method development

[ajey]

what is the procedure to develop a method (formulations) having more than one pKa or containing acidic, basic and neutral analyte?
What will be the pH of buffer solution, if the mixture contains analyte having pKa (consider analyte A having pKa=2.5, analyte B having pKa=5 and analyte C having pKa=7.5)?
Regards,
Ajay Mhatre

[Uwe Neue]

You can use any pH, but pay attention to the reproducibility of the buffer preparation.

Alternatively, you can use pH 10 with ammoniumbicarbonate and a column that is suitable at this pH.

[praveenpaliwal]

U can select the moderate pH near to 3.5-4.0 and used a C-18 column with high carbon loading . Let me know that what is ur compounds structure's

[mohan_2008]

I will start at the pH = 2.0 using a Zorbax SB-C18 column. Make your mobile phase choice with 0.1% TFA which will take you to that pH.
At the low pH, practically all the silanols will remain protonated and you have an additional advantage of a good peak shape.
All the bases (as mentioned) will elute faster at this lower pH, and hence peak tailing with column migration will be less.

Other option is to try out at a high pH. I will avoid this option if I have a choice (Silica columns cannot stand the high pH conditions).

If option one & two dont work, start on an ion-pair method. This being extremely good for highly polar compounds can additional impart selectivity to your separation.