Hydrophilic interaction chromatography/electrospray ionization mass spectrometry for determination of allantoin in human biological samples

Allantoin is a catabolic product of purines, which is formed by the enzymatic reaction of uricase with uric acid. Investigations of the residual existence of allantoin in the human body have revealed the presence of trace levels of allantoin in human biological samples. However, the hitherto available analytical techniques for allantoin in biological samples suffer from such problems as insufficient separation from coexisting substances and a non-specific detection wavelength of 220 nm. Therefore, an accurate, sensitive and selective analytical method for the determination of allantoin in human biological samples using hydrophilic interaction chromatography with mass spectrometry (HILIC/MS) was developed. The following were used for HILIC/MS: Atlantis(TM) HILIC Silica column, electrospray ionization (ESI), negative mode, and selected ion monitoring (SIM) with m/z 157 for allantoin and m/z 159 for allantoin-1,3-(15) N-2. The detection limit (S/N = 3) and the quantification limit (S/N = 10) were 0.03 and 0.1 muM, respectively, and the calibration curve (0.1 similar to 10 muM) showed linearity with a correlation coefficient (r) of 0.999. The average recoveries of allantoin with an internal standard in human serum, urine and saliva samples were 96.9% (RSD: 13.2%), 95.2% (RSD: 9.0%) and 100.3% (RSD: 11.8%), respectively. The proposed HILIC/MS method for the determination of allantoin can be applied to the detection of trace amounts of allantoin in human biological samples.