Intra-assay precision testing

Deflazacourt is an inactive prodrag which after oral administration is rapidly converted to the active 21-hydroxydeflazacort. It has anti-inflammatoiy activity, and is used in the treatment of rheumatoid arthritis. Ifa et al. [87] developed a method for the determination of 21-hydroxydeflazacort in human plasma. The analyte and dexamethasone-21-acetate as the IS are extracted from plasma using a diethyl ether LLE. After evaporation to dryness and reconstitution in mobile phase, the compoimds are analysed using LC-MS with positive-ion APCl in SRM mode, monitoring the transitions m/z 400 124 for the analyte and m/z 435 397 for the IS. The linearity was tested in the range of 1 00 ng/ml. The intra-assay precision and accuracy were better than 5.5% and 7.1%, respectively. 

The repeatability or intra-assay precision is defined as the precision of determinations performed in the same way and within a short space of time. Its determination is of particular interest in the analytical testing for the imiformity of the content of a measmed active ingredient (content uniformity test). 

Intra-assay precision Run samples (S5 replicates) in a single run. Samples used for precision testing may include commercial QC samples, but precision testing should also include species-specific samples of the relevant matrix (e.g., serum, plasma, urine) for the intended study samples Test S3 samples at low, medium, and high concentrations across the calibration curve range Calculate mean, SD, and CV% CV 20%, CV<25% at LLOQ... 

QC (see Section 41.5.9) samples that will be used to monitor the assay during routine use should also be subject to the intra- and inter-assay precision testing outlined earlier during the validation process. 

Precision defines the scatter of repeated analysis, or the coefficient of variation of analytical results. Both intra-assay and inter-assay precision must be investigated. Intermediate precision describes the influence of different analysts, equipment, days and other intra-laboratory variabihty. Inter-laboratory comparison is also of interest in establishing the precision of the method. AU testing on accuracy and precision must be carried out by replicate analyses of a statistically relevant number of samples. Depending on the use of the method, it may be necessary to estabhsh both parameters over the measurable range, or in the case of content determination simply in the range of 80-120% of the nominal value. 

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