Prevalidation statistical design to assess analytical methods. Example of a quick liquid chromatographic assay of itraconazole in serum.

Abstract

(A) An analysis-of-variance spreadsheet program is presented which allows to readily test and/or quantitate in a single run analytical linearity, matrix effect on recovery, repeatability of measurement and of extraction and the ruggedness of these features for up to three sessions. Owing to napierian logarithmic transformation, ANOVA mean squares directly read as relative standard deviations and checking linearity is straightforward. (B) A quick assay for therapeutic drug monitoring of itraconazole and its main metabolite was devised with the help of the program, and subsequently validated according to current quality control recommendations. The assay involves acetonitrile demixing extraction, reversed-phase HPLC and UV detection and shows acceptable performance from 0.06 to 5.0 mg/l (limit of detection about 0.03 mg/l). The prevalidation design fairly predicted precision and accuracy, was more informative about matrix effect and was even more demanding about analytical linearity.