method validation for low (assay) and high load (impurities)

[ntruong]

Hi,
To validate an HPLC method for assaying of an active compound, do I need to validate both for high concentration (to detect impurities) and lower concentration (for assaying of the active compound)? ICH guidelines for method validation parameters such as linearity, accuracy, range, etc... will be followed for the assay part, but I have no idea about the higher load, which is needed to chromatographic impurities. The concentration for high load is about 1 mg/mL and for assay is 0.1mg/mL. Any suggestions?
Thanks in advance,

[Sunjay]

Hi Dear,

In my opinion, the different method validation parameters should cover the whole concentration. Like for linerity it should cover 80% of assay conc to 120% of impurities concentration and like for hte other parameters.

Simillarily precision, ruggedness should be determined both for assay and impurities.

For specificity, higher concentration will be ok, since yuo have to demonstrate non-interferance at RT of impurities also.

Hope this will help.

Pls share any different views.

Regards

[KC]

I will repost a comment I made recently on a similar thread

Just finished an impurity study where I was looking at 4 impurities ranging in conc from 0.5 ug/mL to 12.0 ug/mL in solutions of the active compound at a conc of 1200 ug/mL. In order to quantify all impurities I prepared combined standards of the 4 impurities from 0.5 to 20 ug/mL and analyzed all solutions disregarding the huge active compound peak (way off scale). To quantify the active compound, I did 25X dilutions on all solutions and analyzed the samples again with active compound standards ranging from 10.0 to 100 ug/mL. Worked like a champ, r2 for each of the 5 components was 1.000 and never had a problem with overloading of the detector.

I will also add that each one of the components (active compound and impurities) had to be validated for specificity, accuracy and precision at appropriate concentrations in solutions of the active compound. Impurity validation levels were 0.500 and 20.0 ug/mL to cover the expected concentrations in the solutions and the active compound validated at the higher 1200 ug/mL level. Each validation was performed SEPARATELY by standard addition before all components were quantitated as described above. I will also add that to avoid the 25x dilutions I spoke of, you may be able to just reduce the injection volume when looking at the active compound to make the whole procedure as easy as just analyzing the same samples twice. You have two methods that are basically the same with the only difference being the injection volume and scaling factors.

[ntruong]

Hi Sunjay and KC,
Thanks for sharing your thoughts on the topic. The method that I am going to validate turned out to be for chromatographic purity determination only. Therefore, there are not much requirements as I thought it was. By the way, I checked one of the method that was done by the contract lab for us (similar situation for high and low load) and it wasdone exactly like KC did.
Thanks again,