ABSTRACT
An ion-pair liquid chromatography-electrospray mass spectrometry (LC-ESI-MS) method with in vivo microdialysis for the determination of free-form amphetamine in rat brain has been developed. A microdialysis probe was surgically implanted into the striatum of the rat and artificial cerebrospinal fluid (aCSF) was used as the perfusion medium. Samples were collected and then analyzed off-line by LC-ESI-MS. A reversed phase C18 column was employed for LC separation. Trifluoroacetic acid (TFA) was added in the mobile phase (acetonitrile-water, 10:90, v/v) as an ion-pair reagent. The ion-pair process disguises the protonated amphetamine cations from the ESI-MS electric field as neutral molecules. Post-column addition of volatile organic acid was utilized to minimize TFA signal suppression effect on ESI-MS detection. More than six-fold enhancement of ESI-MS response was achieved by the post-column addition of propionic acid. Good linearity (0.01-1.00 microg/ml, r2 = 0.99) and detection limit (0.002 microg/ml) were determined. Good precision and accuracy were obtained. The applicability of this newly developed method was demonstrated by continuous monitoring of amphetamine concentrations in rat brain after a single 3.0 mg/kg i.p. administration.
Subject(s)
Amphetamine/analysis , Brain/metabolism , Central Nervous System Stimulants/analysis , Amphetamine/cerebrospinal fluid , Amphetamine/pharmacokinetics , Animals , Central Nervous System Stimulants/cerebrospinal fluid , Central Nervous System Stimulants/pharmacokinetics , Chromatography, Ion Exchange , Male , Microdialysis , Rats , Rats, Sprague-Dawley , Reference Standards , Reproducibility of Results , Solutions , Spectrometry, Mass, Electrospray IonizationABSTRACT
A gas-chromatographic method for simultaneously measuring p-hydroxyamphetamine (pOA) against amphetamine (A) in plasma and CSF is presented. The time course of body temperature (Tb), stereotyped behavior (St), and A and pOA levels in plasma and CSF were studied after administration of 0.6 and 1.5 mg/kg p.o. of A to dogs. Stereotyped behavior reached maximal value 2.5 h after A, as did levels of A in CSF. The A levels in CSF decreased steadily in the following hours and simultaneously with the levels of A in plasma. St remained elevated and began to decrease after 6.5 h. The relationship between St and amounts of A was not linear but exponential. This suggest that both A and its metabolite contributed to this effect. In fact, a linear relationship was found between St and the amounts of pOA in CSF. Body temperature had a time course similar to A plasma levels, reaching peak value after 1.5 h and declining thereafter simultaneously with A. A linear relationship was found between Tb and the amounts of A in plasma. Thus Tb seems to be a peripheral A effect related to the presence of the drug in plasma.
