The uncertainty in the determination of nicotinamide in milk powder by ultra-high performance liquid chromatography was evaluated. According to the technical normal “Evaluation and Expression of Uncertainty in Measurement” (JJF 1059.1-2012), a mathematical model was established, and the source of each uncertainty component in the mathematical model was analyzed and calculated. The results showed that the uncertainty in nicotinamide determination mainly came from the dilution of the standard solution. In addition, the average content of nicotinamide in milk powder was 5007 μg/100 g with an expanded uncertainty of about 197 μg/100 g (k = 2).

To establish an ultra performance liquid chromatography (UPLC) analysis method that can simultaneously determine niacin, niacinamide, and vitamin B₆ in formula milk powder. Gradient elution was performed with 10 mmol/L ammonium acetate solution (containing 0.1% formic acid) as mobile phase A and methanol as mobile phase B. The analytes were separated by Poroshell 120 EC-C₁₈ in tandem with ACQUITY^® UPLC HSS T3 column, detected with the UV detector and fluorescence detector, and quantified by an external standard method. Good linearity was observed for niacin concentrations in the range of 0.0-0.8 μg/mL,nicotinamide concentrations in the range of 0.0-20.0 μg/mL, pyridoxamine concentrations in the range of 0.0-0.2 μg/mL, pyridoxal concentrations in the range of 0.0-0.6 μg/mL and pyridoxine concentrations in the range of 0.0-4.0 μg/mL, and the correlation coefficients of standard curves were all greater than 0.999. The limits of detection and quantification met the requirements. The average recoveries of the method were all in the range of 80.5%-104.1% with relative standard deviation (RSD) were 0.37%-2.99%. This method is simple, accurate, can be used for batch detection of niacin, niacinamide, and vitamin B₆ in formula milk powder.