During extraction of fungal secondary metabolites using methylene chloride from culture filtrate, the methylene chloride layer forms lipids and fats that contaminate the extraction process. I need to know how to purify the filtrate from these lipids prior to extraction. I have already used n-hexane as a defatting agent three times but I still found the fats.
try this trick, I have been using for years to separate analyte from the fatty matrix. You shake fat with hexane to dissolve anlayte and fat then shake the hexane layer with acetonitrile. Fat has poor solubility in acetonitrile (~ 1-1.5 %) and your anlayte which should be polar or moderately non-polar will go to acetonitrile. If you stil need to get rid of fat residue in acetonirile, throw some C18 SPE material in the acetonitrile, it will absorb more fat. Good luck
First of all try to identify the lipids using MS. Once you know the lipids, try to identify the mixture of solvents (one of which has to be methylene chloride) which will show you the difference of partition coefficient of those lipids. Also know the partition coefficients of the metabolites too and then by using logic select a combination of liquid which will suit you best. Alternatively, freeze the mixture of metabolites and lipids in methylene chloride and slowly thaw it so that the lipids separate out leaving you with very little contamination. Lastly if you can get a hold of Preparative HPLC, nothing like it. good luck.
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