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How to dissolve unsaturated fatty acids - (Jan/04/2008 )

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Hello all,

I am trying to do gene expression and functional assay of Fatty acyl reducatse (FAR) in transfectd SF9 cells in SF900 medium. My objective is to determine FAR gene expression by checking whether it converts E/Z11-14: COOH to E/Z11-14:OH. My problem is I can’t able to dissolve the E/Z11-14: COOH in SF900 SFM medium. E/Z11-14: COOH is originally dissolved in hexane. For functional assay, at first I tried with E/Z11-14: COOH in Hx directly put in the 60 mm plate and after evaporating the Hx in fume hood, trasfected SF9 cells were plated and incubated. But this would not work out. Later I tried to dissolve E/Z11-14: COOH in 0.9 % NaCl and 10 % BSA, but it is not fully dissolving.

Your valuable suggestions are welcome.

Thanks

-Binu-

whats the chain length of the FA?

-genehunter-1-

some of these might take time to rehydrate so give it 30 - 45 min. You may have to warm it up a little to completely dissolve it.

good Luck !!!

-scolix-

Try its salt form with KOH, or some mild detergents like Cremophor EL.

-genehunter-1-

QUOTE (genehunter-1 @ Jan 5 2008, 02:39 AM)
whats the chain length of the FA?


Its C14 (Myristic acid) derivatives

-Binu-

QUOTE (scolix @ Jan 5 2008, 05:30 AM)
some of these might take time to rehydrate so give it 30 - 45 min. You may have to warm it up a little to completely dissolve it.

good Luck !!!


I tried to dissolve the compound in 0.9% NaCl by vortexing & stirring, later heating at 80oC for 10 min. Still the solution looks think.
any other suggestions?

-Binu-

QUOTE (genehunter-1 @ Jan 5 2008, 08:20 AM)
Try its salt form with KOH, or some mild detergents like Cremophor EL.


I afraid, it may leads to cell death. Because I tried with Tergitol & failed.

-Binu-

Try mixing it with BSA, which complexes fatty acids and makes them less toxic to cells.

-phage434-

QUOTE (phage434 @ Jan 5 2008, 12:38 PM)
Try mixing it with BSA, which complexes fatty acids and makes them less toxic to cells.


How to mix with BSA? please elaborate the protocol.

-Binu-

FA is less soluble in water as an free acid form. However, when converted to K+ salt , FA with 14 C chain length should be water soluble below its CMC (critical micelle concentration). C14 FA should have a CMC in 2-4 mM and its derivatives will have higher CMC. What will be the dosage you are going to use?

Alternatively, BSA is a good carrier. One BSA molecule binds up to 7 FA, so you can have a rough idea how much to use.

-genehunter-1-

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