Here you can find high cited paper: http://www.jlr.org/content/24/1/83.short

As suggested above, any C18 RP with an acetonitrile:water gradient should work. However, I would encourage you to switch to GC (w. FID) instead of LC if you are interested in good quantification, in particular at low sample amounts or concentrations. Only if you need a preparative approach, I would go for LC. But of course I do not know whether you have the options in terms of analytical equipment.

I have developed one method for fatty acids by HPLC, you can try it if it works for you.

Mobile phase: Acetonitrile: Water (80:20), Column: Luna Kinetex C18, 4.6x100 mm, 2.6µ, 100A°.

It works for me and separating more than 10 fatty acid compounds between C18 to C22 carbon chain.

If the number of fatty acids is few then HPLC is a viable approach but I strongly recommend considering GC-FID for quantification. There are several Compendia methods that would be suitable.

I strongly agree with Michael. Partly due to the much more accurate quantification with the FID, but also if you are interested in separating and analyzing position double bond isomers (for example C18:1n-9 and C18:1n-7). In that case GC-FID or GC-MS on a relatively polar column is the only suitable method.

Dear Okky can you send me your

HPLC method about analysis FFA and also I have a big problem to separate phthalats in plastic matter especially in children toys with HPLC metods

Can anyone help about this.

Thanks

Nili

Hi Dilip I want to try HPLC method suggested by you,also can you please tell me the procedure for extraction of Fatty acid

I am trying to do short chain fatty acid (AcOH, PrOH and ButOH) analysis from human feces. Using Perkin Elmer with PDA detector HPLC and column Perkin Elmer 150x4.6 mm with 5uM. It has the limitation of pH from 2.5 to 7.5. NaH2PO4 as a mobile phase with pH 3~. When I inject the standards AcOH, PrOH and ButOH, their retention time are 0.9, 1.1 and 1.6 min. respectively, with 1ml/minute flow rate. They are very close, it will be very difficult to separate with samples. How I can increase the retention time, except increasing the flow rate because recommended flow rate from Column Company is 1ml/min.

Kindly suggest me what could be the problem.

Thanks in advance.

Khemlal

Dear Mr. Nirmalkar

Your short chain fatty acids are less polar, all are not retained by your chromatography. To increase retention reduce polarity of your mobile phase composition. You didn't mention full composition of your mobile phase. fatty acids are ester alcohols and not much affected by pH except higher pH (8-9) so here in your chromatography  i don't feel pH of the buffer is much playing role. You didn't mentioned stationary phase of your HPLC column but i believe it should be C18.

To me, make more simple chromatography. I would suggest to try this.

Mibile Phase: Acetonitrile: Water (80:20) Ratio can be adjusted to adjust peak elution

Stationary Phase: C18, 250X4.6mm, 2.6 to 5µm (Smaller partcle size proffered)

Good luck.

I have done GC-MS of FAME but the abundance i am getting of oleic acid methyl ester peak is way low. I have a feeling that my fatty acid i.e. oleic acid is getting degraded during transesterification reaction to make oleic acid methyl ester. Please suggest me a proper procedure for this as I am struggling from three months. I even tried oleic acid detection through HPLC of Free fatty acid but i am not successful

What is your transesterification reagent and procedure to form FAMEs? Without knowing that, it is hard to judge whether your method may affect oleic acid degradation. Apart from that, I suggest to avoid sample exposure to light and oxygen to avoid an oxidation of the double bonds in your unsaturated fatty acids.

Aim- I need to calculate the uptake rate of oleate by e.coli.I use 0.1% of sodium oleate which is 82% pure from sigma.

Firstly I mix sodium oleate with briz-58(surfactant) in minimal media and make 500ul volume of this solution..

1. To this 500ul of solution 0.66 ml of methanol: toluene: dimethoxypropane: H2SO4 (39:20:5:2) and 0.34 ml of hexane was added.

2. The mixture was incubated at 80`C for 1 h.

3. After heating, the tubes were cooled at room temperature. 500 μl of hexane was added.

4. Two phases formed, the upper one containing the FAMEs. This phase was collected in new tubes and it was evaporated to 50 μl using a vacuum concentrator at room temperature.

I use 3N methanolic HCl (Supelco 33050-U) for 20 min at 70°C for transesterification, which works well. On the other hand, I do not see why your protocol should not result in a quantitative transesterification of oleate. Extraction of FAME with hexane should work well.

Dear all, I am also interested in for the determination of short chain fatty acids (e.g. acetic, butyric, propionic etc.) by chromatographic systems. And I’ve looked through your valuable comments on this issue. As I see, most of you, has suggested to go with GC-FID system instead of HPLC. So, my first option has become the GC method for this analysis. So I am kindly asking you to suggest me a good GC-FID protocol for the VFA analysis, including; pretreatment of the fermented VFA containing liquid samples, thermal program of GC, etc…