I want to analyze Fatty Acid Methyl Esters on GC-MS. Currently i am planning to use 30 m x 0.25 mm-internal diameter 70% cyanopropyl polysilphenylene-siloxane 0.25 μm TR-Fatty Acid Methyl Esters (FAME) stationary phase column for GC. but kindly suggest me any other capillary column which I can also use for FAME?
Hi. If I am not mistaken you are researching on biofuels. The group in my ex department were using a capillary column (DB-5HT) dimensions (15 m 0.32 m 0.1 μm), and a 530 μm inner diameter. The detector is FID for sure. You can refer to their publication through the following link:
http://pubs.acs.org/doi/abs/10.1021/ef2004943?journalCode=enfuem
Misbah,
You may carry out your analysis in DB-23 FSCAP or other similar phases of a min of 30 m length column.
Good Luck
Dr Chanotiya CSIR-CIMAP, INDIA
If you need better resolution for CLA or omega3 fatty acid methyl esters I suggest you to choose longer columns (50-60 m). In my experience the best choice I think is Restek columns specific for FAME analyses.
I agree with Daniele, longer columns give you a better resolution especially when you deal with trans-isomers. Moreover, depending on the matrices you can face with huge number of FAME from the shorter 8:0 or 10:0 up to the long chain 26:0.
Things get complicated when the FAME chain is highly unsaturated (one or more double bonds) or branched, in these cases running times get longer but necessary to resolve as many peaks as possible. Longer column ( 60 and even 100mt) gives you better resolution and separation.
thank u so much everyone for your valuable answers.
I want to see level of free circulating fatty acids in plasma. Daniele and Fulvio your suggest is very good. I will surly consider it and get the column of 60 m or more.
Hello Misbah Hussain,
The choice of column for FAME analysis will depend upon the information you would like to get from the analysis.
Non-polar columns are routinely used to provide results on the amount of saturated versus unsaturated fatty acids.
Example chromatogram on Equity-1:
http://www.sigmaaldrich.com/catalog/analytical/supelco/g004278
Polyethylene glycol based columns are used to provide carbon chain length & degree of saturation resolution.
Example chromatogram on Omegawax:
http://www.sigmaaldrich.com/catalog/analytical/supelco/g004263
The analysis of cis and trans FAMEs, along with resolving their positional isomers, requires the use of highly polar biscyanopropyl polysiloxane columns such as SP-2560 or SLB-IL111 (AOCS Method Ce 1h-05, “Determination of cis-, trans-, Saturated, Monounsaturated and Polyunsaturated Fatty Acids in Vegetable or Non-ruminant Animal Oils and Fats by Capillary GLC” AOCS Official Methods (2005), American Oil Chemists Society.
Example chromatograms:
SLB-IL111: http://www.sigmaaldrich.com/catalog/analytical/supelco/g005367
SP-2560: http://www.sigmaaldrich.com/catalog/analytical/supelco/g005366
Hi Misbah,
I use for FAME analysis ZB-5MS from Phenomenex. You can see more information in my articles from RG. I hope it will be useful for you!
Good luck in your research!
Alex :-)
Hello Misbah Hussain.
The type of GC liquid stationary state depends upon the aim of your work: 3 days ago you have defined your aim as determination of FREE fatty acids in human plasma. But you can achieve this goal analysing TRIMETHYTSILYL esters of FA on non-polar methyl siloxane columns (instead of analysing METHYL esters on polar columns). Such extremely simple, sensitive and selective method is presented for example by Kurkis et al. [Kurkis A., Myher I.I. Marai L., Geher K. Estimation of plasma free fatty acids as the trimethylsilyl esters. Anal. Biochem. 1976, 70(2), 302, or Kurkis A., Myher I.I., Geher K. Quantification of plasma lipids.... J. Lipid res. 1993, 34(6), 1029]. Briefly: internal standard (free 17:0-acid) is added to your sample, then it is extracted using Folch mehod, a portion of chloroform extract is dried and residue is derivatized witn pure bis(trimethylsilyl)trifluoroacetamide (0,05-0,1 mL) or its 1:1-mixture with dry pyridine. There is no need for preliminary separation of free FA from other lipids (TG, PL, CE) because TMS-esters are formed only by free FA, no FAME-artefacts can emerge as a result of trans-esterification of TG, DG, PL with nucleophylic methylating reagents during conversion of free FA into treir ME (this complication is extremely dangerous!). Relatively short capillary columns (10-20 m) may be successfully used (according to my own experience). One can mention than chromatographic effectiveness of nonpolar columns is much higher than that of polar columns - as well as maximum working temperatures! The pairs of saturated and monounsaturated acids (12:0-12:1, 14:0-14:1, 16:0-16:1, 18:0-18:1, 20:0-20:1) are separated very efficiently even on 12-meter HP-1 (0,25 mm x 0,25 mkm) column.
It must be noted certainly that isomeric analysis of monounsatirated FA cannot be done with the help of nonpolar columns: for example, it is impossible to separate oleic and vaccenic acids (typical for human plasma) with such unselective phase. It is necessary to use polar columns of considerable length (>25 m, Supelcowax or AT-1000) to separate such positional isomers.
As to me SUPELCOWAX-10 columns are the best polar CC for FAME analysis (in particular it effectively withstands to corruption with moist samples for several years of intensive work).
Dear researcher, you can use DB-23, SP-2330 and BPX- 70 capillary columns for FAMEs analysis.
Dear Misbah, I am using Thermo Scientific™ TRACE™ TR-FAME GC Column, 60m x 0.25mm x 0.25um, max temperature 250/260 C. This column is a cyanopropylphenyl-based phase specifically designed for the separation of FAMEs. also suitable for any application that requires a high-polarity column for use with MS detection.
Do suggest the GC FID setting for DB-5ms column for FAME analysis
Dear Misbah, i am also require same column for separation of vocs.
so, please, tell me which company or supplier provides this column in india
Hi, from my experience I can say that HP-5MS is a good column for resolving FAME chromatograms. However this column can not separate C18:3 isomers. I tried several years analyzing FAME from microalgae and resolution was very good, for C18:1, C18:2, EPA and DHA, but when I tried with vegetable oils was not possible and DB-23 (60 m) was a very good choice. Best regards from Chile.
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