Compounds were obtained from silica gel column in very small quantity which could not be separated by prep tlc or on another column because one is not sure of getting anything at the end
You haven't given us very much information. Can you scale up your experiment to increase the amount of material you are after. Is degradation (maybe oxidation?) a problem. Would adding an anti-oxidant such as BHT help?
How do you know the two compounds are isomers? Chromatography can't tell you. What additional information do you have - NMR, MS, IR, UV . . . ?
With respect to the chromatography you will get better resolution if you can get access to an HPLC or, better still, a UPLC. Is this possible? Have you tried HPTLC?
I have a vary similar problem. I was fractionating an essential oil distilled form S. officinalis on 2D thin layer chromatography and I did a column chromatography. I did GS-MS before this so I have a reference about the oil composition. Looking at the thujone content of my sample I expected 2 big stains on 2D crom. for alpha and beta thujone but I got 4. Doing the same for column chromatography I didn't got a good separation. Currently my experiment because is paused because I have no time at the moment. Im curious how you will solve this problem.