Plasma allantoin measurement by isocratic liquid chromatography with tandem mass spectrometry: Method evaluation and application in oxidative stress biomonitoring

摘要

Background: Allantoin in human plasma is a specific biomarker of oxidative stress. We describe a sensitive method to measure plasma allantoin using isocratic liquid chromatography and mass spectrometry (LC-MS/MS). Methods: Direct injection of deproteinized plasma into the LC-MS/MS system was performed. The method was technically evaluated. Results on 200 healthy and 35 Type 2 diabetic Chinese subjects were compared. Results: Dose-response of allantoin was linear to at least 21 pmol (20 umol/1 in plasma); LOD was 0.16 pmol; recovery 99.2-100.2% at 1-5 umol/1; accuracy, 98.5-100.8%; within-day and between-day CVs (n = 6), <4.0% (at 5.00-40 umol/1) and <2.0% (at 1-5 umol/1), respectively. Plasma allantoin in diabetic patients was ~8-fold higher than in healthy subjects; mean (SD): 8.82 (7.26) and 1.08 (0.86) umol/1, respectively (p < 0.0001). Allantoin was slightly higher in healthy men than in age- and BMI-matched women: 1.21 (0.99) umol/1, n = 88 compared to 0.97 (0.74) umol/1, n = 112; p < 0.001. No association with age was seen. Gender difference was also seen in the diabetes patients: men, n = 14,11.57 (8.57) umol/1; women, n = 21,6.99 (5.75) umol/1, p < 0.05. Conclusions: Based on 95th percentiles of the healthy subjects, plasma allantoin of > 2.2 umol/1 in women and >3.1 umol/1 in men indicates increased oxidative stress. Allantoin in diabetes subjects is clearly and markedly increased. The method will facilitate future studies of oxidative stress in human biomonitoring studies.