Molecular weight of hyaluronic acid from rabbit skin

Hyaluronic acid was prepared from adult rabbit skin. Defatted skin powder suspended in 0.5 M NaCl was homogenized, and total glycosaminoglycans were precipitated from this 0.5 M NaCl extract with cetylpyridinium chloride, then redissolved successively with increasing concentrations of NaCl and finally 0.5 N NaOH. Hyaluronic acid, the major acid glycosaminoglycan in the 0.5 M NaCl extract, was purified and fractionated by DEAE-Sephadex chromatography. The molecular weights ranged from 1 X 10(4) to 7.2 X 10(4). Alternatively, hyaluronic acid was obtained from adult rabbit skin without mechanical powdering and homogenizing. Defatted skin pieces were suspended in water and heated at 100 degrees, then the extract was digested with pronase followed by DNase [EC 3.1.4.5]. Glycosaminoglycans were excluded in gel filtration with Sephadex G-75. Hyaluronic acid and dermatan sulfate, the two major glycosaminoglycans of this tissue, were separated by gel chromatography on Sepharose 4B. The molecular weight of this hyaluronic acid ranged from 1.6 X 10(5) to 1.3 X 10(6). Yields of hyaluronic acid by these two methods were similar. Hyaluronic acid was probably degraded by the mechanical treatments in the first method. Other factors affecting the viscosity of the tissue extract were examined.