Theoretical Plate Method Validation

[daltore]

I am required to validate the test method of determining the height equivalent theoretical plates unless I am using a compendial method. Is anyone aware of a compendial source for determining HETP? Most sources, i.e. USP just give the equations and don't go into detail about how to perform the method. If I have to validate, what kind of repeatability should I expect from measuring HETP on the same column on multiple days? Thanks for your help!

[DR]

If you eamine the help or other literature that accompanies your data system software, you will probably find that it can do this calculation for you. HETP is (I think - haven't looked it up) a function of k and resolution, so it will vary with your retention times and resolution. I think theat by rearranging the HETP formula, you can get to a theoretical plates equation. Most suitability requirements just include a minimum number for this, which would correspond to a maximum HETP. Look at your HETP results and figure out what they would be if you lowered resolution to a marginally acceptable number and figure out what it would be if all of your retention times became a bit smaller - you will be pretty close to a reasonable spec if you plug these altered numbers into the HETP formula.

[danko]

Daltore wrote:

If I have to validate, what kind of repeatability should I expect from measuring HETP on the same column on multiple days?

First of all you should ask yourself: Why do I need to determine HETP? Here is the equation: HETP = L/N

L ~ column length
N ~ Plate count

Now, your column length obviously remains constant. So the only thing you should keep track on is Plate count!
FDA advises their inspectors to look after anything higher than 2000 (N > = 2000). This number is way too liberal in my mind, but you can use it as your lowest limit when you set a limit for N in your validation protocol.
Another criterion could be the separation of your critical peaks. Just find a chrom. which shows a reasonable resolution, then calculate N for your main peak and there you have it – recommended N for your application.

Finally, if somebody asks you again to calculate HETP, just ask him/her why. Or you might chose to be polite and calculate it according to the equation above.

Best Regards