Measurement of citrate in urine using liquid chromatography tandem mass spectrometry: comparison with an enzymatic method
Measurement of citrate in urine using liquid chromatography tandem mass spectrometry: comparison with an enzymatic method
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DOI：
10.1258/0004563054889963
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发表时间：
2005-09-01
影响因子：
2.2
通讯作者：
Kavanagh, J
中科院分区：
文献类型：
--
作者：
Keevil, BG;Owen, L;Kavanagh, J
背景：尿液柠檬酸盐的测量用于评估进一步形成尿路结石的风险，并评估受影响个体的治疗益处。我们希望开发一种简单快速的液相色谱串联质谱 (LC-MS/MS) 方法来分析尿柠檬酸盐，并将其与我们当前的酶法测定进行比较。方法：对于 LC-MS/MS 测定，通过将 10 μL 尿液和 20 μL 内标添加到 400 μL 水中，在深孔块中制备样品。混合后，将3μL稀释样品注入LC-MS/MS系统。使用 LC 系统，用含有 2 mmol/L 乙酸铵和 0.1% (v/v) 甲酸的 0.4 mL/min 水等度洗脱 C18 柱 (50 x 2.1 mm)。使用含有 2 mmol/L 乙酸铵和 0.1% (v/v) 甲酸的 100% 甲醇逐步梯度洗柱。柠檬酸盐的保留时间为 1.4 分钟，d4-柠檬酸盐的保留时间为 1.4 分钟。注射到注射的循环时间为 4.0 分钟。使用串联质谱仪在多反应监测模式下使用以下离子对进行监测，柠檬酸盐 m/z 191.0 > 111.0 和 d4-柠檬酸盐 m/z 195.0 > 113.0。 结果：在 480-3800/ mu mol/L 范围内，批次内和批次间的变异系数 < 3%。定量下限为24.0μmol/L。回归分析显示 LC-MS/MS = 0.8781（酶测定）+ 102.5，r = 0.964，n = 73。 结论：我们开发了一种简单的 LC-MS/MS 方法用于尿液柠檬酸盐测量，该方法显示出可接受的性能。
Background: Measurement of urine citrate is used to assess the risk of further urinary stone formation and to assess the benefit of treatment in affected individuals. We wanted to develop a simple and rapid liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the analysis of urinary citrate and to compare it with our current enzymatic assay.Methods: For the LC-MS/MS assay, samples were prepared in a deep-well block by adding 10 mu L of urine and 20 mu L of internal standard to 400 mu L of water. After mixing, 3 mu L of the diluted sample was injected into the LC-MS/MS system. An LC system was used to isocratically elute a C18 column (50 x 2.1 mm) with 0.4 mL/min water containing 2 mmol/L ammonium acetate and 0.1% (v/v) formic acid. A step gradient of 100% methanol containing 2 mmol/L ammonium acetate and 0.1% (v/v) formic acid was used to wash the column. The retention times were 1.4 min for citrate and 1.4 min for d4-citrate. Cycle time was 4.0 min, injection to injection. The analytes were monitored using a tandem mass spectrometer operated in multiple reaction monitoring mode using the following transitions, citrate m/z 191.0 > 111.0 and d4-citrate m/z 195.0 > 113.0.Results: Within and between-batch coefficients of variation were < 3% over the range 480-3800/ mu mol/L. The lower limit of quantification was 24.0 mu mol/L. Regression analysis showed LC-MS/MS = 0.8781 (enzymatic assay) + 102.5, r = 0.964, n = 73.Conclusions: We have developed a simple LC-MS/MS method for urinary citrate measurement that shows acceptable performance.