University of Tasmania
Browse

File(s) under permanent embargo

Determination of melatonin and monoamines in rat pineal using reversed-phase ion-interaction chromatography with fluorescence detection

journal contribution
posted on 2023-05-26, 16:20 authored by Mills, MH, Finlay, DC, Paul HaddadPaul Haddad
A method is reported for the ion-interaction, reversed-phase separation of 24 compounds (chiefly monoamines) arising from the metabolism of tyrosine and tryptophan. These compounds were separated as two groups. The first group comprised 3,4-dihydroxyphenylethylene glycol, tyrosine, 3-methoxy-4-hydroxyphenyl glycol, 5-hydroxytryptophan, norepinephrine, 3,4-dihydroxyphenylacetic acid, epinephrine, 5-hydroxyindole-3-acetic acid, homovanillic acid, 5-hydroxytryptophol, dopamine, tryptophan, N-acetylserotonin, N-acetyltryptophan, 5-methoxytryptophan and serotonin. The mobile phase consisted of a 6.8:93.2 (v/v) mixture of acetonitrile and an aqueous solution containing 0.16 M ammonium phosphate, 0.06 M citric acid, 0.15 mM disodium EDTA, 10 mM dibutylamine and 6 mM sodium 1-octanesulphonate at pH 4.50. The second group of compounds comprised 6-hydroxymelatonin, 5-methoxyindole-3-acetic acid, indole-3-acetic acid, 5-methoxytryptamine, tryptamine, 5-methoxytryptophol, melatonin and tryptophol. The mobile phase consisted of a 16:84 (v/v) mixture of acetonitrile and an aqueous solution containing 0.05 M ammonium phosphate, 0.05 M citric acid, 0.15 mM disodium EDTA, 25 mM dibutylamine and 5 mM sodium 1-octanesulphonate at pH 5.30. Detection was by fluorescence measurement (˜í¬™ex = 280 nm, ˜í¬™em = 340 nm). The proposed method exhibited linear calibration over the biochemically significant concentration range, with detection limits in the 10‚Äö-200 pg range. Excellent precision for peak areas and retention times was observed, even over a period of 24 h. The applicability of amperometric detection (at 0.72V) is also demonstrated. The method is applied to the determination of monoamines in individual rat pineals. Low nanogram levels of tyrosine, norepinephrine, 5-hydroxyindole-3-acetic acid, tryptophan, serotonin and 6-hydroxymelatonin, and picogram levels of 5-hydroxytryptophan, 5-hydroxytryptophol, 5-methoxyindole-3-acetic acid, indole 3-acetic acid, 5-methoxytryptophol and melatonin were indicated in most of the samples.

History

Publication title

Journal of Chromatography - Biomedical Applications

Volume

564

Article number

1

Number

1

Pagination

93-102

ISSN

0378-4347

Publication status

  • Published

Rights statement

The definitive version is available at http://www.sciencedirect.com

Repository Status

  • Restricted

Usage metrics

    University Of Tasmania

    Categories

    No categories selected

    Exports

    RefWorks
    BibTeX
    Ref. manager
    Endnote
    DataCite
    NLM
    DC