Proteoglycans represent a special class of glycoproteins that are heavily glycosylated. In nature proteoglycans have a wide variety of biological functions, such as maintenance of protein conformation and stability, immune and surface or intracellular recognition, inflammation, cell adhesion, pathogenicity, metastasis, and other cellular processes [1]. These functions may be dependent on either the protein part or the carbohydrate part, or on both. Aconitum coreanum (Levl.) Rapaics (Guanbaifu in Chinese) is one of the most documented centuries-old Chinese herbs in ancient Chinese medicinal literature. To date, A. coreanum has been used to treat various kinds of disorders such as cardialgia, facial distortion, epilepsia, migraine headache, vertigo, tetanus, infantile convulsion, and rheumatic arthralgia [2, 3]. Some active components from A. coreanum, such as alkaloids and flavonol glycosides, have been widely investigated [4]. However, no specific studies on proteoglycan isolated from A. coreanum have been carried out. In this study a more refined water-soluble proteoglycan was isolated and purified, and some of its properties and chemical compositions were determined. In this study, an AKTA Explorer 100 purification system, equipped with a P-900 series pump, Monitor UV-900, Monitor pH/C-900, Fraction collector Frac-950, Auto-sampler A-900, and various kinds of columns, was employed to purify A. coreanum proteoglycan. It can simplify the experiment and improve the purity of proteoglycan. At the same time, the operations of data collection were driven by UNICORN software, which guarantees quick, simple communications between systems and users and meets the stringent control and data handling procedures of modern laboratories [5]. The yield of the crude water-soluble polysaccharide extracted with hot water from A. coreanum was 4.62%. The main fraction (ACP-I) was purified with DEAE-cellulose anion-exchange and Sepharose CL-6B gel filtration chromatography to yield 45% of the crude polysaccharide. ACP-I appeared as a white powder, [ ]D 20 +148 (c 0.2, H2O). The ACP-I was identified as consisting mainly of polysaccharide and protein by the phenol-sulfuric acid method and the Bradford method. As shown in Table 1, the percentages of polysaccharide and protein were 84.3% and 12.4%, respectively. ACP-I contained 20.3% uronic acid evaluated by the m-hydroxydiphenyl colorimetric method. The average molecular weight of ACP-I was calculated as 24 kDa by HPGPC according to the calibration curve with standard dextrans and glucose. The HPGPC profile also demonstrated that ACP-I had a single and symmetrically sharp peak, revealing that ACP-I was a homogeneous proteoglycan. GC analysis showed ACP-I was composed of five kinds of monosaccharides, namely fucose, arabinose, xylose, glucose, and galacturonic acid with molar ratios of 0.8:0.8:1:3.9:1.6. The results indicated Gal was the predominant monosaccharide. In the UV spectrum of the ACP-I, the intense peak at 209 nm was the characteristic band of double bonds or triple bonds. However, the peak at 278.9 nm should be attributed to protein. The IR spectrum of ACP-I revealed a typical major broad stretching peak around 3440 cm–1 for the hydroxyl group, and a weak band at 2930 cm–1 showing the C–H stretching vibration. The strong absorption peak at around 1630 cm–1 and a weak one near 1430 cm–1 were indicative of the presence of carboxyl groups and carbonyl groups that indicated the characteristic IR absorption of uronic acids. The band at 849 cm–1 was ascribed to -pyranoses in the polysaccharide. These observations further confirmed that the ACP-I was composed of polysaccharide, protein, and uronic acids.
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Mar 22, 2016
Na Wu + 3
Open Access