PLASMA METHOTREXATE AS DETERMINED BY LIQUID-CHROMATOGRAPHY, ENZYME-INHIBITION ASSAY, AND RADIOIMMUNOASSAY AFTER HIGH-DOSE INFUSION

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  • 1 January 1980
    • journal article
    • research article
    • Vol. 26  (6) , 734-737
Abstract
Three techniques for measuring methotrexate [in the blood of children with osteosarcoma undergoing chemotherapy] show various cross reactivities with methotrexate-related compounds: high-pressure liquid chromatography, by principle, is virtually specific for methotrexate; the enzyme-inhibition assay quantitates methotrexate, methotrexate diglutamate and methotrexate triglutamate equally well, but has a 10% cross reactivity with 4-amino-4-deoxy-N10-methylpteroic acid and 1% with 7-hydroxymethotrexate; radioimmunoassay shows an equal cross reactivity with methotrexate, 4-amino-4-deoxy-N10-methylpteroic acid, methotrexate diglutamate and triglutamate, and a 5-10% cross reactivity with 7-hydroxymethotrexate. Radioimmunoassay almost always yielded the highest values for methotrexate, followed by enzyme-inhibition assay, then liquid chromatography. The presence of 2 methotrexate-related compounds, 7-hydroxymethotrexate and 4-amino-4-deoxy-N10-methylpteroic acid, was confirmed in human urine samples and quantitated in patients'' plasma by liquid chromatography, the respective maximum plasma concentrations being 250 and 16 .mu.mol/l. Materials cross reacting with methotrexate in radioimmunoassay of chromatographic fractions from plasma were also noted in fractions corresponding to methotrexate diglutamate and triglutamate peaks, in quantities estimated to be 47 and 30 nmol/l methotrexate equivalents, respectively. 7-Hydroxymethotrexate is eliminated more slowly than methotrexate and its production increases with dosages of methotrexate.