Partial Purification of Rat Urinary Anorexigenic Substance

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Abstract
Partial purification of an anorexigenic component (FMS IA) of fatmobilizing substance extracted from rat urine was accomplished through ultrafiltration, anion exhcange chromatography on DEAE-cellulose and gel filtration on Sephadex G-75. Bioassay consisted of measuring body weight and spontaneous intake of a standardized diet before and after i.p. injection of various materials into male assay rats weighing approximately 200 g. Losses of body weight paralleled reductions of food intake in all cases. Specific anorexigenic activity was defined as percentage reduction in food intake 24 hours after injection/milligram injected. A fraction of FMS IA of approximate molecular weight (m.w.) 30,000 to 50,000, separated by ultrafiltration, showed specific activity 8.2, a nearly threefold improvement over that of the starting material, dialyzed FMS IA, with specific anorexigenic activity 2.8. Anorexic responses to this fraction were linear in the dosage range 0 to 12 mg. A fraction isolated by chromatography on DEAE-cellulose showed specific anorexigenic activity 5.5, a twofold improvement over the starting material. The m.w. of this fraction was not determined. A more potent fraction was obtained by the use of Sephadex G-75. This fraction, of m.w. apparently above 80,000, showed specific anorexigenic activity 27.0 when injected in a dose of 1.5 mg. This represents a nearly tenfold increase in biological activity. There was no apparent relationship between anorexic response and dose injected over the range 0.9 to 3.6 mg. The m.w. of the active material in the above preparations appears to be either about 40,000 or 80,000, or higher, depending upon the method of investigation employed and possible aggregation or disaggregation of subunits. These results demonstrate again the presence of a powerful anorexigenic substance in the urine of rats.

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