Volpi N
Department of Biologia Animale, University of Modena, Italy.
Boll Chim Farm. 1993 May;132(5):153-60.
Heparin was extracted and purified from beef intestinal mucosa. The two components, fast moving heparin and slow moving heparin were purified by selective precipitation as barium salts. Heparan sulfate was extracted and purified from beef spleen. Dermatan sulfate was purified from beef intestinal mucosa and chondroitin sulfate from bovine trachea. The purity of the purified glycosaminoglycans was evaluated by agarose-gel and cellulose polyacetate electrophoresis and by specific optical rotation. The relative molecular masses of glycosaminoglycans were estimated by high performance-size exclusion chromatography and the sulfate to carboxyl ratio by titrimetric analysis. The disaccharide pattern of heparin, fast moving and slow moving heparins and heparan sulfate were determined by specific enzymatic cleavage using heparinase I, II and III; the disaccharide composition of dermatan sulfate and chondroitin sulfate was evaluated by cleavage by chondroitinase ABC. The disaccharides obtained by enzymatic cleavage were qualitatively and quantitatively analysed by strong anion exchange-high performance liquid chromatography. The sulfate to carboxyl ratios of glycosaminoglycans were also determined by this technique and compared with the values obtained by titrimetric analysis.
肝素从牛肠黏膜中提取和纯化。快速移动的肝素和慢速移动的肝素这两种组分通过选择性沉淀作为钡盐进行纯化。硫酸乙酰肝素从牛脾脏中提取和纯化。硫酸皮肤素从牛肠黏膜中纯化，硫酸软骨素从牛气管中纯化。通过琼脂糖凝胶和醋酸纤维素电泳以及比旋光度评估纯化的糖胺聚糖的纯度。通过高效尺寸排阻色谱法估计糖胺聚糖的相对分子质量，并通过滴定分析确定硫酸根与羧基的比例。使用肝素酶I、II和III通过特异性酶切确定肝素、快速移动和慢速移动肝素以及硫酸乙酰肝素的二糖模式；通过软骨素酶ABC切割评估硫酸皮肤素和硫酸软骨素的二糖组成。通过强阴离子交换-高效液相色谱法对酶切获得的二糖进行定性和定量分析。还通过该技术测定糖胺聚糖的硫酸根与羧基比例，并与滴定分析获得的值进行比较。
