黄秋葵花多糖的超声提取及其结构和抗氧化活性研究

发布时间：2018-02-15 15:48

本文关键词： 黄秋葵花多糖超声提取结构表征抗氧化活性　出处：《浙江大学》2017年硕士论文　论文类型：学位论文

【摘要】：黄秋葵(Abelmoschus esculentus(L.)Moench),原产非洲,其花中富含多糖、蛋白质、黄酮、脂肪酸、矿物质元素等,营养保健价值高,具有很大的开发潜力。国内外已有大量对黄秋葵嫩荚多糖的研究报道,但对黄秋葵花中同样富含的多糖类物质研究较少。本文通过比较不同提取方法所得多糖的理化性质以及抗氧化活性,探讨了提取方法对多糖性质的影响,优化了超声提取黄秋葵花多糖的工艺条件,并进一步分离纯化超声提取的多糖,对主要多糖组分的化学结构进行表征。1.以黄秋葵花作为原料,采用了热水、酸法、酶法、超声辅助提取的方法,分别得到四种粗多糖,其中酶法提取率最高。四种多糖的半乳糖醛酸含量在29.34%-52.63%,粗多糖中还检测出少量蛋白质、酚类物质。采用HPLC、红外光谱分析多糖的分子量、单糖组成和结构特征,结果表明得到的四种样品都具有多糖的典型红外光谱特征;四种多糖的单糖组成类型相同,主要由鼠李糖、半乳糖醛酸、半乳糖组成,但单糖的摩尔比有所不同;酶法和热水提取的多糖样品具有较高的分子量且特性粘度较大,而超声提取显著降低了多糖的分子量和特性粘度,同时使多糖分子量的分布更加均一。扫描电镜观察的结果显示,多糖的微观表面形态主要呈现不规则的薄层状。通过DPPH自由基清除能力、氧自由基吸收能力等5种经典体外抗氧化实验,评价除去了色素等杂质后多糖的抗氧化活性,结果表明四种多糖具有不同程度的抗氧化活性且具有剂量依赖性。其中,超声辅助法提取的多糖抗氧化活性最强。2.采用单因素试验考察超声功率、提取温度、提取时间、料液比等因素对黄秋葵花多糖提取率的影响,再通过响应面分析法优化黄秋葵花多糖的提取工艺,最终优化的提取工艺:提取温度55℃,提取时间30 min,超声功率85.5 W,在此条件下多糖的提取率为12.62%。3.采用DEAE-Q Sepharose Fast Flow色谱柱对超声提取的多糖进行分离纯化。其中,0.5 mol/LNaCl溶液洗脱下来的组分含量较高,且为均一多糖,命名为UOFP-2。用HPLC、高碘酸氧化-Smith降解法,结合FT-IR、1D-NMR和2D-NMR图谱对UOFP-2的理化性质及化学结构进行研究,发现UOFP-2由鼠李糖、半乳糖醛酸、半乳糖、阿拉伯糖和甘露糖组成,单糖摩尔比为32.11:20.67:44.41:1.51:1.30,多糖的主链由→2)-α-Rha-(1 →4)-α-GalA-(1 →交替的重复单元和同聚鼠李糖组成,以1→4、1→3连接的β-Gal为支链,连接在α-Rha的O-4位。多糖的构象采用SEC-MALLS-IR-VIS联用法研究和AFM观测,得到UOFP-2 的特性粘度为 1.78 dL/g,Mw 为 217 kDa,Mark-Houwink 方程中,特性常数α=0.31,表明UOFP-2在溶液中表现为球形构象,与AFM观测到的结果相符。4.多糖组分UOFP-2表现出了明显的羟基自由基清除活性,IC50为5.89 mg/mL,具有剂量依赖;UOFP-2的ORAC值为30.38μmol Trolox/g,CAA值为 342.98μumolQE/100g。
[Abstract]:Okra (Abelmoschus esculentus (L.) Moench), originating in Africa, the flowers are rich in polysaccharides, protein, flavonoids, fatty acids, mineral elements, high nutritional value, it has great potential for development. There are lots of okra pods polysaccharide research reports, but less study of polysaccharides also rich in okra. Through the comparison of physicochemical properties of the extraction methods of polysaccharide and antioxidant activity of different extraction methods on the effects of the nature of the polysaccharide, ultrasonic extraction of okra polysaccharide process conditions were optimized, and further purification of Polysaccharide by ultrasonic extraction, the main polysaccharide the chemical structure of the group.1. were characterized with okra as raw material, by hot water, acid method, enzymatic method, ultrasonic extraction method, four kinds of polysaccharide were obtained, of which the highest rate of enzymatic extraction. Four kinds of polysaccharides of galactose The content of uronic acid in 29.34%-52.63%, crude polysaccharide was also detected in a small amount of protein, phenolic compounds. Using HPLC, infrared spectroscopy analysis of the molecular weight of the polysaccharide, monosaccharide composition and structural characteristics, results show that the typical characteristics of infrared spectra of four samples are obtained with four kinds of polysaccharide; Monosaccharide composition of the polysaccharide of same type. Is mainly composed of rhamnose and galacturonic acid, galactose, but the molar ratio of different monosaccharide; enzyme method and hot water extraction of polysaccharide samples with high molecular weight and viscosity characteristics, and ultrasonic extraction significantly reduced the polysaccharide molecular weight and viscosity, and the distribution of molecular weight of the polysaccharide more uniform scanning electron microscopy. The results showed that the surface morphology of polysaccharides mainly showed irregular lamellar. By DPPH free radical scavenging capacity, oxygen radical absorbance ability of 5 kinds of classic antioxidant experiments, evaluation of removal The pigments and other impurities after polysaccharide antioxidant activity, the results show that four kinds of polysaccharides with different degrees of antioxidant activity in a dose-dependent manner. The ultrasound assisted extraction method of polysaccharides of the strongest antioxidant activity of.2. single factor test was used to investigate the ultrasonic power, extraction temperature, extraction time, ratio of material to liquid on extraction rate effect of okra polysaccharides by extraction process analysis to optimize the okra polysaccharide response surface, the optimized extraction process: extraction temperature 55, extraction time 30 min, ultrasonic power 85.5 W, the extraction rate of polysaccharides was 12.62%.3. by DEAE-Q Sepharose Fast Flow column for the extraction of polysaccharides by ultrasonic method the separation and purification of this condition. Among them, 0.5 mol/LNaCl solution eluted fractions were higher, and homogeneous polysaccharide, named UOFP-2. HPLC, periodate oxidation, -Smith degradation, combined with FT-IR, 1D-NMR Study on UOFP-2 and 2D-NMR. The chemical structure of UOFP-2, found by Li Tang rat, galacturonic acid, galactose, glucose and mannose in Arabia. The molar ratio of 32.11:20.67:44.41:1.51:1.30, the polysaccharide backbone by alpha - 2) -Rha- (1, 4) - -GalA- (1 - alpha alternating repetition unit and together with rhamnose, from 1 to 4,1 and 3 connected -Gal beta branched chain, connected to the alpha -Rha O-4. The polysaccharide conformation by SEC-MALLS-IR-VIS spectrometry study and AFM observation, UOFP-2 viscosity was 1.78 dL/g, Mw 217 kDa, Mark-Houwink equation and characteristic constant alpha =0.31 UOFP-2, that is in the form of spherical conformation in solution, and AFM observed results of.4. polysaccharide fraction UOFP-2 showed obvious hydroxyl radical scavenging activity, IC50 was 5.89 mg/mL, with dose dependence; UOFP-2 ORAC = 30.38 mol Trolox/g, CAA The value is 342.98 mu umolQE/100g.

【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TQ281

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