Selective solid-phase extraction of urinary 2,3-dinor-6-ketoprostaglandin F1 alpha for determination with radioimmunoassay.
This paper describes a method for selective two-step solid-phase extraction of urinary 2,3-dinor-6-ketoprostaglandin F1 alpha for reliable determination with radioimmunoassay. In the immunoreactivity profile of non-selectively extracted urine after HPLC separation, over 90% of the total 2,3-dinor-6-ketoprostaglandin F1 alpha immunoreactivity consisted of interfering material coeluting with 6-ketoprostaglandin F1 alpha and 2,3-dinor-6-ketoprostaglandin F1 alpha. Among the alkyl silica sorbents studied (methyl, butyl, octyl, and octadecyl), an efficient separation of 2,3-dinor-6-ketoprostaglandin F1 alpha from 6-ketoprostaglandin F1 alpha and the lowest immunoreactive concentration of analyte were achieved in extraction on the methyl silica sorbent by elution of 2,3-dinor-6-ketoprostaglandin F1 alpha with chloroform: hexane (85:15, v/v) from the cartridge. The proportion of specific immunoreactivity could be further increased by two-step extraction of sample on methyl silica cartridges, first at pH 3 and then at pH 10 using diethyl ether:hexane (85:15, v/v) and chloroform as eluent, respectively. After this, a high correlation was found with concentrations of samples determined by radioimmunoassay using three different antisera. A significant correlation of values was also observed between samples measured by radioimmunoassay and those measured by GC-MS. The values of 12-h excretion of 2,3-dinor-6-ketoprostaglandin F1 alpha in eight volunteers (268 +/- 204 ng/g creatinine, mean +/- SD) as well as the inhibitory effect of acetylsalicylic acid (74 +/- 12%) are in accordance with those reported in the literature. This selective extraction procedure provides a high validity in radioimmunoassay without requiring subsequent TLC or HPLC purification.
Riutta
A
A
Department of Biomedical Sciences, University of Tampere, Finland.
Nurmi
E
E
Weber
C
C
Hansson
G
G
Vapaatalo
H
H
Mucha
I
I
D015121:6-Ketoprostaglandin F1 alpha / Q000031:analogs & derivatives* / Q000302:isolation & purification / Q000652:urine; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D008297:Male; D011863:Radioimmunoassay / Q000379:methods; D011865:Radioisotope Dilution Technique; D012016:Reference Values; D012997:Solvents; D014316:Tritium
D007202:Indicators and Reagents; D012997:Solvents; D014316:Tritium; D015121:6-Ketoprostaglandin F1 alpha; C028290:2,3-dinor-6-ketoprostaglandin F1alpha