Stability of nicotinamide adenine dinucleotide immobilized to cyanogen bromide activated agarose

The stability of NAD(H) immobilized to a crosslinked agarose support (Sepharose(R)-4B) was examined in buffer solutions at a pH of 7.0 and 8.5. Specifically, this study investigated particle attrition and ligand leakage rates from a cyanogen bromide activated agarose support. Particle attrition did not occur under the experimental conditions. Ligand leakage rates were found to be first order in immobilized ligand concentration with two labile populations of ligand. The two-population model is consistent with the cyanogen bromide coupling chemistry, which results in both an isourea and imidocarbonate ligand linkage. The rate of ligand leakage was found to occur over a time scale of days, with first order rate constants ranging from 0.007 to 0.15 d(-1), depending on solution pH. (c) 1997 John Wiley & Sons, Inc.