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Clinical Chemistry 28: 29-31, 1982;
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Clinical Chemistry, Vol 28, 29-31, Copyright © 1982 by American Association for Clinical Chemistry

Liquid-chromatographic determination of the total thiamin content of blood

M Kimura, T Fujita and Y Itokawa

A liquid-chromatographic method for determining the total thiamin content of blood is presented. Blood is deproteinized and incubated with Aspergillus oryzae carboxyl proteinase (EC 3.4.23.6; Takadiastase) to convert thiamin phosphate esters to free thiamin. An aliquot of the sample is applied to the column (Shodex OH-Pak M-414) of a high- performance liquid chromatograph. A 100 mg/L solution of potassium ferricyanide in 150 g/L sodium hydroxide is added to the column effluent with a proportioning pump, to convert thiamin into a fluorophore. The intensity of the fluorophore is measured with a spectrofluorophotometer and recorded graphically. The total thiamin in each blood sample appears as a single peak, and no co-eluting substance was detected. This method is simple, highly reproducible, and rapid, and its sensitivity is sufficient for determination of the thiamin content of 0.1-mL blood samples.


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Copyright © 1982 by the American Association for Clinical Chemistry.