Analytical and Bioanalytical Chemistry

, Volume 406, Issue 20, pp 4861–4874 | Cite as

Optimization and validation of a label-free MRM method for the quantification of cytochrome P450 isoforms in biological samples

  • Ahmad Al Ali
  • David TouboulEmail author
  • Jean-Pierre Le Caër
  • Isabelle Schmitz-Afonso
  • Jean-Pierre Flinois
  • Catherine Marchetti
  • Isabelle De Waziers
  • Alain Brunelle
  • Olivier Laprévote
  • Philippe Beaune
Research Paper


Cytochromes P450 (CYPs) play critical roles in oxidative metabolism of many endogenous and exogenous compounds. Protein expression levels of CYPs in liver provide relevant information for a better understanding of the importance of CYPs in pharmacology and toxicology. This work aimed at establishing a simple method to quantify six CYPs (CYP3A4, CYP3A5, CYP1A2, CYP2D6, CYP2C9, and CYP2J2) in various biological samples without isotopic labeling. The biological matrix was spiked with the standard peptides prior to the digestion step to realize a label-free quantification by mass spectrometry. The method was validated and applied to quantify these six isoforms in both human liver microsomes and mitochondria, but also in recombinant expression systems such as baculosomes and the HepG2 cell line. The results showed intra-assay and interassay accuracy and precision within 16 % and 5 %, respectively, at the low quality control level, and demonstrated the advantages of the method in terms of reproducibility and cost.


Calibration curve in complex matrix for CYPs quantification


Cytochrome P450 Mass spectrometry Multiple reaction monitoring Label-free quantification Liver microsomes Baculosomes 



Financial support of the Institut de Chimie des Substances Naturelles (CNRS-ICSN) is gratefully acknowledged.

Supplementary material

216_2014_7928_MOESM1_ESM.pdf (145 kb)
ESM 1 (PDF 144 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Ahmad Al Ali
    • 1
    • 2
  • David Touboul
    • 1
    Email author
  • Jean-Pierre Le Caër
    • 1
  • Isabelle Schmitz-Afonso
    • 1
  • Jean-Pierre Flinois
    • 2
  • Catherine Marchetti
    • 2
  • Isabelle De Waziers
    • 2
  • Alain Brunelle
    • 1
  • Olivier Laprévote
    • 3
    • 4
  • Philippe Beaune
    • 2
  1. 1.Centre de Recherche de GifInstitut de Chimie des Substances Naturelles, CNRSGif-sur-Yvette CedexFrance
  2. 2.Médecine Personnalisée, Pharmacogénomique et Optimisation Thérapeutique, INSERM UMR-S1147, Centre Universitaire des Saints-PèresUniversité Paris DescartesParis Cedex 06France
  3. 3.Chimie-Toxicologie Analytique et Cellulaire, UMR CNRS 8638, Faculté des Sciences Pharmaceutiques et BiologiquesUniversité Paris DescartesParisFrance
  4. 4.Service de Toxicologie BiologiqueHôpital LariboisièreParisFrance

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