Colorimetric assay for free and bound L-fucose.

M A Cohenford, A Abraham, J Abraham, J A Dain

Index: Anal. Biochem. 177(1) , 172-7, (1989)

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Abstract

A novel, rapid, and reliable colorimetric method for measuring L-fucose has been developed. This method utilizes NADH formed from the interaction of L-fucose with fucose dehydrogenase and NAD to generate color in a reaction involving CuSO4 and neocuproine. NADH reduces Cu2+ to Cu1+ and the latter interacts with neocuproine to yield a complex with a maximal absorption at 455 nm. The reaction of NADH with copper-neocuproine is immediate and under the conditions of the assay the color formed remains stable for at least 2 h. When the assay is used to determine levels of L-fucose, the absorbance is found to be linearly proportional to exogenously added fucose concentrations from 16 to 179 nmol with resulting molar extinction coefficient of 13,660. Using this procedure, L-fucose released by acid hydrolysis from porcine submaxillary mucin, and by alpha-L-fucosidase from p-nitrophenyl-alpha-L-fucopyranoside, was quantitated.