Assay of serum allantoin in humans by gas chromatography–mass spectrometry

• Published in: Clinica chimica acta 2002-04, Vol.318 (1), p.63-70
• Main Authors: Pavitt, Darrell V, de Fonseka, Sue, Al-Khalaf, Nisrin, Cam, Janice M, Reaveley, David A
• Format: Article
• Language: English
• Subjects: Aged; Allantoin; Allantoin - blood; Arteriosclerosis - blood; Atherosclerosis (general aspects, experimental research); Biological and medical sciences; Biomarkers; Blood and lymphatic vessels; Calibration; Cardiology. Vascular system; Female; Free radical; Free Radicals - metabolism; Gas Chromatography-Mass Spectrometry; Humans; Lipids - blood; Male; Mass spectrometry; Medical sciences; Middle Aged; Reference Values
• ISSN: 0009-8981; 1873-3492
• DOI: 10.1016/S0009-8981(01)00805-1

Background: The small amount of allantoin present in human serum results from free radical (FR) action on urate and may provide a stable marker of free radical activity in vivo. We describe a gas chromatography–mass spectrometry (GC–MS) assay for serum allantoin and report a reference range in healthy individuals. Methods: Fasting blood samples were obtained from 134 healthy middle-aged volunteers (56 men, mean age 55, range 45–72; 78 women, mean age 55, range 50–72) Allantoin was assayed using 15N 2 allantoin as an internal standard. After isolation from aqueous standards or serum by extraction onto an anion exchange column (AG-MP1), allantoin was derivatised with N-methyl- N-(tert-butyldimethylsilyl) trifluoroacetamide (MTBSTFA). Derivatives were injected onto an HP-1 column and analysed using a Mass Selective Detector with Single Ion Monitoring at 398 and 400 m/ z. Results: The distribution of serum allantoin concentrations in men and women was non-Gaussian and log transformation was used for the analysis of data. Women (10.8±1.7 μmol/l (mean±S.D.)) had significantly lower serum allantoin levels than men (13.4±1.6 μmol/l, p=0.015). Reference ranges (95% CI) for middle-aged healthy subjects were 7.4–46.8 μmol/l (men) and 3.7–31.2 μmol/l (women). Conclusion: Gas chromatography–mass spectrometry provides a reliable and accurate method for the determination of serum allantoin.