This kind of work is usually trial and error type. So you can start with the easiest procedure and gradually improve

Hi Chai.

Your method development will be depend of matrix complexity, quantification limit and tocopherol concentration in your samples. Link below show a compedium methodology for quantification alpha tocoferol in tablets.It is a good starting point for your methodology, but could be necessary make necessary changes for your matrix.

Regards

http://jpdb.nihs.go.jp/jp14e/14data/Part-I/Tocopherol.pdf

Your method depends on the oil (specificity) and expected concentration (LOQ + linearity). Check the AOCS (American Oil Chemists' Society) for an method you can adapt or the internet. Read over ICH Q2(r1) for validation parameters.

Not much point re-inventing the wheel!

Thanks for the feedback everyone !

Looks like detection of tocopherols are still widely debatable for different methods

As I recall, it was a C18 column with 95% MeOH/5% Water flowing at 0.5 mL/min. (for a 250 mm column) and detection at 285 nm.

1.0 mL/min. for a 50 mm column. Depends on how you will extract the oil! I would just dilute in IPA, ultrafilter, and shoot!

That's roughly what I've been reading as well Bruce.

Good idea too to start based on your recommendation.

Time to give it a go and shoot ! Thanks :)

Dear Chai Yee Ho, as you have a C18 column, I believe you want to work with HPLC. But I would like to suggest some reflections before performing the validation of the method. First, do you need to know the purpose of the validated method (purity, quantification, indentification)? Secondly consider which official guides reference methodologies for alpha-toeopherol analysis? See if these methodologies apply or need to be adapted. Normally the complexity of the matrix is ​​the main aspect that prevents the direct application of a method (there are several articles dealing with matrix effect). Look at the structure of your molecule and see which chromatographic systems can be used to analyze (HPLC, GC). If you need high sensitivity either because of the purpose of the method (limiting assays) or due to matrix constraints (low concentrations), you can use GCMS as a technique to validate the method. It can even work with alpha-toeopherol chemical derivatization. Another precious suggestion is that you work hard on developing a sample pretreatment methodology that ensures clean analysis of (which influence the purity of the chromatographic peaks) and free of greases (which decrease the useful life of the columns) and finally provide preconcentration of analytes in the sample analyzed.

Good job.

Thanks for the reply Valmir.

Looks like there's quite a number of things to consider as well based on your reply.

Much appreciated !