An Improved Gas Chromatographic Method for the Determination of Urinary Acetylpolyamines

Abstract

Our previous method for the determination of acetylpolyamines in human urine by gas chromatography has been improved. A urine sample fortified with the internal standard (monoacetylnorspermidine) was directly treated with ethyl chloroformate in an alkaline medium, thereby converting the acetylpolyamines to the N-ethoxycarbonyl derivatives. After extraction of the resulting derivatives with chloroform, the combined extract was applied to a silica gel minicolumn to remove interfering substances. The derivatives of acetylpolyamines thus isolated were determined by gas chromatography with nitrogen-selective detection under temperature-programmed conditions. Calibration curves for acetylpolyamines were linear over the range of 0.2-10 nmol examined. The sensitivity of this method is over 100-fold higher than that of the previous method, so that the acetylpolyamines in human urine could be accurately determined in a sample as small as 200 μl. The use of a nitrogen-selective detector and concomitant procedural modifications greatly reduced the time required for analysis.