Looking to validate RP-HPLC

Basic questions from students; resources for projects and reports.

3 posts Page 1 of 1
Hi,

I searched a lot in this subject but I really lost into details especially regulatory guidelines.

Frankly I developed a method and I need now to validate it.

Have you come a crosse any peer review paper that I can follow and get the paper published.

Sample: in water, soil extract etc
The book "Validating Chromatographic Methods: A Practical Guide", by David M. Bliesner is a close to a validation "cookbook" as you can find. Here's a link to it on Amazon:
http://tinyurl.com/cxmr6zy
-- Tom Jupille
LC Resources / Separation Science Associates
tjupille@lcresources.com
+ 1 (925) 297-5374
Hi, I would have to know what I was validating before I started a validation. As a chromatographer, you are establishing the proof that you have thought out your separation: sampling, sample prep, instrument functioning, and matrices. Don't stop here, these are just the topics off the top of my head. If you are doing a validation for the FDA, you will document literature research, interviews, and any other source of information that may cause your method to fail. That is, you need to show that your protocol tested the likely challenges to a successful separation and analysis. Your protocol, based on your knowledge of what you are validating, will have sections covering: abstract, introduction, experimental procedures, data calculations, analyses, discussion, conclusions, schedule, personnel, and sign-off. At least these topics and you may need more sections. The protocol very specifically directs the work. A validation is not a research project, for which results may dictate experiments that were not planned. A well-written protocol writes the report. For example, the protocol says what will be discussed and how- the report discusses what the protocol ordered.
If your substance is a small molecule, the literature will tell you how it could be synthesized, The precursor molecules should be used to challenge the separation. Stability limits some separation accuracy, not just in getting the sample to the chromatograph, but degradation products interfere with the separation. Use heat, acid, base, etc. to degrade you samples and see if the separation still works well.
As part of a validation, you will show the method is working each time it is used. A 'proof' sample, a sample you specify will have to pass specifications you establish for a successful separation. Retention time, resolution, tailing etc. make good parameters to evaluate the quality of each separation. During the validation you will repeat assays, usually five or more times to show you get the same result each time. (Declare the first or first few assays are not to be used, if you think you need to avoid failure before the equipment is up to speed.)
I think I could ramble on for days. A concrete example of your assay would make it easier to address a validation for it. Your regulatory requirements dictate the form of the validation. A validation is just a new buzzword for developing and showing a method works.
Jim
3 posts Page 1 of 1

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