Determination of 2H-enrichment of rat brain interstitial fluid and rat plasma by headspace-gas-chromatography – quadrupole-mass-spectrometry

Abstract

2H2O as nonradioactive, stable marker substance is commonly used in preclinical and clinical studies and the precise determination of 2H2O concentration in biological samples is crucial. However, aside from isotope ratio mass spectrometry (IRMS), only a very limited number of methods to accurately measure the 2H2O concentration in biological samples are routinely established until now.In this study, we present a straightforward method to accurately measure 2H-enrichment of rat brain interstitial fluid (ISF) and rat plasma to determine the relative recovery of a cerebral open flow microperfusion (cOFM) probe, using headspace-gas-chromatography – quadrupole-mass-spectrometry. This method is based on basic-catalyzed hydrogen/deuterium exchange in acetone and detects the 2H-labelled acetone directly by the headspace GC-MS. Small sample volumes and limited number of preparation steps make this method highly competitive. It has been fully validated. 2H enriched to 8800 ppm in plasma showed an accuracy of 98.9% and %Relative Standard Deviation (RSD) of 3.1 with n = 18 over three days and with two operators. Similar performance was obtained for cerebral ISF enriched to 1100 ppm (accuracy: 96.5%, %RSD: 3.1). With this highly reproducible method we demonstrated the successful employment of 2H2O as performance marker for a cOFM probe.