Topic: HPLC for Phospholipid and Fatty Acid Analysis (Read 9322 times)

DeltaForce · « **on:** April 20, 2006, 03:38:16 PM »

Hello

I'm just working through my revision and wondered why can't HPLC be used to analyse lipids and fatty acids? Apparently there are some "major limitations" that need to be overcome before we can use HPLC as an analytical technique for small amounts of fatty acids and lipids, but for the life of me I can't think or find out why

Please *delete me*

DeltaForce

Yggdrasil · « **Reply #1 on:** April 20, 2006, 09:01:11 PM »

Well, fatty acids and other lipids aren't water soluble, and as far as I know, most HPLC systems are designed to use aqueous solutions as the mobile phase.

DeltaForce · « **Reply #2 on:** April 22, 2006, 08:18:00 AM »

So the solution would be to use a solvent such as Methanol for mobile phase?? All the solvents I've ever used have been "SOLVENT:Water" 50:50.

Yggdrasil · « **Reply #3 on:** April 22, 2006, 03:02:00 PM »

I'm not an expert on fatty acid analysis, but I don't think a 50:50 water:methanol solution would be able to even solublize most fatty acids. Most of the time, whenever I've seen chromatographic analysis of fatty acids, its been a GC of methyl esters of fatty acids. However, I will reiterate that I'm not an expert on this topic and I may be wrong.

waleed · « **Reply #4 on:** May 28, 2006, 10:57:33 AM »

well.. i think u can find many work based upon sepration of fatty acids using HPLC,,, but u hve to remember that you have polar and non-polar compounds,,, most recent studies use n-hexane - propane-2-ol and water. or acetonitrile methanol ..water .. as a mobile phase... but u hve to controle the conditions... column , detectors mainly...
ok?
about myself i ddnt get a method for separation of phospholipids using HPLC,,, cz it have many things to be affected with... so if u can help me too to provide me a method i can use .. i will be thankful..

GSK · « **Reply #5 on:** October 01, 2006, 06:58:06 PM »

you can absolutely analyze fatty acids with RP-HPLC. Detect at 205-210nm and use MeOH/water/AcOH or ACN/water/TFA as the mobile phase, for simple fatty acids like arachidonic, EPA, DHA, etc. For oxylipins and other derivatives, you can adjust the monitoring wavelength or the phase (NP with IPA/Hx/AcOH)

chiralic · « **Reply #6 on:** October 05, 2006, 11:56:40 PM »

Hello DeltaForce:

You can find on internet a lot papers about separation and quantitation of Lipids (mainly Phospholipds) using HPLC (Nornal mode and Reversed-Phase mode). For example: phospatidylcholine, phosphatidylethanolamine, phosphatidylinositol, phosphatidic acid, phosphatidylserine, shingomyelim, cerebrosides and cardiolipin...these separations is based on differences in chain length and of course on degree of unsaturation. Also you can find many separations on differents HPLC detectors.

Of course, the succefull separation and quantitation of Lipids depend in part on the complexity of your sample and the HPLC equipment ("type and conditions") available in your lab.

Now is better use Reversed-phase HPLC over normal HPLC because your can avoid use large amounts of samples needed for analysis, peak broadening and tailing, long analysis time, low detection sensitivity, working pH range limited to 2-7,and volatile toxic organic solvents.

If you need more information or details, please send me a private message...

Regards,

Chiralic