Chromatography Forum

Hi, maybe someone can tell me how this is possible...a friend in an other company is developing a UPLC method for an API. I know she works with Waters Aquity Systems with a PDA as detector, and today came a weird question. She asked how it is posible to have different absorption values but still the same concentration?

She says the concentrations are correct and there is no significant RT or lambda max shifting. Peak symmetry is constant. I dont know if this is enough information but i dont have more, maybe some one can explain it?

Lots of possible reasons (though we do not have enough information to narrow it down). (1) Different detectors may have been used (different models). (2) Different flow cell path-lengths or volumes. (3) Use of different wavelength bandwidths (larger bandwidth gives less selectivity but more signal). (4) Incorrectly using the Reference Wavelength software feature on the Waters PDA.

In addition, (as I recall) the injection volume of a UPLC is ~1 uL while HPLC can be ~100 uL. The smaller particle size of a UPLC packing (<2 um) will give the analyst a sharper and higher peak with a lower LOQ.