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Nondestructive Measurement of Carotenoids in Plant Tissues by Fluorescence Quenching

USDA-ARS, Dale Bumpers National Rice Research Center, Stuttgart, AR 72160-1090

View larger version (17K): [in a new window]   Fig. 1. Illustration of the mathematical integrations of the fluorescence spectra for milled rice (having no carotene) and milled rice spotted with ß-carotene. The addition of ß-carotene (B) causes a shift downward (arrow) of the spectral envelope at the lower wavelengths since carotenoids, in general, absorb light of wavelengths less than about 525 nm. In the presence of carotenoids, the area above the baseline curve (dark shading in A and B) becomes greater. Changes in the curves of the spectral envelope are measured in comparison to the straight line of the corresponding baseline spectrum by subtracting the area under the baseline from the area under the spectral evelope, thus producing the net integrated fluorescence intensity.

View larger version (14K): [in a new window]   Fig. 2. Demonstration of the changes in autofluorescence that occur during fluorescence quenching. The natural autofluorescence of milled white rice is the white rice line. The scan of white rice spotted with carotene was subtracted from the scan of white rice. The resulting subtraction line is the absorption spectrum of the ß-carotene in the dry endosperm matrix. The net integrated fluorescence intensity is obtained from the 505- to 565-nm region of a fluorescence spectrum.

View larger version (21K): [in a new window]   Fig. 3. Dose response curves of net integrated fluorescence intensities from fluorescence quenching measurements of known carotenoids. Measurements were made of white rice kernels spotted with 1 µL of five different concentrations of three carotenoids. Bars indicate standard deviations.