【作者】 赖富饶；

【导师】 温其标； 吴晖；

【作者基本信息】 华南理工大学 ， 粮食、油脂与植物蛋白工程， 2010， 博士

【摘要】 本文以豆类加工的副产品-豆皮为原料,对豆皮果胶多糖的提取、分离纯化、理化性质、结构分析、体外抗氧化活性、胆酸结合能力以及体内降血脂功能进行了较为系统的研究,研究结果表明:豆皮的主要成分为不溶性纤维、可溶性纤维和蛋白质。在单因素试验的基础上,对大豆皮果胶多糖的提取工艺进行二次通用旋转组合优化,得到最佳提取条件为pH值5.5、水料比25:1(mL:g)、提取温度90℃、提取时间68 min、提取2次,大豆皮粗多糖SCP的得率为10.17%;将响应面分析法用于优化绿豆皮多糖的超声辅助提取工艺,对绿豆皮多糖得率影响的因素依次为:超声功率>pH值>超声时间,最适提取条件为:pH值4.6、超声功率155 W、超声时间40 min,重复3次,绿豆皮粗多糖MCP的得率为8.54%。将初步纯化后的豆皮多糖SHP和MHP经DEAE-纤维离子交换柱分离和Sephadex G-100凝胶柱层析,SHP得到3个纯化组分:SHP1、SHP2、SHP3,其得率分别为24.31%、49.70%和11.52%,总回收率85.53%。MHP得到2个组分:MHP1、MHP2,其得率分别27.63%、59.21%,总回收率为86.84%。经纯度鉴定,5个多糖组分都为均一性多糖,采用凝胶渗透色谱(GPC)法测定多糖组分的分子量,得SHP1、SHP2、SHP3、MHP1、MHP2的分子量分别为149342 Da、72799 Da、45554 Da、83236 Da、45012 Da。通过对豆皮粗多糖和纯化组分的体外抗氧化活性的测定评价两种豆皮多糖的抗氧化能力,结果显示,MHP、MHP1和MHP2的体外抗氧化活性随浓度的增大而增强,且具有比SHP及其组分更强的还原能力、抗氧化性、O ?2?、? OH和DPPH自由基清除能力以及邻苯三酚自氧化的抑制效果。另外,纯化组分MHP1表现出比粗多糖MHP更强的抗氧化活性。比较了粗多糖与纯化组分的体外胆酸结合能力,结果显示粗多糖具有较强的胆酸结合能力,分级纯化后,多糖组分的体外胆酸结合能力呈下降趋势,表明分离纯化过程会对多糖中的其他活性造成损失从而影响其体外胆酸结合的能力,多糖与胆酸的结合与其自身的物化结构及与胆酸的活性结合位点有关。采用红外光谱(FT-IR)、核磁共振(NMR)图谱分析、气相色谱(GC)、部分酸水解、高碘酸氧化-Smith降解、甲基化分析等对SHP1、SHP2和SHP3进行了结构分析,显示SHP1、SHP2和SHP3为弱酸性或酸性杂多糖,糖醛酸含量分别为17.31%、57.36%和26.71%,SHP1的中性糖主要为甘露糖和半乳糖,SHP2主要由鼠李糖、半乳糖和阿拉伯糖组成,SHP3则主要由阿拉伯糖、甘露糖、半乳糖和鼠李糖组成。经分析,SHP2的主链由→2)Rhap(1→4)GalA(1→2)Rhap→重复单元构成,侧链主要由→1)Araf(3,5→)Araf(3→和→1)Galp(6→3)Araf(1→)Galp(6→组成。豆皮多糖具有明显的调节血脂作用。对高脂饮食小鼠每天灌胃50 mg/kg·bw和180 mg/kg·bw的大豆皮多糖组分SHPb和绿豆皮多糖组分MHPa,可有效抑制高脂饮食小鼠体重的过快增长,显著降低血清TC、TG含量,提高HDL-C水平,并能降低动脉粥样硬化指数;能有效抑制肝组织中TC、TG水平。另外,MHPa能显著降低高脂饮食小鼠血清和肝脏的MDA浓度,提高SOD和GSH-Px活力;进食豆皮多糖可增加小鼠体内的胆固醇和胆酸的排泄,降低高脂血症发生的风险。更多还原

【Abstract】 The extraction, purification, physicochemical properties, structural analysis, in vitro antioxidant activity, of polysaccharide from soybean and mung bean hull, as well as in vitro bile acid binding capacity and hypolipemic effects were investigated in this study. The main contents and results are as follows:The main compounds in soybean hull are insoluble fiber, soluble fiber and protein. On the basis of single-factor experiments, the extraction technique of soybean hull polysaccharide was optimized by quadratic general rotary unitized design. The results showed that the optimum extraction conditions of soybean hull polysaccharide were pH of 5.5, ratio of water to solid of 25:1 (mL:g), extraction temperature of 90℃, extraction time of 68 min. Under these conditions, the yield of soybean hull crude polysaccharide (SCP) was 10.17% by two-time extraction. Similarly, the response surface analysis (RSA) was used to optimize the extraction conditions of mung bean hull crude polysaccharide (MCP). The best extraction conditions were pH value of 4.6, ultrasonic power of 155 W, extraction time of 40 min, and repeated 3 times. In this case, the yield of MCP was up to 8.54 %.After deproteinization by Sevag reagent and decoloration by macroporous absorbent resin, the pre-purified polysaccharide SHP and MHP were obtained. Subsequently, SHP and MHP were fractioned by DEAE-cellulose anion exchange column and purified by Sephadex G-100 gel permeation column. There were three fractions obtained from SHP marked SHP1, SHP2 and SHP3, and two from MHP marked MHP1 and MHP2. The yields of five fractions were 24.31%, 49.70%, 11.52%, 27.63%, and 59.21%, with molecule weights of 149342 Da, 72799 Da, 45554 Da, 83236 Da, 45012 Da, respectively.The evaluation of antioxidant activity of polysaccharide fractions was carried out. The results indicated that the antioxidant capacities of MHP, MHP1 and MHP2 increased with the concentration. Furthermore, The reducing power, antioxidant activity, scavenging activity of O ?2?, ? OH and DPPH free radical, as well as inhibition effects of self-oxidation of 1,2,3-phentriol of MHP, MHP1 and MHP2 were stronger than those of SHP, SHP1, SHP2 and SHP3. Besides, MHP1 exhibited the best antioxidant activity among these three substances. The comparison of the ability to bind bile acid in vitro of seven substances demonstrated that SCP and MCP could bind more bile acid than purified fractions. These results indicated that the ability to bind bile acid of polysaccharide was affected by the purification process, physical and chemical structures of purified fractions or active binding sites.The Fourier transformation infrared spectroscopy (FT-IR), nuclear magnetic resonance (NMR) analysis, gas chromatography (GC), partial acid hydrolysis, periodate oxidation– Smith degradation and methylation analysis were adopted to characterize the structures of SHP1, SHP2 and SHP3. The purified fractions were acid heteropolysaccharide without protein or nucleic acid. The content of uronic acid was 17.31%, 57.36%, 26.71%, respectively. Moreover, the neutral sugar of SHP1 mainly was mannose and galactose, while SHP2 consists of rhamnose, galactose and arabinose, SHP3 contains arabinose, mannose and rhamnose.Methylation analysis indicated that the main chain of SHP2 was the repeated units of→2)Rhap(1→4)GalA(1→2Rhap(1,4→, and the main branched chain of SHP2 was glycan composed by→1)Araf(3,5→)Araf(3→and→1)Galp(6→3)Araf(1→)Galp(6→.The polysaccharides from soybean and mung bean hull could adjust the blood lipid level of mice with high fat diet significantly. After lavage of fraction SHPb and fraction MHPa at the dose of 50 mg/kg·bw and 180 mg/kg·bw for 4 weeks, respectively, the excessive increase of body weight was inhibited. The levels of serum TC, TG, LDL-C and atherosclerotic index (AI) reduced while HDL-C increased significantly. And, the levels of hepatic TC, TG were effectively maintained in a healthy status. Additionally, MHPa could lower the serum and liver malondialdehyde (MDA) concentration of mice with high fat diet and increase activity of SOD and GSH-Px. Obviously, the polysaccharides from soybean and mung bean hull could promote the excretion of TC and bile acid, and lessen the risk of hyperlipoidemia.更多还原