Chromatography Forum

I run amino acid separation with buffer A: 50 mm sodium acetate (pH 5.45); buffer B: 70% acetonitrile containg 32 mm sodium phosphate (pH 6.1).

Last week, everything worked allright and I got good separation.

This week, I remade buffer A and adjusted pH meter. HPLC baseline drift upward with increased buffer B. Separation still is good , but with upward baseline. Is this due to pH adjustment for Buffer A?

Buffer A run a straigt baseline if no buffer B.


Please give me suggestions!! Thank you!

Dear Zhi Ju,

1 - You didn´t write if you are using gradient or isocratic procedure on both weeks.

2 - Have you been performing your HPLC analysis with UV-Vis detector? What is the wavelength?

3 - Did you add the same acid solution to adjust the Buffer A pH?

4 - Are you sure that your Buffer B remaind good on this week? (Remember that Phosphates in Acetonitrile can floculate or microfloculate with considerable probability, when you introduce this Buffer on your analysis can you verify any pressure value increment?).

I are wait your comments.

Take care and Have Good Elutions!!

Carlos Teixeira
teixeiracs@yahoo.com