Chromatography Forum

Hi, now i am doing fatty acid GC method development, later i need validate this method

our product include vegetable oil and fish oil, my methodology is following:
1. i use the simplest method for transesterification of triglycerides, KOH/METHANOL
2. I use the internal standard and standards to correct and quantitate the transesterication, the reaction is impossible complete 100%
3. i add the internal STD into the sample from the extraction beginging
4. I use the ethyl STD, proceed the same extraction procedure as of sample
from these procedure, I think i can recover the any loss of extraction, because the STD also experience the same procedure, I assume it has similar loss as sample.
I am new to FAMEs, thank you for your support.
by the way, during validation stage, what kind of specificity test is specific to GC and the FAMEs analysis, and how can I detect the peak purity? I only have FID detector, I use Agilent DB-23 column, which is high polar, with the 50% cyano-propyl as the stationary phase.

I don't fully understand what your internal standard is, and I've been doing FAMEs assays for 30 years. If I was going to use an internal standard for this, I'd use an odd-chain, not an "ethyl STD" (the term I don't understand. I'm also wondering why you need to validate such a common and universally-used procedure.

Aren't there ASTM methods which require no validation?

best wishes,

Rod

What does it mean ASTM?
thanks
best wishes,

Sorry Elaine,

I did intend to type AOCS but typed ASTM, similar but not the same.

American Society for Testing and Materials, a group which proposes standard methods of analysis.

best wishes,

Rod

Thanks!
from the methods decribed from those authorites, they derivertise the triglycerides into methyl ester, but they use ethyl standard, why?
best wishes,

Rod[/quote]

An internal standard works best if it separates from the components you wish to measure and has a similar response to the detection method that is used.

PS Do not expose the ethyl ester to the methyl ester reagent or you will convert your ethyl ester to methyl ester.

best wishes,

Rod

If the AOCS method is anything like the EP method then something like C23:0 would be a good standard to use.

As far as i am aware there should be very few peaks (if any) around C23:0. We also test veg and marine oil and C23 is used in quantitative analysis of oils.

hope this helps

steve

The AOCS has produced GC methods that work well, are accepted worldwide, have standard mixtures and internal standards ( eg C21:0 for Ce-1b ) that are readily available from most suppliers, and cover the FAME profiles of most oils. The AOAC will also methods relating to foods, and pharmacopeia have methods for drugs and supplements.

Why would you not just follow one of those methods?. A few examples are below, but they have many others, as a search of their site will reveal. Using ethyl esters as Internal Standards is no longer preferred by most authorities for analysis of commonly-available oils.

AOCS Official Method Ce 1b-89 ( revised 2005 )
Fatty Acid Composition by GLC Marine Oils

AOCS Official Method Ce 1h-05 ( revised 2005 )
Determination of cis-, trans-, Saturated, Monounsaturated and Polyunsaturated Fatty Acids in Vegetable or Non-ruminant Animal Oils and Fats by Capillary GLC

AOCS Official Method Ce 1-62
Fatty Acid Composition by Gas Chromatography
- The standard packed column method.

Bruce Hamilton

Hi, thank you very much.
where can i find those official method?