it is essential to give at least the compounds you are working with, otherwise you are wasting other peoples time as well as your own. there is thousands of antibiotics and all have different issues - a lot need pH adjustment.
also you should give the amount you inject on column as well as the injection volume
I agree with Kai, you don't give us enough information i.e. structure, physical property, and chromatogram to diagnose. It is like you tell me my car would not start, what is the fix. I need more information.