Validation of UPLC-MS/MS Method for Determination of Urinary Lactulose/Mannitol
Validation of UPLC-MS/MS Method for Determination of Urinary Lactulose/Mannitol
复制标题
DOI：
10.3390/molecules23102705
复制
发表时间：
2018-10-01
期刊：
影响因子：
4.6
通讯作者：
Persichilli, Silvia
中科院分区：
文献类型：
--
作者：
Gervasoni, Jacopo;Primiano, Aniello;Persichilli, Silvia
尿乳果糖/甘露醇的测定是评价肠屏障功能最常用的试验之一。高效液相色谱（HPLC）分离与电喷雾电离串联质谱保证了高水平的选择性和重现性。在本文中，我们报告了以前公布的液相色谱串联质谱法的升级，引入更可靠的内标物和超高效液相色谱与乙烯桥接的混合酰胺柱。超高效液相色谱法提供了一个有效的色谱分离的两种糖在5分钟内，相比15分钟，使用以前的方法。甘露醇的定量限为10 μ g/mL，乳果糖的定量限为2.5 μ g/mL，甘露醇和乳果糖的定量限分别为1000 μ g/mL和1000 μ g/mL。运行内精密度和准确度范围分别为0.7 - 2.9%和97.2 - 101.2%。运行间精密度和准确度范围分别为1.9 - 4.7%和94.8 - 97.5%。乳果糖和甘露醇的回收率均高于90.2%，基质效应均低于15%。通过这种新方法，我们在准确性和再现性方面有了真实的改进，确保在更短的时间内获得结果。对先前方案的更改使此方法非常适合常规用途。
Determination of urinary lactulose/mannitol is one of the most used tests to evaluate intestinal barrier function. High-performance liquid chromatography (HPLC) separation with electrospray ionization tandem mass spectrometry guarantees high levels of selectivity and reproducibility. In this paper we report an upgrade of the previous published liquid chromatography tandem mass spectrometry method, introducing more reliable internal standards and ultra-performance liquid chromatography with ethylene bridged hybrid amide columns. The ultra-performance liquid chromatography provided an efficient chromatographic separation of the two sugars in 5 min, compared to 15 min using the previous method. The limit of quantification was 10 mu g/mL for mannitol and 2.5 mu g/mL for lactulose, and the assay was linear up to 1000 mu g/mL for mannitol and 1000 mu g/mL for lactulose. The within-run precision and accuracy ranged from 0.7 to 2.9% and 97.2 to 101.2%, respectively. The between-run precision and accuracy ranged from 1.9 to 4.7% and 94.8 to 97.5%, respectively. Recovery was higher than 90.2% for both lactulose and mannitol, and the matrix effect for both lactulose and mannitol was lower than 15%. With this new method we have a real improvement in terms of accuracy and reproducibility, ensuring results in shorter time. The changes to the previous protocol make this method excellent for routine purposes.