A simple, sensitive, and fast matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) method was developed for the determination of acetylcholine (ACh) and choline (Ch) in microdialysis samples. An optimized dried droplet preparation with the common matrix α-cyano-4-hydroxycinnamic acid (CHCA) was used. Limits of detection [signal-to-noise ratio (S/N) = 3] of 0.3 fmol/?L for ACh and 20 fmol/?L for Ch were found using standards diluted in artificial cerebrospinal fluid (aCSF). The limit of quantification (LOQ) for ACh was 1 fmol/?L, and excellent linearity (R2 = 0.9996) was maintained over the range of 1?1000 fmol/?L. Choline was quantified over the range of 0.1?50 pmol/?L, also with excellent linearity (R2 = 0.9995). Good accuracy and precision were obtained for all concentrations within the range of the standard curve. The developed method was successfully used for the determination of ACh and Ch in mouse brain microdialysis samples. The samples were quantified by a calibration curve and also by the method of standard addition. Despite the high salt content of the perfusion fluid (>150 mM), a direct measurement was possible. To the authors? knowledge, this is the first published method to determine acetylcholine and choline in microdialysis samples by MALDI-TOF MS. The method presented significantly reduces the needed analysis time, as only approximately 10 s/sample is required, and it is also possible to improve the temporal resolution, because only ?1 ?L of sample is needed.
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