Chromatography Forum

hi, i'm trying to elute hexaminolevulinate HCl on C18 TLC with 9:1 MeOH:H2O and I am getting poor elution and streaking.

I need to stick with C18 stationary phase too. I thought about adding TEA to decrease column interaction and possibly get rid of streaking.

Any suggestions?

All X18s are not created equal, with older-technology ("Type A") columns giving much more tailing than newer columns ("Type B"). Tailing can often be improved by lowering the pH and making sure you have enough buffer (25 mM would be typical, but you can try going up to 50 or even 100 mM and see what happens. TEA is sort of a last resort, but it does work.

I would suggest that you use a sodium acetate buffer instead of water for your mobile phase. You might also wish to reduce the amount of methanol and add some ethyl acetate to the mobile phase to compensate for the increased amount of water.

I would adjust the pH of the buffer to acidic using acetic acid. Just an idea.

best wishes,

Rod

Thanks guys. What worked was buffering the aqueous.

I used pH 4.75 acetate.

MestizoJoe

Scary how you read my mind.

But of course, you had the right idea on your own.

Do you understand WHY that worked?

Please share your analysis of the problem if you have the time.

best wishes,

Rod

I think it's that without buffer the salt may be broken on the stationary phase and the free base form of the compound is much more non polar. amines in hplc also tend to tail which in talc would manifest itself as streaking.

Regards

Ramon