Abstract: A method for the quantitation of nucleotides (cytidine 5’-monophosphate, uridine 5’-monophosphate, adenosine 5’-monophosphate, inosine 5’-monophosphate, and guanosine 5’-monophosphate) in infant formula was developed by sulfoalkyl betaine-based zwitterionic hydrophilic interaction liquid chromatography. The nucleotides in samples were extracted by two-dimensional solid-phase extraction with weak anion exchange and strong cation exchange, data acquisition was achieved using a UV detector. Chromatographic separation was performed by isocratic elution with a mobile phase consisting of 0.001% formic acid-acetonitrile and 10 mmol/L sodium dihydrogen phosphate solution at pH 5. Under the optimized conditions, good linearity was observed for these nucleotides, with a limit of quantification in the range of 0.10 to 0.15 mg/100 g. The method recovery was above 95% and the precision (expressed as relative standard deviation, RSD) was within 10%. This method was fast, sensitive and accurate and could be applied for the determination of nucleotides in infant formula samples.

Key words: nucleotides; sulfoalkylbetaine-based zwitterionic hydrophilic interaction liquid chromatography; two-dimensional solid-phase extraction; ultra-high performance liquid chromatography; infant formula