HILIC-MS/MS Method for the Quantitation of Nucleotides in Infant Formula and Adult Nutritional Formula: First Action 2011.21

Official Method(SM) 2011.21 is for the quantitation of the following nucleotides: adenosine 5'-monophosphate (AMP), guanosine 5'-monophosphate (GMP), uridine 5'-monophosphate (UMP), cytidine 5'-monophosphate (CMP), and inosine 5'-monophosphate (IMP) in infant formula and adult/pediatric nutritional formula. It uses hydrophilic interaction liquid chromatography with tandem mass spectrometry (HILIC-MS/MS). Preparation of the internal standards was conducted using centrifugal ultrafiltration and the standards are AMP-C-13(10), N-15(5); GMP-C-13(10), N-15(5); UMP-C-13(9), N-15(2); and CMP-C-13(9), N-15(3). Data were collected by using multiple reaction monitoring of the product ions of protonated molecules of the five nucleotides generated by positive-electrospray ionization. The HILIC conditions were conducted with ammonium formate (30 mmol/L) in water (pH 2.5, adjusted with formic acid) and methanol. The LOD and LOQ of the standard solution were 0.005-0.01 and 0.01-0.03 mu g/mL, respectively. Recovery data were collected for intraday and interday testing and ranged from 98.1 to 108.9% with an RSD of 0.7-5.4%. The analytical range of the method is between 0.04 to 5 mu g/mL for standard solution.