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NK ☆ India, 2025-07-28 11:49 (337 d 21:49 ago) Posting: # 24417 Views: 3,233 |
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Dear All, How should the specificity experiment be designed and performed during bioanalytical method validation for Fixed Dose Combination (FDC) products using LC-MS/MS? As per the ICH M10 section 3.2.2, Specificity, “In the case of LC-MS based methods, to assess the impact of such substances, the evaluation may include comparing the molecular weight of a potential interfering related substance with the analyte and chromatographic separation of the related substance from the analyte.” In FDC studies involving multiple analytes, What is the recommended approach to ensure that the presence of one analyte at high concentration (e.g., Cmax or ULOQ) does not interfere with the accurate detection and quantification of another analyte at low concentration (e.g., LLOQ)? Should specificity testing requires only spiking blank biological matrix samples with additional analytes at high concentrations to assess potential cross-talk or interference or is it appropriate to evaluate specificity using LLOQ-level samples for one analyte in the presence of co-administered analytes at their respective Cmax concentrations? Regards NK Edit: Guideline linked. [Helmut] |
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Ohlbe ★★★ France, 2025-07-29 10:55 (336 d 22:43 ago) @ NK Posting: # 24419 Views: 2,640 |
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Dear NK, ❝ Should specificity testing requires only spiking blank biological matrix samples with additional analytes at high concentrations to assess potential cross-talk or interference or is it appropriate to evaluate specificity using LLOQ-level samples for one analyte in the presence of co-administered analytes at their respective Cmax concentrations? On a regulatory point of view, ICH M10 states: Responses detected and attributable to interfering components should not be more than 20% of the analyte response at the LLOQ and not more than 5% of the IS response in the LLOQ sample. So it looks like regulators still only consider the risk of interference in the same old-fashioned way as in the days of HPLC-UV: visible peaks. In this case, it is sufficient to spike blank plasma with analyte A and to make sure that no peak appears at the retention time and m/z transitions of analyte B and its IS. Personally with LC-MS/MS I am more concerned about the non-visible interferences (e.g. ion suppression or enhancement). So in addition to spiking blank plasma, I would spike blanks with A at ULOQ / B at LLOQ and A at LLOQ / B at ULOQ. If you want to be extra-cautious and to consider competition for ionisation, which may only be visible at high concentrations of both, you can also spike A at ULOQ / B at ULOQ. — Regards Ohlbe |
Ing. Helmut Schütz