MEASUREMENT OF ALLANTOIN IN URINE AND PLASMA BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH PRECOLUMN DERIVATIZATION

A method is reported for determination of allantoin in urine and plasm a based on high-performance liquid chromatography (HPLC) and pre-colum n derivatization. In the derivatization procedure, allantoin is conver ted to glyoxylic acid which forms a hydrazone with 2,4-dinitrophenylhy drazine. The hydrazone appears as syn and anti isomers at a constant r atio. These derivatives are separated by HPLC using a reversed-phase C 18 column from hydrazones of other keto acids possibly present in urin e and plasma and then monitored at 360 nm. All components were complet ely resolved in 15 min. Both the reagents and derivatization products are stable. Recovery of allantoin added to urine and plasma was 95 +/- 3.7% (n = 45) and 100 +/- 7.5% (n = 64), respectively. The lowest all antoin concentration that gave a reproducible integration was 5 mumol/ l. The between-assay and within-day coefficients of variation were 2.8 and 0.6%, respectively.