Determination of Pyridoxine in Dietary Supplements by Liquid Chromatography with UV, Fluorescence, and Mass Spectrometric Detection: Single-Laboratory Validation

Abstract

A single-laboratory validation was performed for a method that determines pyridoxine, one of the B6 vitamers, in dietary supplements using LC and UV, fluorescence, or MS detection. The method was adapted for use with either HPLC or ultra-performance LC (UPLC). Pyridoxine is extracted from samples using 0.1 M formic acid, and specific conditions are adjusted for each of the different types of supplement materials examined. Reversed-phase chromatography with C18-based columns is used in both HPLC and UPLC. Fluorescence detection, often used in chromatographic analyses of vitamin B6 in foods, was successfully used here, but offered no great advantages over UV detection in the supplement materials tested. MS detection was also satisfactory, although use of an internal standard was required. Accuracy of the method was demonstrated in several ways, including use of a standard reference material. Precision and repeatability of the method were found acceptable by analysis of variance and HorRat repeatability calculations.