DETERMINATION OF 5-HYDROXYTRYPTAMINE, 5-HYDROXYINDOLEACETIC ACID AND TRYPTOPHAN IN PLASMA AND URINE BY HPLC WITH FLUOROMETRIC DETECTION

Using native fluorescence detection, 5-hydroxytryptamine (5-HT), 5-hyd roxyindoleacetic acid (5-HIAA) and tryptophan were resolved from thems elves and other naturally occurring compounds using reversed-phase HPL C within 5 min. Deproteinated platelet-poor plasma (PPP) and crude dil uted urine were injected directly into the chromatograph, Careful sele ction of the HPLC column is important and various octadecyl silica (OD S) and base deactivated silic (BDS) columns were evaluated. Pre-treatm ent of an ODS column with tetrabutylammonium ions gave good selectivit y. Between pH 5 and 6 the compounds were well resolved from each other . The limit of quantitative detection of 5-HT and 5-HIAA was 3.5 nmol/ L. The overall chromatogram obtained using native fluorescence is clea ner than that obtained with the more commonly employed electrochemical (EC) systems although the chromatography is effectively the same. For analysis of 5-HT in plasma, collection in EDTA was more efficient tha n lithium heparin. Plasma 5-HT in healthy volunteers was mean 61 (SD = +/- 73) nmol/L, n = 20; urine 5-HIAA gave mean 28.95 (SD = +/- 0.98) mu mol/L, (n = 12). Whole blood 5-HT analysis is unreliable in compari son with platelet-poor plasma.