Liquid-chromatographic determination of the total thiamin content of blood.
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.
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods; D006801:Humans; D008722:Methods; D011817:Rabbits; D051381:Rats; D012016:Reference Values; D013831:Thiamine / Q000097:blood*
D013831:Thiamine