Optimization and validation of an LC-fLD method for biotin in infant formula, infant cereals, cocoa-malt beverages, and clinical nutrition products

A fast and simple chromatographic method to determine biotin in foods is presented. Biotin is extracted using papain (60 degrees C, 1 h). After pH adjustment and filtration, biotin is determined by LC with fluorescence detection using postcolumn reagent avidin-FITC (avidin labeled with fluorescein isothiocyanate). The method has been validated in a large range of products: milk- and soy-based infant formulas, cereals, cocoa-malt beverages, and clinical nutrition products. The method showed recovery rates of 98.1 +/- 5.7% (average +/- SD) in a large range of concentrations. Biotin concentrations determined in infant formula standard reference materials 1846 and 1849 were in agreement with reference values. RSD of repeatability (RSDr) varied from 2.0 to 4.5%, and intermediate reproducibility (RSD(iR)) from 5.8 to 9.4%. LOD and LOQ were 3.0 and 5.0 microg/100 g, respectively. The proposed method is suitable for routine analysis of biotin in fortified foods (infant formulas, infant cereals, cocoa-malt beverages, and clinical nutrition products). It can be used as a faster, more selective, and precise alternative to the classical microbiological determination, and is easily transferable among laboratories.