We discuss the use of 'high-performance' liquid chromatography with electrochemical detection for the quantification of serotonin (5-hydroxytryptamine) and metabolically related products (i.e., 5-hydroxyindoleacetic acid and 5-hydroxytryptophan) in biological matrices. Two methods are described: one for routine quantification of nanogram amounts or more; the other a two-column purification-separation ('trace enrichment') technique for determining subnanogram amounts of these compounds. Various factors affecting the liquid-chromatographic separation and quantification of 5-hydroxyindoles are detailed, including selection of appropriate internal standard, sample purification, and effects of changes in mobile phase composition, e.g., ion-pair reagent, organic modifier content, and pH. We describe in detail the trace-enrichment method for determinations in the range of 25 pg to 25 ng, and discuss the general factors to consider when developing or modifying an assay for specific applications.
All Science Journal Classification (ASJC) codes
- Clinical Biochemistry
- Biochemistry, medical