A novel approach to quantitative LC-MS/MS: therapeutic drug monitoring of clozapine and norclozapine using isotopic internal calibration

Lewis Couchman*, Sarah Belsey, Simon Handley, Bob Flanagan

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

17 Citations (Scopus)

Abstract

Therapeutic drug monitoring (TDM) requires timely results in order to be clinically helpful. Such assays, when carried out using mass spectrometry-based methods, typically involve a batched sample approach with multipoint calibration. Isotopic internal calibration offers the possibility of open-access mass spectrometric analysis with consequent shortening of turnaround times. We measured plasma clozapine and N-desmethylclozapine (norclozapine) concentrations in (1) external quality assessment (EQA) samples (N = 22) and (2) patient samples (N = 100) using liquid chromatography-tandem mass spectrometry with isotopic internal calibration (ICAL-LC-MS/MS). Analyte concentrations were calculated from graphs of the response of three internal calibrators (clozapine-D-4, norclozapine-D-8, and clozapine-D-8) against concentration. Precision (% RSD) and accuracy (% nominal concentrations) for the ICAL-LC-MS/MS method were <5 % and 104-112 %, respectively for both analytes. There was excellent agreement with consensus mean and with 'spiked' values on analysis of the EQA samples (R (2) = 0.98 and 0.97, respectively, inclusive of clozapine and norclozapine results). In the patient samples, comparison against traditionally calibrated HPLC-UV and LC-MS/MS methods showed excellent agreement (R (2) = 0.97 or better) with small albeit significant mean differences (<0.041 and <0.042 mg/L for clozapine and norclozapine, respectively). These differences probably reflect discrepancies in the in-house preparation of calibrators and/or interference in the UV method. Internal calibration offers a novel and attractive alternative to traditionally calibrated batch analysis in analytical toxicology. The method described has been validated for use in the high-throughput TDM of clozapine and norclozapine, and allows for (1) same-day reporting of results and (2) significant cost savings.

Original languageEnglish
Pages (from-to)9455-9466
Number of pages12
JournalANALYTICAL AND BIOANALYTICAL CHEMISTRY
Volume405
Issue number29
DOIs
Publication statusPublished - Nov 2013

Keywords

  • Clozapine
  • Norclozapine
  • Isotopic internal calibration
  • LC-MS/MS
  • TDM
  • HEMOLYZED WHOLE-BLOOD
  • HUMAN PLASMA
  • ATYPICAL ANTIPSYCHOTICS
  • LIQUID-CHROMATOGRAPHY
  • ORAL FLUID
  • SERUM
  • HPLC
  • BIAS
  • MS

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