In-house validation of a liquid chromatography–tandem mass spectrometry method for the determination of selective androgen receptor modulators (SARMS) in bovine urine

ABSTRACT

A sensitive and robust LC–MS/MS method allowing the rapid screening and confirmation of selective androgen receptor modulators in bovine urine was developed and successfully validated according to Commission Decision 2002/657/EC, chapter 3.1.3 ‘alternative validation’, by applying a matrix-comprehensive in-house validation concept. The confirmation of the analytes in the validation samples was achieved both on the basis of the MRM ion ratios as laid down in Commission Decision 2002/657/EC and by comparison of their enhanced product ion (EPI) spectra with a reference mass spectral library by making use of the QTRAP technology. Here, in addition to the MRM survey scan, EPI spectra were generated in a data-dependent way according to an information-dependent acquisition criterion. Moreover, stability studies of the analytes in solution and in matrix according to an isochronous approach proved the stability of the analytes in solution and in matrix for at least the duration of the validation study. To identify factors that have a significant influence on the test method in routine analysis, a factorial effect analysis was performed. To this end, factors considered to be relevant for the method in routine analysis (e.g. operator, storage duration of the extracts before measurement, different cartridge lots and different hydrolysis conditions) were systematically varied on two levels. The examination of the extent to which these factors influence the measurement results of the individual analytes showed that none of the validation factors exerts a significant influence on the measurement results.

KEYWORDS:

• Selective androgen receptor modulators
• LC–QTRAP–MS/MS
• enhanced product ion scan
• matrix-comprehensive in-house validation
• factorial analysis
• residue control

Acknowledgments

The author wishes to thank Ms. M. Grunow (BVL) and Ms. D. Holzkamp (BVL) for their technical assistance in the in-house validation study. Dr. Jens Dahlmann (Sciex) is gratefully acknowledged for helpful discussions with regard to recording of the EPI spectra and the generation of reference mass spectral libraries.

Disclosure statement

No potential conflict of interest was reported by the authors.