The utilization of bathocuproinedisulfonic acid as a reagent for determining D-glucose and D-galactose levels in glycoconjugates |
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Authors: | Sovic David M Lester Leanna R Murray Elizabeth E Cohenford Menashi A |
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Affiliation: | Department of Integrated Science and Technology, Marshall University, One John Marshall Drive, Huntington, WV 25755, USA |
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Abstract: | A sensitive, rapid, and reliable method for measuring d-glucose and d-galactose levels in glycoconjugates has been developed. In this method, the NAD(P)H produced from the enzymatic oxidation of the monosaccharides is reacted with a CuSO4-bathocuproinedisulfonic acid reagent (Cu-BCS) to produce a color complex absorbing maximally at 486 nm. With galactose dehydrogenase and glucose dehydrogenase serving as the model enzymes, graphs of absorbance versus varying d-glucose or d-galactose concentrations yielded a linear plot from 2.5 to 250 nmol of sugar. Using this procedure, sugar released by acid hydrolysis from lactose, porcine submaxillary mucin and raffinose was quantified. When p-nitrophenyl-α-d-glucopyranoside and p-nitrophenyl-β-d-galactopyranoside were acid hydrolyzed and assayed with the Cu-BCS reagent, the amount of sugar released from each of the p-nitrophenyl compounds was found to be equal to the levels of p-nitrophenol in solution. This method is easy to use and with minor modifications can be employed for the quantification of d-glucose and d-galactose in other glycoconjugates. |
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Keywords: | Nicotinamide adenine dinucleotide Nicotinamide adenine dinucleotide phosphate Glucose dehydrogenase Galactose dehydrogenase Raffinose Lactose Porcine submaxillary mucin p-Nitrophenyl-α- smallcaps" >d-glucopyranoside p-Nitrophenyl-β- smallcaps" >d-galactopyranoside Bathocuproinedisulfonic acid |
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