A High-Performance Liquid Chromatography Method for the Detection of Diaminopimelic Acid in Urine

✍ Scribed by Gabriel Borruat; Claude-Alain H. Roten; Laurent-Bernard Fay; Dimitri Karamata

• Publisher: Elsevier Science
• Year: 2001
• Tongue: English
• Weight: 73 KB
• Volume: 291
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.2001.5016

✦ Synopsis

We describe a quantitative assay for diaminopimelic acid (DAP) in urine. It involves (i) hydrolysis of urine samples, (ii) purification by several different liquid chromatography steps, and (iii) analysis by high-performance liquid chromatography on a reversed-phase C18 column. Tritiated-DAP, the internal standard, allows one to precisely follow the DAP-containing fractions and to determine the yield during purification. Sensitive and relatively accurate quantification of DAP, with a threshold of 50 fmol, is based on ion-pairing properties of eluants and ortho-phthaldialdehyde derivatization. The presence of DAP in relevant fractions was confirmed by combined gas chromatography and mass spectrometry. The DAP concentration in adult human urine pooled over 24 h ranges from 0.69 to 2.01 microM, a result in fair agreement with previously published values obtained by ninhydrin derivatization or gas chromatography.