Use of Coloumn in HPLC - (Mar/05/2008 )
Hi, I am a JRF at Avesthagen, Bangalore, India. I had used hydroxylapatite column for separation of plant alkaloids but got very little eluate (0.012ng). I had run for 30min. and coloumn length was standard. I had used ethanol 95% as a solvent. Is there any other coloumn that will produce a greater yield for plant alkaloids?
What standards did you use and did they come through the column ok?
Alkaloids are such a diverse group of molecules you may need to modify your approach to isolatate the fractions. A C18 column in reverse phase mode, 150mm x 4.6 should be fine but do check your solvent. You could try acetonitrile (25%) in 75% water. Also add some TEA (0.027M) to unhibit interaction with the column.
What detection system are you using? UV 280 would probably be fine.