Nucleotides play important roles in numerous intracellular biochemical processes
and are used in infant formulas and other dairy products. However, domestic
analytical methods for assessing nucleotide content in products have not yet
been established, and therefore, methods for determining nucleotide content are
urgently required. A rapid and simple analytical method for determining the
content of five types of nucleotides in dairy products was improved using solid
phase extraction clean-up and high-performance liquid chromatography with diode
array detector. The extraction solvent used in the AOAC method was not well
dissolved and was changed to hydrophilic EDTA-Na. In addition, the results
obtained using the isocratic elution method and a single wavelength were similar
to those obtained using the AOAC method, and the time taken for analysis was
shortened from 40 min to 25 min. The process of method validation revealed the
following parameters: accuracy (84.69%–102.72%), precision
(1.51%–6.82%), linearity (0.999), and limit of detection
(cytidine 5′-monophosphate, 0.09 mg/L; uridine 5′-monophosphate,
0.11 mg/L; adenosine 5′-monophosphate, 0.12 mg/L; guanosine
5′-monophosphate, 0.11 mg/L; and inosine 5′-monophosphate, 0.14
mg/L). The method was also used to determine the nucleotide concentration in 25
samples (infant formulas, 1.99–29.39 mg/100 g; and cow milk,
0.28–0.83 mg/100 g). The newly improved method was appropriate for
analyzing nucleotides in infant formulas and other dairy products faster when
compared to conventional methods.