Based on the chromatogram of the blank hexane, it shows that there are ghost or unknown peaks in that chromatogram even though most of the parts of the GC have been changed to new prior analyzing the blank hexane.
Plus, the running of another blank hexane showed only one peak which can be confirmed as the hexane peak.
Is it possible the first blank hexane is contaminated?
Another question is:
Is it possible if the retention time of different concentrations FAME standard are different in big margins as compared with the retention time in the Certificate of Analysis ( COA- less than 20 minutes, all peaks were observed, prepared FAME Standard- more than 70 minutes).
If it is possible, how can we identify which compound is which because of the FAME standard retention time is very different compared with the COA?
Mr. Ali,
Please, consider detail on the following discussion:
[https://www.researchgate.net/post/How_to_seperate_two_components_by_RP-HPLC_having_very_near_pKa_values_ie_1298_and_1338].
Retention times do not change with changing concentration, they are always stable.
If your blank was the first injection of the day then you are probably seeing septum bleed that has accumulated on the head of the column or in the injection liner. Very common - one of the reasons you always run a blank injection before you inject actual samples.
If you are trying to match a COA then you have to use EXACTLY the same conditions and column as are shown on the COA. Even then you may not be successful - I had a standard from a major reference standards manufacturer that showed pentane eluting after octane on the COA. Uhhhhh.......
Given the fact that your FAME analysis took 70 minutes and the COA shows 20 tells me you need to match conditions first. Try that and see if your retention issues don't go away.
Concerning your first question, if changing the solvent the ghost peaks disappeared, then the first solvent is contaminated, since the change is discarding contamination of the GC column or system contamination.
If the retention times of the FAME are different compared to the certification test, you should study it.
It could be related to changes in the method or factors such as column, flow, thermal profile. I suggest you to develop your own calibration curve for the full FAME kit. Then you will accurately know the qualitative identification of compounds.
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