反相高效液相色谱法测定尿中吡啶醚和脱氧吡啶醚

Analysis of the Pyridinoline and Deoxypyridinoline in Urine by Reversed Phase High Performance Liquid Chromatography (RP HPLC)

尿中吡啶醚（ｐｙｒｉｄｉｎｏｌｉｎｅ，ＰＹＤ）和脱氧吡啶醚（ｄｅｏｘｙｐｙｒｉｄｉｎｏｌｉｎｅ，ＤＰＤ）是骨代谢特异的生化指标。应用高效液相色谱法（ＨＰＬＣ）建立了尿中ＰＹＤ和ＤＰＤ的测定方法。尿液用６ｍｏｌ／ＬＨＣｌ水解后，以纤维素ＣＦ１小柱提取，然后用ＨＰＬＣ测定；色谱系统为ＳｐｈｅｒｉｓｏｒｂＣ１８反相色谱柱，流动相组成为１５％甲醇添加０．１％七氟丁酸，流速为１．２ｍＬ／ｍｉｎ。系统的检测限：ＰＹＤ为１０ｎｍｏｌ／Ｌ，ＤＰＤ为７ｎｍｏｌ／Ｌ；回收率：ＰＹＤ为９１．５％，ＤＰＤ为１０６．１％；日内变异系数：ＰＹＤ为１．３９％，ＤＰＤ为０．１６％；日间变异系数：ＰＹＤ为３．７１％，ＤＰＤ为１．３２％。结果表明，方法具有简单、准确和特异的优点。

A method for the simultaneous and rapid determination of pyridinoline and deoxypyridinoline in urine using RP HPLC has been established. After hydrolysis for 18 hours at 107℃ in 6mol/L HCl, the urine sample was prefractionated by partition chromatography using a column packed with cellulose CF1. The appropriate fractions were freeze dried. The samples were analyzed by HPLC with a 5μm Spherisorb C18 column, a mobile phase of MeOH/H2O containing 01% heptafluorobutyric acid at a rate of 12mL/min and fluorescence detector monitoring at λex 290nm and λem 400nm. The limit of a detection for PYD was 10nmol/L and DPD 7nmol/L. The recovery of PYD was 915% and that of DPD was 1061%. Within day CV for the two compounds were 139% and 016% respectively and day to day CV 371% and 132% respectively.