A new method of quantifying the contents of riboflavin (RF) and pyridoxine (PY) in their mixed solution was introduced in this study. A mathematical model was established to calculate the actual concentration of PY (Z) based on the apparent concentrations of PY (Y) and RF (X), which were quantified directly when RF and PY were mixed together. First, a linear relationship was found between Y and Z with a high coefficient, which defines fluorescence quenching efficiency. Second, a curvilinear equation was established between the apparent concentration of X and the fluorescence quenching efficiency (k) of PY. The actual concentration of PY could be obtained by using the two equations. The established mathematical model was verified, and the relative error of the calculated PY value was below 2.5%. The upper limit of fluorescence spectrophotometry quantification was up to 20 μg/mL for both RF and PY. Compared with RP-HPLC, this method is convenient in terms of sample pretreatment, as well as saves organic solvents and time.

Xiaofei Li, Zhihua Wu, Shenguang Tong, Ping Tong, Xin Li, Anshu Yang and Hongbing Chen