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Spectrofluorometric Method for the Determination of Sulfhydryl and Disulfide Groups in Milk Proteins

Research and Development Division, National Dairy Products Corporation, Glenview, Illinois

ABSTRACT

A spectrofluorometric method has been applied to the determination of sulfhydryl (-SH) and disulfide (-SS-) groups of milk proteins. The method is based on the decrease (quenching) of the fluorescence of fluorescein mercuric acetate (FMA) by -SH, at neutral or basic pH's and by -SS-at highly alkaline pH's, which is proportional to the concentration of these groups. FMA produces fluorescence when subjected to excitation by light at 499 mµ, which can be measured at 520 mµ by the use of a spectrofluorometer.

The method has been tested on model systems consisting of mixtures of reduced (GSH) and oxidized (GSSG) glutathione as well as milk protein systems with and without added known amounts of GSH and GSSG. A high degree of accuracy in the recovery of known amounts of -SH and -SS- was attained from these systems, proving that the method is highly dependable.

¹ Present address: Department of Food Science, Ohio State University, Columbus, Ohio.