Use of an immobilized enzyme reactor for the analysis of residues of 17 alpha-methyltestosterone in trout by high-performance liquid chromatography.
We have recently observed in trout that 48 h after ingestion of a single dose of [14C]-17 alpha-methyltestosterone ([14C]-MT), 25% of the radioactivity was still in the carcass, corresponding to metabolites of 17-MT. These compounds have no appreciable chromophore, fluorophore or electrophore, therefore the usual detection systems was not satisfactory for their analysis. Consequently a method was developed for high-performance liquid chromatographic separation and detection of two of the major tissue metabolites of 17-MT: 5 alpha-androstane-17 alpha-methyl-3 alpha,17 beta-diol and 5 beta-androstane-17 alpha-methyl-3 alpha,17 beta-diol. A column of immobilized 3 alpha-hydroxysteroid dehydrogenase was prepared and used for detection. The NADH produced from 3-hydroxysteroids by this immobilized enzyme reactor was monitored fluorimetrically. The detection limit of this method, as obtained from the calibration curve, was at the picomole level; the limit of quantification in muscle was 1 microgram/kg, at a signal-to-noise ratio of 4.
Cravedi
J P
JP
I.N.R.A.-Laboratoire des Xénbiotiques, Toulouse, France.
Delous
G
G
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004800:Enzymes, Immobilized; D008777:Methyltestosterone / Q000032:analysis* / Q000378:metabolism; D009132:Muscles / Q000737:chemistry / Q000378:metabolism; D014337:Trout / Q000378:metabolism*
D004800:Enzymes, Immobilized; D008777:Methyltestosterone