Determination of retinol, alpha-tocopherol, and beta-carotene in serum by liquid chromatography with absorbance and electrochemical detection.
We describe a method for the determination of retinol, alpha-tocopherol, and beta-carotene in serum, using a liquid-chromatographic separation with wavelength-programmed ultraviolet/visible absorbance and amperometric electrochemical detection with a glassy carbon electrode. After protein denaturation and addition of an internal standard, tocol, 250-microL samples are twice extracted with hexane. The reversed-phase, gradient-elution chromatographic separation provides baseline resolution of: the all-trans isomer of retinol from the cis isomers, alpha- from gamma-tocopherol, and all-trans-beta-carotene from alpha-carotene and from cis-beta-carotene isomers. The linearity of response and the detection limits for the two detectors for the three analytes are measured. A comparison of the values obtained for serum extracts shows good agreement between the absorbance and electrochemical detectors.
D002338:Carotenoids / Q000097:blood* / Q000302:isolation & purification; D002853:Chromatography, Liquid / Q000379:methods; D004563:Electrochemistry; D006801:Humans; D007202:Indicators and Reagents; D007536:Isomerism; D013053:Spectrophotometry; D014801:Vitamin A / Q000097:blood* / Q000302:isolation & purification; D014810:Vitamin E / Q000097:blood* / Q000302:isolation & purification; D019207:beta Carotene
D007202:Indicators and Reagents; D019207:beta Carotene; D014801:Vitamin A; D014810:Vitamin E; D002338:Carotenoids