Hi,
I need to analyze the bicyclic compounds which do not have any chrmophore group. The compound contains 5 member ring with 1 nitrogen,(pyrrolidine) and 6 member ring with double bond or with OH. I have been trying ODS column, UV 220 nm, flow rate 1.0 mL/min, temp. 40C, mobile phase is CH3CN/ 0.1% H3PO4, using gradient method. The base line is curving and the product is eluted right on the portion where the baseline is shift upward!
I am not familiar and specialised in HPLC therefore, would like to know if it is normal for such curvy baseline observed in gradient method? and which method or column should I try to analyses these types of compounds?
thank you in advance.

The curvy baseline indicates that the mobile phase you are using is contaminated. In that case this curving will usually start when you increase the concentration of mobile phase.

As mentioned previously, contaminated acetonitrile could be a problem. Are you using HPLC grade acetonitrile? Another factor may be the sensitivity range you are using. I find that the baseline using a methanol: water gradient (0.1%TFA in both solvents) rises quite steeply when I'm analysing low amounts of analyte, with peak heights <1mAU.
I realise that you may be stuck with the equipment you currently have, but it would be worth trying an ELSD (Evaporative Light Scattering detector)if you can - we have trialled several on samples without UV chromaphores (pyrrolizidine alkaloids) and have been impressed with the results. They seem to be good, non selective detectors and cope happily with gradient elution - great to use for method development for our LC-MS.

Hi Neil,
thank you very much for your advice. I always use HPLC grade solvent.
I do not know anything about ELSD so I will look on internet to inform myself about such detector.I wonder how easy and how expensive to get such detector?
I might ask more questions later concerning ELSD if you do not mind.
Thank you very much again.
CTP