Posted 30 June 2009 - 01:09 AM
I am looking at analysing the Free fatty acid profile of cell culture media (DMEM) using a GC. From what I understand, the free fatty acids are not volatile under the 220 *C temperature of the GC so they must be derivitized to FAME's (Fatty acid methyl esters). The protocol I have uses large quantities of sample (150ml) where im looking at about 200 ul. This change in volume renders most of the protocol and its apparatus useless.
In brief: It suggests dissolving sample in 2:1 chloroform:methanol soln, removing supernatant, drying, derivitize using acetylCl, dry and then dissolve once more in chloroform for injection.
Does anybody have any good GC or lipid extraction resources for me to figure out the best method to use? most that I find are vague and very general ie fatty acids or lipids not free fatty acids specifically.
Any help at all here would greatly increase my confidence in moving forward.
Thanks in advance.
Posted 30 June 2009 - 05:53 AM
you can't figure out how to scale this down?
The protocol I have uses large quantities of sample (150ml) where im looking at about 200 ul. This change in volume renders most of the protocol and its apparatus useless.
where it says "ml" substitute "ul" and use smaller vessels (small glass tubes). there should be no problem as long as there is no distillation step.
Edited by mdfenko, 30 June 2009 - 05:56 AM.
genius does what it must
i do what i get paid to do