Shimada E, Matsumura G
J Biochem. 1977 Jan;81(1):79-91. doi: 10.1093/oxfordjournals.jbchem.a131453.
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.
透明质酸是从成年兔皮肤中制备的。将脱脂皮肤粉末悬浮于0.5M氯化钠中进行匀浆，然后用十六烷基吡啶氯化物从该0.5M氯化钠提取物中沉淀出总糖胺聚糖，接着依次用浓度递增的氯化钠以及最终的0.5N氢氧化钠重新溶解。0.5M氯化钠提取物中的主要酸性糖胺聚糖透明质酸，通过DEAE - 葡聚糖凝胶色谱法进行纯化和分级分离。其分子量范围为1×10⁴至7.2×10⁴。另外，无需机械粉碎和匀浆即可从成年兔皮肤中获得透明质酸。将脱脂皮肤块悬浮于水中并加热至100℃，然后提取物先用链霉蛋白酶消化，接着用脱氧核糖核酸酶[EC 3.1.4.5]处理。糖胺聚糖在Sephadex G - 75凝胶过滤中被排除。该组织的两种主要糖胺聚糖透明质酸和硫酸皮肤素，通过在Sepharose 4B上的凝胶色谱法分离。这种透明质酸的分子量范围为1.6×10⁵至1.3×10⁶。这两种方法制备透明质酸的产率相似。第一种方法中的机械处理可能使透明质酸发生了降解。还研究了影响组织提取物粘度的其他因素。
