Determination of pyridinoline and deoxypyridinoline in urine, with special attention to retaining their stability.

Urinary excretion of the pyridinium crosslinks pyridinoline (Pyr) and deoxypyridinoline (Dpyr) is used as a biochemical marker of bone resorption. The present study was undertaken to determine the long-term stability of these compounds in stored urine, using the HPLC method. Systematic investigation of their chemical stability in urine demonstrated that both the free and conjugated forms of Pyr and Dpyr are extremely stable: No significant changes were observed after 6 weeks at -20 degrees C storage (e.g., free Pyr 9.6 +/- 1.2 mumol/mol creatinine (before) and 10.6 +/- 3.2 (after); free Dpyr 2.3 +/- 0.2 mumol/mol creatinine (before) and 2.5 +/- 1.2 (after)). These results predict stability of urines stored for 10-20 years at -20 degrees C in the dark. Also, freezing and thawing as many as 10 times had no effect on the concentrations of the crosslinks. Study of the stability of the excretion pattern in healthy women showed substantially higher variations in excretions of free and total Dpyr (18% and 13%, respectively) than of Pyr (10% for both forms).