Optimisation of the determination of thiamin, 2-(1-hydroxyethyl)thiamin, and riboflavin in food samples by use of HPLC
2008
Jakobsen, J.
The aim of this study was first to optimise and validate a method using an enzyme-mixture to liberate protein- and phosphate-bound thiamin and riboflavin in food by the use of ultrasonication and HPLC, and second to include the quantitation of the vitamin B₁ active compound 2-(1-hydroxyethyl)thiamin (HET). The enzyme-mixture consisted of α-amylase, proteinase, and phosphatase. The use of ultrasonication in the enzyme treatment enabled the results for vitamin B₁ and B₂ to be obtained in 1 day. In consequence of an incomplete release of phosphate-bound thiamin of some of the batches of enzymes used, thiamin was quantitated as the sum of thiaminmonophosphate and thiamin. The vitamin B₁ active compound, HET was detected and quantitated separately. The standard deviations for the method were 3.7%, 4.7%, and 13.3% for thiamin, riboflavin, and HET, respectively. The relative bioactivity of HET is similar to the bioactivity of thiamin. In the samples of animal origin the content of HET represented 7-24% of the content of thiamin, while in dried yeast the content of HET was 37% of the content of thiamin. Quantitation of vitamin B₁ in food by a post-column derivatisation is recommended to include separate quantitation of thiamin and HET.
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