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TOPIRAMATE DOSAGE IN DRIED BLOOD SPOTS BY A FAST LIQUID CHROMATOGRAPHY-TANDEM MASS SPECTROMETRIC METHOD FOR NEWBORNS
  1. G laMarca1,
  2. S Malvagia1,
  3. L Filippi2,
  4. P Fiorini2,
  5. G Fontanelli2,
  6. G Pieraccini3,
  7. G Moneti3,
  8. G Cavallaro4,
  9. R Guerrini1
  1. 1Clinic of Pediatric Neurology, A. Meyer University Children’s Hospital, Florence, Italy
  2. 2Neonatal Intensive Care Unit, A. Meyer University Children’s Hospital, Florence, Italy
  3. 3CISM, Mass Spectrometry Center, University of Florence, Florence, Italy
  4. 4Neonatal Intensive Care Unit, Carlo Poma Hospital, Mantova, Italy

Abstract

Objective Topiramate is used to treat a wide range of epileptic disorders in children, including newborns, thanks to its inhibitory activity against glutamate receptors. Its clinical effect correlates better with blood level than with doses, which provide the rationale for topiramate monitoring.

Methods Up to now topiramate has been dosed by HPLC, mass spectrometry (MS), GC/MS, capillary electrophoresis/UV, or fluorescence polarization immunoassay (FPIA). Chromatography is time demanding and sample volumes required are in the range of 0.5–2 mL, which represent a limitation in newborns. We validated a liquid chromatography tandem mass spectrometry (LC-MS/MS) test for the analysis of topiramate in dried blood spot (6 microliters).

Results Using as internal standard D12-topiramate, the calibration curve was linear in the concentration range of 0.0166–1.66 µg/mL, corresponding to 0.5–50 µg/mL in dried blood spot before the 1:30 dilution. The regression equation for the chromatographic method was y  =  32700x + 2370; R2 = 0.9982. The CVs were in the range 2.12–12.37%, and the accuracy ranged from 91.06% to 113.16% in the range of 0.5–50 µg/mL. The analysis was performed in the SRM in which topiramate was detected through transitions from the precursor to the product ion: m/z 340 to m/z 264; m/z 352 to m/z 270 for D12-topiramate. The retention time was 2.2 min in a 3 min analysis. The Bland-Altman test showed a high correlation between the FPIA and the LC-MS/MS assay.

Conclusions We demonstrate that minimal sample preparation, high sensitivity/specificity, and minimal instrument maintenance make this method a candidate for large-scale routine task in newborns.

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