ION-EXCHANGE HPLC DETERMINATION OF PYRIDINIUM CROSS-LINKS IN URINE ASMARKERS OF BONE-RESORPTION

Citation
P. Spacek et al., ION-EXCHANGE HPLC DETERMINATION OF PYRIDINIUM CROSS-LINKS IN URINE ASMARKERS OF BONE-RESORPTION, Journal of liquid chromatography & related technologies, 20(12), 1997, pp. 1921-1930
Citations number
22
Categorie Soggetti
Chemistry Analytical","Biochemical Research Methods
ISSN journal
10826076
Volume
20
Issue
12
Year of publication
1997
Pages
1921 - 1930
Database
ISI
SICI code
1082-6076(1997)20:12<1921:IHDOPC>2.0.ZU;2-V
Abstract
The aim of this study was to improve the published method(1) for rapid and sensitive determination of the urinary pyridinoline as well as de oxypyridinoline, i.e. the crosslinking elements of bone and cartilage collagen. This method is based on the HPLC analysis of previously prep ared urinary hydrolysates. Urine hydrolysate (6 N HCl, 110 degrees C, 16 hours) was purified by selective fractionation using washing with a mixture of water -acetic acid-butanol (1-1-4 by volume) on small disp osable columns filled with microgranular cellulose. Liquid chromatogra phy was based on ion-exchange counteractions and was properly performe d with the use of a strong cation exchanger as a stationary phase and with a buffered solution at pH=3.35 as the mobile phase, by keeping th e oven temperature at 48 degrees C and with the injection of a volume of 10 mu l. The time desired for HPLC determination alone didn't excee d 15 min and by using a fluorescence detector (Shimadzu RF 535) set at 297 and 400 nm for excitation and emmision wavelengths, respectively, it reached a sensitivity of about 200 femtomoles for both the crossli nks.