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During the last few years, knowledge has developed concerning the biosynthesis of the he teropol ysaccharides, A biosyn the tic pathway for hyaluronic acid has been established (1); from glucose this involves the formation of uridinediphospho-D-glucuronic acid and uridinediphospho -Nacetyl-Dglucosamine, which are precursors of the heteropoly-It has been shown that D-glucosamine-l-C14 injected into rats and rabbits(3,4) is incorporated into the glycoproteins of the body tissues and fluids. In this investigation, we have studied the incorporation of D-glucosamine-1 -C14 into the acid mucopolysaccharides of rabbit liver.Experimental. Fasting young white male rabbits weighing 1504 (1342-1666) g were injected intraperitoneally with 60 microcuries of D-glucosamine-l-C14. At intervals of 0.5 to 72 hours following the injection, the animals were sacrificed by exsanguination from the abdominal aorta. The liver was removed, perfused with ice cold saline, lyophilized and extracted with n-butanol and acetone. The acid mucopolysaccharides were extracted from 2 g of this preparation by a modification of the method described by Schiller et aZ. (5). This involved extraction with 0.5 N NaOH, neutralization, ethanolic precipitation, digestion with proteolytic enzymes, trichloroacetic acid precipitation of protein, and dialysis followed by passage of the acid-free preparation through Dowex 50 resin at 0'. Mucopolysaccharides were precipitated with cetylpyridinium chloride and fractionated by selective solubilization of the cetylpyridinium salts of the mucopolysaccharides (Scott, 6). By this technique the cetylpyridinium salts were resolved into three fractions: those soluble in 0.5, 1.5 and 2.0 molar Supported in part by a predoctoral fellowship from Nat. Inst. Health. Heparitin sulfate was kindly supplied by Dr. J. A. Cifonelli and kcratosulfate by sodium chloride. Cetylpyridinium chloride was removed by precipitation with KCNS ( 7 ) ; soluble salts were then removed by dialysis.The resulting polysaccharide preparations were analyzed for uronic acid by a modification of the carbazole method(8), and for hexosamine (9).For further studies of the hexosamine constituents, samples of the polysaccharide preparations were hydrolyzed in 4N HCl in closed tubes at 100°C for 6 hours. After removal of HCl in vacuo at room temperature, the hydrolysates were dissolved in water, again taken to dryness, and finally dissolved in 10% isopropanol. Aliquots were subjected to descending paper chromatography on Whatman No. 1 paper using an ethyl acetatepyridine-water ( 12 : 5 : 4) solvent system which separates glucosamine and galactosamine. Amino sugars were detected by dipping the chromatogram in a 0.2% ninhydrin solution (in acetone with 4% acetic acid), followed by development at room temperature. Spots with the mobility of glucosamine were found in all hydrolysates; however, galactosamine was never detected. Radioisotope studies of the glucosamine areas of the paper chromatograms were made by cutting out the appropriate areas and placing them dire...