Abstract
ObjectivesVitamin B12 dietary supplement can be critical to the alleviation strategies against micronutrient malnutrition and food insecurity. A high-performance liquid chromatography with ultraviolet detection (HPLC-UV) method has been developed and validated, for the quantitation of four bioactive forms of vitamin B12 (adenosylcobalamin, cyanocobalamin, hydroxocobalamin, methylcobalamin) from dietary ingredients and supplements. MethodsA Plackett-Burman factorial study was used to identify factors that contributed to the extraction of cobalamins. Significant factors were selected to produce the improved HPLC method for cobalamin separation. This method was then subjected to a single-laboratory validation according to the AOAC International guidelines for linearity, suitability, detection limits, accuracy, and precision. ResultsThe method achieves chromatographic baseline resolution of vitamin B12 forms on a modern column platform without the expensive requirement of an ultra-high pressure liquid chromatography and/or mass spectrometry. The method has a wide analytical range (0.0005% w/w - 85% w/w), high precision (repeatability relative standard deviations ranged from 1.08% to 3.06%), and high accuracy (>96% spike recovery rate). The method detection and quantification limits are less than 0.16 and 0.52 μg/mL, respectively. ConclusionsTo our best knowledge, the method is simpler, less time-consuming, and more economical than other published methods for its intended uses. Funding Sources1Eurofins Supplement Analysis Center, Eurofins Scientific, Inc.2MilliporeSigma Supporting Tables, Images and/or Graphs▪
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