Factors affecting a cyanogen bromide-based assay of thiamin.
We analyzed extensively a modified thiochrome method for thiamin analysis. Acid phosphatase (EC 3.1.3.2) from potato was superior to either alpha-amylase or acid phosphatase from wheat germ as a dephosphorylating agent. Timing of cyanogen bromide exposure was important, but the assay had good precision and accuracy. The standard curve was linear from 10 to 3000 nmol/L. The within-run and between-run coefficients of variation for total thiamin in whole blood were 3.6% and 7.4%, respectively. Analytical recoveries for low, intermediate, and high additions of thiamin to whole blood were 93-109%. Sample yield was increased by 41% (+/- 29% SD) with pre-assay freezing. Samples were stable for two days at room temperature, for seven days when refrigerated, and for two years when frozen. Previously unreported interference was seen with penicillin derivatives, and with several commonly used diuretic and antiepileptic medications. This assay may be suitable for population screening; 200 samples could be analyzed weekly at a cost of +0.20 per sample.
Wyatt
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DT
Department of Pediatrics, Medical College of Wisconsin, Milwaukee 53226.
Lee
M
M
Hillman
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RE
D000135:Acid Phosphatase; D000328:Adult; D000927:Anticonvulsants / Q000097:blood; D003488:Cyanogen Bromide; D004232:Diuretics / Q000097:blood; D006801:Humans; D010084:Oxidation-Reduction; D010406:Penicillins / Q000097:blood; D010766:Phosphorylation; D011786:Quality Control; D012016:Reference Values; D013050:Spectrometry, Fluorescence; D013831:Thiamine / Q000097:blood*; D014238:Trichloroacetic Acid
D000927:Anticonvulsants; D004232:Diuretics; D010406:Penicillins; D014238:Trichloroacetic Acid; D000135:Acid Phosphatase; D003488:Cyanogen Bromide; D013831:Thiamine