Solid-phase extraction of urinary 11-dehydrothromboxane B2 for reliable determination with radioimmunoassay

Asko Riutta, I. Muchá, Heikki Vapaatalo

Research output: Contribution to journalArticle

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Abstract

In this paper we elaborate a one-step procedure for the selective extraction of urinary 11-dehydrothromboxane B2 on octylsilyl silica cartridges for reliable determination with radioimmunoassay. The immunoreactivity profile of nonselectively extracted urine after HPLC separation showed that as much as 70% of the total 11-dehydrothromboxane B2 immunoreactivity comigrates with polar interfering material. Its amount could be considerably decreased using acetonitrile:water (18:82, v/v) as wash solvent before elution of 11-dehydrothromboxane B2 from the cartridge. Alternatively, very high immunoreactive purity was achieved without the preceding wash step by selective elution of the analyte with dichloromethane:hexane (70:30). After both optimized steps in the extraction procedure were combined, immunoreactivity was found only in HPLC fractions corresponding to the retention volume of authentic 11-dehydrothromboxane B2. The homogeneity of this immunoreactivity was confirmed by two-step HPLC separation. A significant correlation of values was observed between samples measured after extraction and those measured after subsequent HPLC purification. A high correlation was also found with concentrations determined by radioimmunoassay using four different antisera. The values of 24 h excretion of 11-dehydrothromboxane B2 in 10 male volunteers (595 ± 114 ng/g creatinine, mean ± SD) as well as the inhibitory effect of acetylsalicylic acid (80 ± 13%) closely correspond with those reported in the literature. This selective extraction procedure provides a high validity in radioimmunoassay without requiring any further purification step.

Original languageEnglish
Pages (from-to)299-305
Number of pages7
JournalAnalytical Biochemistry
Volume202
Issue number2
DOIs
Publication statusPublished - May 1 1992

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Solid Phase Extraction
Radioimmunoassay
High Pressure Liquid Chromatography
Purification
Methylene Chloride
Hexanes
Silicon Dioxide
Aspirin
Immune Sera
Volunteers
Creatinine
11-dehydro-thromboxane B2
Urine
Water

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Solid-phase extraction of urinary 11-dehydrothromboxane B2 for reliable determination with radioimmunoassay. / Riutta, Asko; Muchá, I.; Vapaatalo, Heikki.

In: Analytical Biochemistry, Vol. 202, No. 2, 01.05.1992, p. 299-305.

Research output: Contribution to journalArticle

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