吡啶
化学
脱氧吡啶啉
色谱法
高效液相色谱法
吡啶
尿
生物化学
酶
有机化学
碱性磷酸酶
骨钙素
作者
Elena Monticelli,C.S. Aman,Maria L. Costa,Paola Rota,Daniela Bogdan,Pietro Allevi,Giuliana Cighetti
标识
DOI:10.1016/j.jchromb.2011.07.044
摘要
Pyridinoline (Pyr), deoxypyridinoline (D-Pyr), galactosyl-pyridinoline (Gal-Pyr) and glucosyl–galactosyl pyridinoline (GluGal-Pyr) are enzymatic mature pyridinium crosslinks. Generally, only total Pyr and D-Pyr urinary amounts (free + bound forms) are evaluated by HPLC as indices of bone resorption. This report describes the validation of an HPLC-fluorescence method for the simultaneous evaluation of free Pyr and D-Pyr, together with GluGal-Pyr and Gal-Pyr, in urine of healthy women (n = 20, aged 27–41) and girls (n = 20, aged 5–10). The use of an unnatural D-Pyr homologue, here proposed for the first time as internal standard, and of pure Pyr, D-Pyr, GluGal-Pyr and Gal-Pyr synthesized to be used as primary calibrators, guarantees method specificity and correct crosslink quantification. Urine, spiked with IS, was solid-phase extracted prior to HPLC analysis. Total Pyr and D-Pyr amounts were also evaluated after urine hydrolysis. The HPLC method was validated for selectivity, sensitivity, linearity, precision, accuracy, recovery and stability for all measured crosslinks. Both free and total Pyr and D-Pyr as well as GluGal-Pyr and Gal-Pyr amounts were significantly higher in girls than in women (p < 0.0001), indicating an increased collagen turnover rather than only bone turnover. Gal-Pyr, for the first time evaluated in girls, was under its lower quantification limit (
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