The utilization of bathocuproinedisulfonic acid as a reagent for determining D-glucose and D-galactose levels in glycoconjugates.
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-alpha-D-glucopyranoside and p-nitrophenyl-beta-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.
Sovic
David M
DM
Department of Integrated Science and Technology, Marshall University, One John Marshall Drive, Huntington, WV 25755, USA.
Lester
Leanna R
LR
Murray
Elizabeth E
EE
Cohenford
Menashi A
MA
D000818:Animals; D003124:Colorimetry; D005690:Galactose / Q000032:analysis*; D005947:Glucose / Q000032:analysis*; D006001:Glycoconjugates / Q000737:chemistry*; D007202:Indicators and Reagents; D008722:Methods; D009249:NADP; D010618:Phenanthrolines / Q000737:chemistry*
D006001:Glycoconjugates; D007202:Indicators and Reagents; D010618:Phenanthrolines; D009249:NADP; C028559:bathocuproine sulfonate; D005947:Glucose; D005690:Galactose