Full Short Communication

Chromatographia

, Volume 68, Issue 1, pp 101-105

Measurement of Acetylcholine in Rat Brain Microdialysates by LC–Isotope Dilution Tandem MS

  • Laszlo ProkaiAffiliated withDepartment of Molecular Biology and Immunology, University of North Texas Health Science Center Email author 
  • , Petr FryčákAffiliated withDepartment of Molecular Biology and Immunology, University of North Texas Health Science Center
  • , Stanley M. StevensJr.Affiliated withDepartment of Molecular Biology and Immunology, University of North Texas Health Science Center
  • , Vien NguyenAffiliated withDepartment of Molecular Biology and Immunology, University of North Texas Health Science Center

Rent the article at a discount

Rent now

* Final gross prices may vary according to local VAT.

Get Access

Abstract

An LC–MS–MS method was developed for measuring acetylcholine (ACh) in an aqueous medium using reversed-phase ion-pair chromatography, electrospray ionization on a quadrupole ion trap instrument and a tetradeuterated analogue (ACh-1,1,2,2-d4) as an internal standard. A rapid separation was achieved on a 5-cm long octadecylsilica column (2.1 mm i.d.) by employing heptafluorobutyric acid (0.1% v/v) as an ion-pairing agent and requiring 10% v/v acetonitrile in 20 mM ammonium formate buffer under isocratic elution at 200 μl min−1 flow rate. The instrument’s response was calibrated with samples containing known mole ratios of ACh and ACh-1,1,2,2-d4 in an artificial cerebrospinal fluid, which afforded the conclusion that analyte concentrations could be determined by multiplying the measured analyte to internal standard ion-current ratio with the known molar concentration of the ACh-1,1,2,2-d4 added. The rapid and simple assay was tested by measuring the basal neurotransmitter concentration in rat brain microdialysates without the use of a cholinesterase inhibitor upon sample collection.

Keywords

Column liquid chromatography–tandem mass spectrometry Electrospray ionization Isotope dilution In vivo intracranial microdialysis Acetylcholine