Validation of an LC-MSMS Method for Urinary Lactulose and Mannitol Quantification: Results in Patients with Irritable Bowel Syndrome

Abstract EN

Aim.

Lactulose/mannitol ratio is used to assess intestinal barrier function.

Aim of this work was to develop a robust and rapid method for the analysis of lactulose and mannitol in urine by liquid chromatography coupled to tandem mass spectrometry.

Lactulose/mannitol ratio has been measured in pediatric patients suffering from irritable bowel syndrome.

Methods.

Calibration curves and raffinose, used as internal standard, were prepared in water : acetonitrile 20 : 80.

Fifty μL of urine sample was added to 450 μL of internal standard solution.

The chromatographic separation was performed using a Luna NH2 column operating at a flow rate of 200 μL/min and eluted with a linear gradient from 20% to 80% water in acetonitrile.

Total run time is 9 minutes.

The mass spectrometry operates in electrospray negative mode.

Method was fully validated according to European Medicine Agency guidelines.

Results and Conclusions.

Linearity ranged from 10 to 1000 mg/L for mannitol and 2.5 to 1000 mg/L for lactulose.

Imprecision in intra- and interassay was lower than 15% for both analytes.

Accuracy was higher than 85%.

Lactulose/mannitol ratio in pediatric patients is significantly higher than that measured in controls.

The presented method, rapid and sensitive, is suitable in a clinical laboratory.