The analysis of 5′-mononucleo tides in infant formulae by HPLC

A method is described for the determination of four 5'-mononucleotides (cytidine 5-monophosphate, uridine 5'-monophosphate, adenosine 5-monophosphate and guanosine 5'-monophosphate) in infant formulae. Nucleosides which may be formed during processing can also be analysed simultaneously. This method is based on deproteinisation of samples and direct analysis by ion-pair HPLC using two Nucleosil 120-C-18 columns in series, followed by diode-array detection. This method gives good recoveries of 5'-mononucleotides from spiked infant formula products. However, some chromatographic interferences were observed when analysing hypoallergenic infant formulae containing hydrolysed proteins which made peak quantification difficult. To overcome this problem a strong anion-exchange solid-phase extraction (SPE) column was used. Four SPE columns from different suppliers were evaluated, but the best recoveries of all four 5'-mononucleotides and highest reproducibility of results were obtained with Bakerbond(R) quaternary amine columns. Nucleosides, which may occur in very low concentrations in hypoallergenic products, are not retained on the SPE columns and so cannot be analysed by this technique. (C) 2001 Elsevier Science Ltd. All rights reserved.