葡萄多糖的提取纯化及物性、抗氧化性研究
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摘要
葡萄(Vitis vinifera L.)系葡萄科葡萄属植物,既是著名的水果和酿酒原料,也作为药材被药典收载。本研究以新疆鲜无核白葡萄为原料,对葡萄粗多糖进行了提取及初步的纯化研究,并对其物性进行了评价,为今后葡萄多糖更深入的研究与开发奠定了理论基础。研究结果如下:
1.比较了Sevag法、酶法、酶法与Sevag法结合脱蛋白的效果,结果表明酶法与Sevag法结合脱蛋白效果最佳,其最佳处理条件为酶用量0.l4%,pH6.5,于60℃下保温2h,再经Sevag脱蛋白4次去除残留蛋白质,去蛋白效果最理想此时蛋白脱除率为72.14%,多糖损失率为l0.91%。
2.采用HPD-300型树脂,30℃下在摇床中吸附3h,树脂用量24g/100ml时,对葡萄多糖的脱色效果最好,该树脂再生后脱色效果良好,可重复利用。
3.透析去除小分子杂质和无机离子后得到分子量大于8000D的葡萄多糖VLP40和VLP80,在经过DEAE-52纤维素柱层析纯化,得到葡萄多糖VLP40-1和VLP80-3。
4.将超声波技术应用于葡萄多糖的提取,并用正交分析法优化提取工艺,得到了葡萄多糖的最优提取工艺:超声功率70W,在40℃下提取20min,料液比为1:2时,葡萄粗多糖提取率为3.04%,其多糖含量为6.83%,超声波辅助提取时,粗多糖提取率和多糖含量均高于传统热水浸提,综合考虑时间和能耗因素,超声波提取更具优势。
5.对VLP40和VLP80的物性:溶解性、吸油性、起泡性、泡持性、吸湿性、保湿性进行了研究。葡萄多糖VLP40和VLP80均能溶于水,不溶于乙醇、丙酮、乙酸乙酯等高浓度的有机溶剂。室温条件下,VLP40和VLP80的溶解度分别为0.14g/ml和0.05g/ml,且溶解度随温度的升高而增大。
6.葡萄多糖具有吸油性和起泡性,添加NaCl和蔗糖对其吸油性和起泡性影响不是很大;在相对湿度为43%和81%的环境中,VLP40的吸湿性均好于VLP80;在相对湿度为43%的环境中,VLP80的保湿性能要优于VLP40,而在干燥的硅胶环境中,VLP40的保湿性能与VLP80相差不大。
7. VLP40和VLP80物性的研究结果表明,VLP40各项性能指标均优于VLP80,所以实际生产中建议采用VLP40开发葡萄多糖类食品和保健品,特别是面包焙烤中具有很强的应用潜力。
8.抗氧化活性试验研究了葡萄多糖VLP40和VLP80对O_2~-·和·OH的清除能力以及其还原能力,结果表明VLP40和VLP80都具有O_2~-·和·OH的清除能力和一定的还原能力,其清除率随浓度的增加都有所增强,VLP40对O_2~-·和·OH的清除能力略强于VLP80。
Vitis vinifera L. is the famous fruit and wine stuff which is recorded as medicine by pharmacopoeia. This article studied on the extraction and purification of polysaccharides isolated from Xinjiang Vitis vinifera L.. Physical properities were also evaluated for the further research and develop of Vitis vinifera L. polysaccharides .The main results were shown as follows:
1. Sevag,enzymatic hydrolysis and combination of enzyme and Sevag were used to remove protein from polysaccharides of Vitis vinifera L. The results showed that with concentration of papain 0.14%, the polysaccharide solution was enzymatic hydrolysis for 2 hours in 60℃water and pH6.5,and after 4 times treatment of sevag method ,the removal percentage of protein was 72.14%,and the loss of polysaccharide was 10.91%.The combination of enzyme and Sevag was an effective and alternative for the common methods which demonstrated higher removal of percentage of protein and lower loss of polysaccharide.
2. The deproteinized polysaccharide was decolored with macroporous resin. The best way to decolor the Vitis vinifera L.polysaccharide was using the HPD-300 macroporous resin, and the macroporous resin amount used was 24g/100ml, adsorbing 3h at 30℃.The decolorization and recycle effect of HPD-300 were well.
3. VLP40 and VLP80 were isolated after deproteinized by enzyme combined with Sevag and dialysised with distilled water and the average molecular weight is more than 8000D. VLP40 and VLP80 were further purified by DEAE-52 cellulose column chromatography and collected VLP40-1 and VLP80-3.
4. Vitis vinifera L. polysaccharides was obtained by using ultrasonic wave extraction method .On the baisis of orthogonal test, the optimum condition of the extraction was carried out: ultrasonic power 70W, temperature 40℃for 20min and the ratio of solid-liquid was 1:2.The extraction rate of polysaccharides was 3.04% .Considered time and resources using , ultrasonic wave extraction method had more advantages than water extraction method.
5. Solubility, capacities of oil absorption, frothing, froth maintenance, moisture、absorption, moisture retention, of VLP 40and VLP80 were studied in the research. VLP 40 and VLP80 could dissolve in water but not in high concentration of organic solvent. With the increasing of temperature, their solubility increased. Solubility of VLP40 and VLP 80 was 0.14g/ml and 0.05/ml.
6.Vitis vinifera L. polysaccharides had capacities of oil absorption and frothing, but NaCI and sucrose both had little effect on the capacities of oil absorption and froth of VLP40 and VLP80. In the environment of relative humidity was 43% and 81%, Capacity of moisture absorption of VLP40 was better than that of VLP80. In the environment of relative humidity is 43%, Capacity of moisture retention of VLP80 was better than that of VLP40,and capacities of moisture retention of VLP 40 and VLP80 were almost the same in the environment of silica gel.
7.Physical properities of VLP40 were better than those of VLP 80, So VLP 40 could be used in food or healthy food under the consideration of commercial and practical food processing, especially in bread-baking industry.
8. The antioxidative activity of Vitis vinifera L. polysaccharides: VLP40 and VLP80 had the ability of scavenging oxygen free radicals, hydroxyl free radical and some reducing ability. With the increasing of concentration, Oxygen free radicals and hydroxyl free radical scavenging ability of VLP40 was higher than VLP80.
1.比较了Sevag法、酶法、酶法与Sevag法结合脱蛋白的效果,结果表明酶法与Sevag法结合脱蛋白效果最佳,其最佳处理条件为酶用量0.l4%,pH6.5,于60℃下保温2h,再经Sevag脱蛋白4次去除残留蛋白质,去蛋白效果最理想此时蛋白脱除率为72.14%,多糖损失率为l0.91%。
2.采用HPD-300型树脂,30℃下在摇床中吸附3h,树脂用量24g/100ml时,对葡萄多糖的脱色效果最好,该树脂再生后脱色效果良好,可重复利用。
3.透析去除小分子杂质和无机离子后得到分子量大于8000D的葡萄多糖VLP40和VLP80,在经过DEAE-52纤维素柱层析纯化,得到葡萄多糖VLP40-1和VLP80-3。
4.将超声波技术应用于葡萄多糖的提取,并用正交分析法优化提取工艺,得到了葡萄多糖的最优提取工艺:超声功率70W,在40℃下提取20min,料液比为1:2时,葡萄粗多糖提取率为3.04%,其多糖含量为6.83%,超声波辅助提取时,粗多糖提取率和多糖含量均高于传统热水浸提,综合考虑时间和能耗因素,超声波提取更具优势。
5.对VLP40和VLP80的物性:溶解性、吸油性、起泡性、泡持性、吸湿性、保湿性进行了研究。葡萄多糖VLP40和VLP80均能溶于水,不溶于乙醇、丙酮、乙酸乙酯等高浓度的有机溶剂。室温条件下,VLP40和VLP80的溶解度分别为0.14g/ml和0.05g/ml,且溶解度随温度的升高而增大。
6.葡萄多糖具有吸油性和起泡性,添加NaCl和蔗糖对其吸油性和起泡性影响不是很大;在相对湿度为43%和81%的环境中,VLP40的吸湿性均好于VLP80;在相对湿度为43%的环境中,VLP80的保湿性能要优于VLP40,而在干燥的硅胶环境中,VLP40的保湿性能与VLP80相差不大。
7. VLP40和VLP80物性的研究结果表明,VLP40各项性能指标均优于VLP80,所以实际生产中建议采用VLP40开发葡萄多糖类食品和保健品,特别是面包焙烤中具有很强的应用潜力。
8.抗氧化活性试验研究了葡萄多糖VLP40和VLP80对O_2~-·和·OH的清除能力以及其还原能力,结果表明VLP40和VLP80都具有O_2~-·和·OH的清除能力和一定的还原能力,其清除率随浓度的增加都有所增强,VLP40对O_2~-·和·OH的清除能力略强于VLP80。
Vitis vinifera L. is the famous fruit and wine stuff which is recorded as medicine by pharmacopoeia. This article studied on the extraction and purification of polysaccharides isolated from Xinjiang Vitis vinifera L.. Physical properities were also evaluated for the further research and develop of Vitis vinifera L. polysaccharides .The main results were shown as follows:
1. Sevag,enzymatic hydrolysis and combination of enzyme and Sevag were used to remove protein from polysaccharides of Vitis vinifera L. The results showed that with concentration of papain 0.14%, the polysaccharide solution was enzymatic hydrolysis for 2 hours in 60℃water and pH6.5,and after 4 times treatment of sevag method ,the removal percentage of protein was 72.14%,and the loss of polysaccharide was 10.91%.The combination of enzyme and Sevag was an effective and alternative for the common methods which demonstrated higher removal of percentage of protein and lower loss of polysaccharide.
2. The deproteinized polysaccharide was decolored with macroporous resin. The best way to decolor the Vitis vinifera L.polysaccharide was using the HPD-300 macroporous resin, and the macroporous resin amount used was 24g/100ml, adsorbing 3h at 30℃.The decolorization and recycle effect of HPD-300 were well.
3. VLP40 and VLP80 were isolated after deproteinized by enzyme combined with Sevag and dialysised with distilled water and the average molecular weight is more than 8000D. VLP40 and VLP80 were further purified by DEAE-52 cellulose column chromatography and collected VLP40-1 and VLP80-3.
4. Vitis vinifera L. polysaccharides was obtained by using ultrasonic wave extraction method .On the baisis of orthogonal test, the optimum condition of the extraction was carried out: ultrasonic power 70W, temperature 40℃for 20min and the ratio of solid-liquid was 1:2.The extraction rate of polysaccharides was 3.04% .Considered time and resources using , ultrasonic wave extraction method had more advantages than water extraction method.
5. Solubility, capacities of oil absorption, frothing, froth maintenance, moisture、absorption, moisture retention, of VLP 40and VLP80 were studied in the research. VLP 40 and VLP80 could dissolve in water but not in high concentration of organic solvent. With the increasing of temperature, their solubility increased. Solubility of VLP40 and VLP 80 was 0.14g/ml and 0.05/ml.
6.Vitis vinifera L. polysaccharides had capacities of oil absorption and frothing, but NaCI and sucrose both had little effect on the capacities of oil absorption and froth of VLP40 and VLP80. In the environment of relative humidity was 43% and 81%, Capacity of moisture absorption of VLP40 was better than that of VLP80. In the environment of relative humidity is 43%, Capacity of moisture retention of VLP80 was better than that of VLP40,and capacities of moisture retention of VLP 40 and VLP80 were almost the same in the environment of silica gel.
7.Physical properities of VLP40 were better than those of VLP 80, So VLP 40 could be used in food or healthy food under the consideration of commercial and practical food processing, especially in bread-baking industry.
8. The antioxidative activity of Vitis vinifera L. polysaccharides: VLP40 and VLP80 had the ability of scavenging oxygen free radicals, hydroxyl free radical and some reducing ability. With the increasing of concentration, Oxygen free radicals and hydroxyl free radical scavenging ability of VLP40 was higher than VLP80.
引文
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