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Microbore high-performance liquid chromatographic method for measuring acetylcholine in microdialysis samples:
1Department of Biological Research, Boehringer Ingelheim KG, Germany.
Summary
This study details a high-performance liquid chromatography method for measuring acetylcholine. Electrode sensitivity decreases over time, requiring regular cleaning and calibration for accurate acetylcholine quantification.
Area of Science:
- Analytical Chemistry
- Neurochemistry
Background:
- Acetylcholine measurement is crucial in microdialysis studies.
- High-performance liquid chromatography (HPLC) with electrochemical detection is a common technique.
Purpose of the Study:
- To validate an HPLC method for acetylcholine measurement in microdialysis samples.
- To investigate the cause of decreased acetylcholine peak height during method validation.
Main Methods:
- Microbore HPLC with electrochemical detection.
- Analysis of acetylcholine standards and microdialysis samples.
- Electrode cleaning with ethanol and methanol.
Main Results:
- An excellent linear relationship was observed between acetylcholine concentration and peak height (r=0.99998).
- A decrease in electrode sensitivity, indicated by reduced acetylcholine peak height and a brown electrode coating, was identified.
- Electrode sensitivity loss was confirmed with a second HPLC system.
Conclusions:
- The platinum electrode's sensitivity decreases over time, impacting acetylcholine measurement accuracy.
- Periodic cleaning of the platinum electrode with ethanol or methanol is recommended.
- Regular calibration with external acetylcholine standards is essential for reliable quantitation.