MEASUREMENT OF TOTAL AND SEPARATE STEREOISOMERS OF DIAMINOPIMELIC ACID IN RUMEN BACTERIA BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

Citation
Am. Elwaziry et al., MEASUREMENT OF TOTAL AND SEPARATE STEREOISOMERS OF DIAMINOPIMELIC ACID IN RUMEN BACTERIA BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY, Journal of chromatography B. Biomedical applications, 677(1), 1996, pp. 53-59
Citations number
17
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Chemistry Analytical","Biochemical Research Methods
Journal title
Journal of chromatography B. Biomedical applications
ISSN journal
1572-6495 → ACNP
Volume
677
Issue
1
Year of publication
1996
Pages
53 - 59
Database
ISI
SICI code
Abstract
New high-performance liquid chromatographies were examined and applied for the analyses of the total (with one peak) and separate three ster eoisomers of 2,6-diaminopimelic acid (DAP) in hydrolysed mixed rumen b acteria. The methods start with the reaction of DAP with 1-fluoro-2,4- dinitrophenylalanine amide for derivatisation. A mixture of 0.05 M tri ethylamine phosphate (pH 3.0) and acetonitrile (72:28, v/v) was used a s an isocratic mobile phase for the total DAP determination (method 1) , and a mixture of both solutions (78.5:21.5, v/v) for the determinati on of the separate three stereoisomers of DAP (method 2). The flow-rat e was 1 ml/min; column, Merck LiChrospher 100 RP-18 (250 X 4 mm I.D.) of 5 mu m particle size; column temperature, 40 degrees C; wavelength of detector, 325 nm. The retention times were 17.5 min for total DAP ( method 1), and 39.4, 63.1 and 77.7 for meso-, LL- and DD-DAP, respecti vely (method 2). Lysine can also be determined by method 2 and the ret ention time was 122.8 min. The minimum detectable limit was 2.5 mu M. The average analytical recoveries were 98.6% for total DAP and 99.1%, 100.4% and 100.2% for meso-, LL- and DD-DAP, respectively. The content of total DAP of the hydrolysed rumen bacteria collected from three go ats fed lucerne cube and concentrate ranged from 25.55 to 27.36 mu mol /g bacterial DM (method 1). The contents of the separate stereoisomers of DAP in the hydrolysed rumen bacteria from the three goats ranged f rom 19.64 to 22.06 and from 4.98 to 5.21 mu mol/g bacterial DM for mes o- and LL-DAP, respectively (method 2). DD-DAP was not detected.