Internal standard method for the measurement of choline and acetylcholine by capillary electrophoresis with electrochemical detection

Abstract

An internal standard method has been developed for the determination of the neurotransmitter acetylcholine and/or its metabolic precursor choline. This approach couples the high separation efficiency of capillary electrophoresis with the sensitivity and selectivity of electrochemical detection at an enzyme-modified electrode. Indirect electrochemical detection is accomplished at a 25 μm platinum electrode modified by cross-linking the enzymes choline oxidase and acetylcholinesterase with glutaraldehyde. Although in this simple form of electrode fabrication there is a gradual loss of response from the electrochemical detector with time, accurate quantitation is achieved by the addition of butyrylcholine, which is also a substrate for acetylcholinesterase, as an internal standard. A linear response is achieved between 0 and 125 μ M with a limit of detection of 2 μ M (25 fmol). The utility of this method was demonstrated by monitoring the kinetics of choline uptake in synaptosomal preparations.