Solid phase extraction and simultaneous high performance liquid chromatographic determination of antipyrine and its major metabolites in urine

Abstract

A reversed phase gradient high performance liquid chromatographic method utilizing solid phase extraction has been described for the simultaneous determination of antipyrine (AP), 4-hydroxyantipyrine (COHAP), norantipyrine (NorAP) and 3-hydroxymethylantipyrine (3-OHMAP) in human urine after hydrolysis with pglucuronidase. The C-18 sorbent cartridges were conditioned and urine samples were applied, washed with 1 X 4 mL of phosphate buffer and eluted with 3 X 100 pL of 20% v/v of acetonitrile in methylene chloride. The eluent was evaporated to dryness, reconstituted in 100 pL phosphate buffer and injected. The calibration ranges were 2.0-250 pg/mL (AP), 2.5-250 pg/mL (NorAP), 2.0-250 pg/mL (3-OHMAP) and 5.0-500 pg/mL (4-OHAP) with regression coefficients of 0.998 or greater. Specificity was indicated by the absence of interferences in chromatogram of blank urine from normal as well as cirrhotic patients. The average recovery was 86.7% for AP, 90.5% for NorAP, 85.2% for 4-OHAP and 74.2% for 3-OHMAP. The within-assay precision as indicated by the reproducibility of the assayed spiked urine was less than 9% in all cases and the between-assay precision was less than 12%. The method was applied to studies on antipyrine metabolism in stable cirrhotic patients. Following administration of a single oral dose of about 1000 mg to nine stable cirrhotic patients and eight age-matched healthy volunteers, the cumulative account excreted in the urine up to 48 h for AP and the three metabolites was comparable to other literature reports.

Pharmacokinetics of minoxidil in patients with cirrhosis and healthy volunteers

“…High-performance liquid chromatography with UV detection was used to determine concentrations of antipyrine in plasma, and antipyrine and three metabolites (4-hydroxyantipyrine, norantipyrine, and 3-hydroxymethylantipyrine) in urine [16,17]. Antipyrine concentrations were linear between 0.2 and 40 mg mL − 1 in plasma, and the between-run CV was 5 7.2%.…”