Characterization of Food Binding of Vitamin B-6 in Orange Juice

There is substantial evidence that natural vitamin B-6 exists largely in a bound form. It has been shown that the principal modes of binding represents an interaction of pyridoxal and/or pyridoxal phosphate with protein moieties. The nature and extent of binding of different forms of the vitamin in orange juice was investigated. Studies employing ultracentrifugation, ultrafiltration and, membrane dialysis of this food product suggested binding to a molecule of less than 3,500 daltons molecular weight. Binding was resistant to protease treatment although susceptible to heat-acid hydrolysis. Separation of pyridoxal, pyridoxine, and pyridoxamine species of the vitamin by ion exchange chromatography was followed by fluorometric analysis with and without preparative hydrolysis. Extensive and equal binding of both pyridoxal and pyridoxine was observed by this method. Thus, the binder(s) of vitamin B-6 in orange juice is a small dialyzable molecule which is heat stable, binds both pyridoxal and pyridoxine and is non-protein in nature.