

I concur with Hollow. Many decades ago HPLC columns were very fragile and ODS-silanol groups could be easily broken by alkaline mobile phases. Many molecules are ionized in the acidic form or alkaline form depending on its pKa (where it's 50/50). In order to achieve just 1 peak from 1 form it was decided to use only acidic conditions.

But 'newer' specialty columns you can uses alkaline conditions (if so specified by the manufacturer).

I will give you an example of what I said. Vitamin C comes in 2 ionic forms, the protonated Ascorbic Acid (the organic acid), and the deprotonated Ascorbate (its ionized anion, and also a chelate). Unless you adjust the pH of your mobile phase to >2 pKa units of the molecule you cannot eliminate their respective concentrations since they are interchangeable, and thus their resulting chromatographic peaks.

Proteins are even worse since every amino acid can be protonated or deprotonated (their organic acid moiety or/and amine moiety).