High-performance liquid-chromatographic separation and fluorescence measurement of biogenic amines in plasma, urine, and tissue

Thomas P Davis, C. W. Gehrke, C. W. Gehrke, T. D. Cunningham, K. C. Kuo, K. O. Gerhardt, H. D. Johnson, C. H. Williams

Research output: Contribution to journalArticle

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Abstract

The authors describe a high-performance liquid-chromatographic method for measuring histamine, norepinephrine, octopamine, normetanephrine, dopamine, serotonin, and tyramine in plasma (2 ml), brain (0.2 g), or urine. These amines are modified by pre-column derivatization with o-phthalaldehyde, which stabilizes the molecules, facilitates extraction, and improves detection of nanogram amounts. Before separation, samples were neutralized with KOH and immediately derivatized and extracted into ethyl acetate, in which derivatives were stable for longer than 24 h. Interfering amino acids were removed from ethyl acetate by partitioning twice with Na 2HPO 4 buffer (pH 10.0). Separation was complete in about 90 min on a 'μ Bondapak/phenyl' column, with which a stepwise gradient of methanol/phosphate buffer (pH 5.1) was used. A variable-wavelength fluorometer was used (exciting wavelength, 340 nm; emission wavelength, 480 nm). Amount and response were linearly related from 1 to 200 pmol. Precision (CV) for retention times was 1%, for derivatization and injection 2.5%. Analytical recoveries of the seven amines from 2 ml of plasma fortified with 200 pmol averaged 65% (CV ~ 8%). Data on rat-brain tissue samples are compared with results by the trihydroxyindole method. Application of the method to urine from normal persons and a patient with a brain tumor is demonstrated.

Original languageEnglish (US)
Pages (from-to)1317-1324
Number of pages8
JournalClinical Chemistry
Volume24
Issue number8
StatePublished - 1978
Externally publishedYes

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Biogenic Amines
Brain
Fluorescence
Urine
Tissue
Plasmas
Wavelength
Amines
Buffers
Liquids
Normetanephrine
Fluorometers
o-Phthalaldehyde
Octopamine
Tyramine
Brain Neoplasms
Histamine
Methanol
Rats
Tumors

ASJC Scopus subject areas

  • Clinical Biochemistry

Cite this

Davis, T. P., Gehrke, C. W., Gehrke, C. W., Cunningham, T. D., Kuo, K. C., Gerhardt, K. O., ... Williams, C. H. (1978). High-performance liquid-chromatographic separation and fluorescence measurement of biogenic amines in plasma, urine, and tissue. Clinical Chemistry, 24(8), 1317-1324.

High-performance liquid-chromatographic separation and fluorescence measurement of biogenic amines in plasma, urine, and tissue. / Davis, Thomas P; Gehrke, C. W.; Gehrke, C. W.; Cunningham, T. D.; Kuo, K. C.; Gerhardt, K. O.; Johnson, H. D.; Williams, C. H.

In: Clinical Chemistry, Vol. 24, No. 8, 1978, p. 1317-1324.

Research output: Contribution to journalArticle

Davis, TP, Gehrke, CW, Gehrke, CW, Cunningham, TD, Kuo, KC, Gerhardt, KO, Johnson, HD & Williams, CH 1978, 'High-performance liquid-chromatographic separation and fluorescence measurement of biogenic amines in plasma, urine, and tissue', Clinical Chemistry, vol. 24, no. 8, pp. 1317-1324.
Davis, Thomas P ; Gehrke, C. W. ; Gehrke, C. W. ; Cunningham, T. D. ; Kuo, K. C. ; Gerhardt, K. O. ; Johnson, H. D. ; Williams, C. H. / High-performance liquid-chromatographic separation and fluorescence measurement of biogenic amines in plasma, urine, and tissue. In: Clinical Chemistry. 1978 ; Vol. 24, No. 8. pp. 1317-1324.
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