For inhibitor EPZ-5676 assay/related substances methods, the active peak should be adequately resolved from all impurity/degradant peaks, placebo peaks, and sample blank peaks. Resolution from impurity peaks could be assessed by analyzing a spiked solution with all known available impurities present or by injecting individual impurities and comparing retention to that of the active. Placebo and sample matrix components should be analyzed without the active present in order to identify possible interferences. If syringe filters are to be used to clarify sample solutions, an aliquot of filtered sample diluent should be analyzed for potential interferences. If the impurities/degradants are unknown or unavailable, forced degradation studies should be performed.
Forced degradation studies of the active pharmaceutical ingredient (API) and finished product, using either peak purity analysis or a mass spectral evaluation, should be performed to assess resolution from potential degradant products. The forced degradation studies should consist of exposing the API and finished product to acid, base, peroxide, heat, and light conditions, until adequate degradation of the active has been achieved. An acceptable range of degradation may be 10�C30% but may vary based on the active being degraded. Overdegradation of the active should be avoided to prevent the formation of secondary degradants. If placebo material is available, it should be stressed under the same conditions and for the same duration as the API or finished product.
The degraded placebo samples should be evaluated to ensure that any generated degradants are resolved from the analyte peak(s) of interest. Evaluation of the forced degraded solutions by peak purity analysis using a photodiode array detector or mass spectral evaluation must confirm that the active peak does not co-elute with any degradation products generated as a result of the forced degradation. Another, more conservative, approach for assay/related substances methods is to perform peak purity analysis or mass spectral evaluation on all generated degradation peaks and verify that co-elution does not occur for those degradant peaks as well as the active peak. Whereas the selectivity experiments for the first approach can be performed during a prevalidation phase (no need for quantification), those for the second approach are usually performed together with the precision and accuracy experiments during the main validation phase.
At this point it must be mentioned Brefeldin_A that the term specificity is used interchangeably with selectivity, although in a strict sense specificity refers to methods, which produce a response for a single analyte, whereas selectivity refers to methods that produce responses for a number of chemical entities, which may or may not be distinguished. Selective multianalyte methods (e.g.