Determination of Vitamin a in Dried Human Blood Spots by High-Performance Capillary Electrophoresis with Laser-excited Fluorescence Detection


We have developed a high-performance capillary electrophoresis (HPCE) method to analyze the retinol (vitamin A) concentration as retinol-retinol binding protein (holo-RBP) from microvolumes of serum (5-10 µl) or one to two drops (~20 µl) of blood collected and air-dried on blood collection filter paper. A 0.64-cm diameter disk was cut from the dried whole blood specimens and the samples were dissolved in a pretreatment buffer and filtered. Filtrate was injected onto the HPCE column for analysis. The separation was carried out in a 60 cm x 50 µm I.D. fused-silica capillary and the running voltage was 20 kV. A He-Cd laser with a wavelength of 325 nm was used for excitation, and the fluorescence of the holo-RBP complex was monitored at 465 nm by a photodiode. A virtual linear relationship was obtained for the retinol concentrations between HPCE and HPLC for 28 serum samples, 19 dried venous blood samples and 9 capillary dried blood spot samples, indicating that valid measures of serum retinol can be obtained from one to two drops of capillary blood collected on filter paper. The absolute detection limit for retinol by HPCE is below 3 µg/l. The method is very useful for vitamin A level screening, especially for children and premature new-born babies.



Keywords and Phrases

Retinol; Blood Analysis; High Performance Liquid Chromatography; Human Tissue; Intermethod Comparison; Microanalysis; Priority Journal; Sampling; Zone Electrophoresis; Chromatography, High Pressure Liquid; Electrophoresis; Human; Lasers; Paper; Spectrometry, Fluorescence; Support, Non-U.S. Gov't; Support, U.S. Gov't, Non-P.H.S.; Vitamin A

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