In vitro and in vivo microdialysis calibration for the measurement of carbamazepine and its metabolites in rat brain tissue using the internal reference technique

J Neurosci Methods. 1993 Sep;49(3):167-73. doi: 10.1016/0165-0270(93)90120-g.


Microdialysis, as in vivo sampling technique, can be used for determining exogenous compounds in the extracellular fluid of freely moving animals and in humans. Usually, calibration of the microdialysis probe is determined by in vitro relative recovery (RR) (dialysate extraction fraction). However, due to different diffusion properties of the compound in tissue, the RR in vivo is different from the RR in vitro. In this study, the evaluation of the internal reference technique as in vivo calibration method was established. To determine the RR in vivo, the relative loss (RL) was defined as the loss of a compound from the perfusate. RL was determined in vitro and in vivo by adding an internal standard (IS) to the perfusate. This internal reference technique was applied for the determination of carbamazepine (CBZ) and its 2 major metabolites, carbamazepine-10,11-epoxide (CBZ-EPO) and trans-10,11-dihydroxy-10,11-dihydro-carbamazepine (CBZ-DIOL) using 2-methyl-5H-dibenz(b,f)azepine-5-carboxamide (m-CBZ) as IS. In vitro and in vivo, the loss of m-CBZ and the recovery of CBZ are identical. The ratios of the RR of CBZ-EPO and CBZ-DIOL to the RL of m-CBZ are constant, in vitro and in vivo. Therefore, m-CBZ can be used as IS for CBZ, CBZ-EPO and CBZ-DIOL determinations in brain tissue. It is shown that the internal reference technique is a useful method to estimate the true concentration of exogenous compounds in the extracellular space of tissues.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Brain / metabolism*
  • Calibration
  • Carbamazepine / analogs & derivatives
  • Carbamazepine / metabolism*
  • In Vitro Techniques
  • Male
  • Microdialysis / methods*
  • Rats
  • Rats, Wistar
  • Reference Standards


  • Carbamazepine
  • 10,11-dihydro-10,11-dihydroxy-5H-dibenzazepine-5-carboxamide
  • carbamazepine epoxide