We are working on Microalgae biodiesel. For FAME analysis GC MS was carried out without standard run and Peak area and RT is given in data. So from peak area and RT how we can measure the concentration of particular compound.
In case you did not add an IS prior to cell disruption/extraction, then it is virtually impossible to determine the fatty acid concentrations in your original sample. Only in case you have a very reproducible extraction method, you could try to estimate the fatty acid concentrations in your samples based on comparison with additional analyses of similar samples (that include an IS). Find attached a copy of our manuscript describing our fatty acid analysis protocol.
Also, the response factors of your IS and the fatty acids in your sample may differ a bit, so it is also advisable to run standards for all fatty acids (N.B. theoretical calculation of response factors relative to that of your IS are possible...I do not have a reference at hand).
Generally, you should use internal standard which is a compound similar to that expected in your sample but never really occur therein, added to the sample in a an amount known in advance at the stage of extraction (e.g. margaric acid, 17:0). Then you can compare area of IS with any unknown component and deduce the amount of the latter. Actually, all GC/MS software allow you to select the IS peak and carry out all calculations automatically.
Alternatively, IF you do not use IS (which is not so precise as the quantification with IS), you will need a calibration curve (this is called external standard method). constructed from the peak area from chromatograms with several known concentrations of the external standard.
thanks for your answer that my question i do not have the external standard thats why the problem occurred.
In case you did not add an IS prior to cell disruption/extraction, then it is virtually impossible to determine the fatty acid concentrations in your original sample. Only in case you have a very reproducible extraction method, you could try to estimate the fatty acid concentrations in your samples based on comparison with additional analyses of similar samples (that include an IS). Find attached a copy of our manuscript describing our fatty acid analysis protocol.
Also, the response factors of your IS and the fatty acids in your sample may differ a bit, so it is also advisable to run standards for all fatty acids (N.B. theoretical calculation of response factors relative to that of your IS are possible...I do not have a reference at hand).
You can use calibration curves of compounds with the same ritention time of the compounds that are in your samples. You can buy the FAME mix (Sigma Aldrich) to compare the retention times and can use the methyl decanoate like internal standard or other compounds with a low carbon atoms (C10-C14).
thanks Zuccaro and lamers for your paper i will try and lets see what i will get.
It may have been too obvious that no one mentioned what is measured as the response! .Since GC and GC MS peaks have a very small base, peak height will give the response. No need to calculate peak area
To answer Priti's question : Peak height is the scale reading of the GC peak on the y-axis of your GC output. The peak width ( x -axis) should be very small in a good GC analysis. Hope this helps!
Peak area is height (y-axis) x width (x-axis). so, to calculate the fatty acid concentration we are using peak area. as suggested by lamers through adding one internal standard (usually the one that not appear in your sample) to the sample extraction you can calculate the fatty acid concentration.
I am a little bit confused regarding calculation. I calculate the concentration of FAME from grapes by a linear equation.
I am not sure about the final unit after the calculation. so what is the unit?
I used 2 g sample (1:1 ratio of grape sample and silica sand), after that, I adopt the extraction procedure, methylation and drying under the nitrogen blow. Then it dissolves in 990 µL hexane +10 µL internal standard and run on GC/MS.
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