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玛咖真伪、有效成分研究及质量评价
陈金金
学位类型博士
导师赵兵
2015-09
学位授予单位中国科学院研究生院
学位授予地点北京
学位专业生物化工
关键词玛咖 真伪鉴别 膳食纤维 玛咖酰胺 质量评价
其他摘要

玛咖(Lepidium meyenii)是生长在高海拔地区的十字花科草本植物,其根部具有丰富的营养价值和药用价值,被长期作为食物和民族药使用。2011年我国卫生部批准玛咖粉为新资源食品,使玛咖产品得以合法进入我国市场。随着玛咖市场的快速兴起,迫切需要相关技术以解决玛咖真伪鉴别、废弃资源利用、活性成分分析及质量安全评价等问题。为解决上述问题,本论文建立了玛咖真伪鉴别、高活性膳食纤维制备及玛咖标志性成分玛咖酰胺的合成及检测方法,研究了不同样品中玛咖酰胺组成和含量,全面分析了玛咖样品中感官、理化及微生物指标,为玛咖资源利用和质量安全评价奠定了基础。本文取得的主要研究结果如下:(1)建立了基于ITS序列的玛咖真伪鉴别方法。用SDS法分别提取了43份不同玛咖样品的总DNA,扩增得到玛咖ITS序列,经过测序和比对分析,结果表明,玛咖ITS序列全长621 bp,无变异位点,具有种内一致性;NCBI数据库搜集了19种玛咖伪品ITS序列,比对分析发现,玛咖ITS序列与其伪品的ITS序列存在5-332 bp的变异位点,具有种间特异性。因此,建立了以ITS序列作为DNA条形码的玛咖真伪鉴别方法,并利用掺杂10%芜菁的玛咖粉验证了方法的可靠性。(2)利用玛咖泡酒残渣制备了高活性膳食纤维。以玛咖泡酒残渣为原料,通过酶法和化学法分别制备了膳食纤维。分析了两种膳食纤维的表面形态结构、红外谱图、基础化学组成及单糖组成,并进一步研究了溶胀率、持水率、持油率、葡萄糖吸附率和葡萄糖阻滞率。结果表明酶法制备的玛咖膳食纤维具有更疏松的空间结构,更高的总纤维含量(81.10%),更丰富的单糖组分(鼠李糖:阿拉伯糖:木糖:甘露糖:葡萄糖:半乳糖为1.84:1.53:0.29:0.26:1.63:1),以及更高的溶胀率(26.17 mL/g),持水率(16.29 g/g),持油率(5.79 g/g),葡萄糖吸附率(905.91 μmol/g)和葡萄糖阻滞率(24.59%)。(3)合成玛咖酰胺对照品,建立HPLC检测方法并优化了提取方法。采用EDC/HOAt法合成了九种玛咖酰胺,并经HPLC-UV,HPLC-MS2及NMR进行结构和纯度检测,表明该法合成的玛咖酰胺得率高(≥90%),纯度高(>95%);采用正交实验优化了超声辅助石油醚提取玛咖酰胺的方法,确定了提取条件,超声功率200 W,温度50 oC,时间15 min,液料比40:1;建立了玛咖酰胺等度HPLC检测方法并进行了方法学考察,采用C18色谱柱,流动相为乙腈:水(含0.2%甲酸)为90:10,流速0.6 mL/min。验证试验显示9种玛咖酰胺对照品线性关系良好(r2 ≥ 0.9990),日内和日间相对标准偏差分别≤2.12%和≤3.21%,证明该方法具有良好的重复性和准确度。(4)研究了不同玛咖样品中酰胺组成和含量。研究了不同颜色、不同部位、不同大小、不同来源、不同产品类型和不同干燥方式的玛咖样品中酰胺种类组成和含量差异。结果表明,不同玛咖样品中酰胺组成相似,但含量差异较大。总酰胺含量黑根(871.72 μg/g)>黄根 (704.58 μg/g)>紫根(676.83 μg/g),须根(1942.90 μg/g)>主根(704.58 μg/g)>叶片 (40.54 μg/g)>种子(0),小根(928.92 μg/g)>大根 (435.79 μg/g),国内(41.24-502.97 μg/g)>进口(578.42-2773.92 μg/g),精片(1235.67 μg/g)>干片(1036.94 μg/g)>鲜根(264.15 μg/g),风干(280.78μg/g)>冻干(58.70 μg/g),且不同干燥样品中总酰胺含量随保存时间明显升高。(5)制定了《玛咖干制品》云南省地方标准。根据《食品安全法》要求,检测了云南省79份玛咖样品的感官指标、理化指标和微生物指标,将玛咖酰胺含量作为玛咖干制品(果、片、粉)质量分级指标,制定出《云南省食品安全地方标准 玛咖干制品》,该标准于2015年9月27日正式施行。

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Maca (Lepidium meyenii) is a herbaceous plant of Brassicaceae family that grows in high plateaus. Its root has been used as both food and folk medicine for high nutritional and medical value for a long time. In 2011, maca powder was authorized by China's national ministry of health to be a new resource food, which legalized the maca products. With the rapid development of maca industry, technologies for identification, residue utilization, bioactive constituent analysis and quality and safety evaluation of maca are quite needed. Focusing on problems mentioned above, in this work, methods were established for maca identification, high-activity dietary fiber preparation, and the marker compounds, macamides, synthesis and detection. Macamide composition and content were studied in different maca samples. Furthermore, sensorial, physicochemical and microbial properties of maca samples were analyzed comprehensively. This work laid the basis for the resource utilization and quality and safety evaluation of maca. The major findings are as follows:(1) A method for maca identification was established based on the ITS (Internal Transcribed Spacer) sequence.Total DNA of 43 different maca samples were extracted respectively using SDS method. Then the ITS sequences were amplified, sequenced and aligned. The result showed that the ITS sequence of maca was 621 bp long and without any variation site, which indicated the intraspecies consistent of maca ITS. Furthermore, nineteen ITS sequences of maca adulterants were collected in NCBI and aligned. The result showed that there were 5 to 332 bp variation sites betweem ITS seqences of maca and its adulterants, which indicated the interspecies specific of maca ITS. Therefore, a method for maca identification was established using the ITS sequence as the DNA barcoding and verified by maca powder adulterated with 10 % of turnip.(2) High-activity dietary fiber was prepared using maca liquor residue.Using maca liquor residue as material, dietary fibers were prepared by enzymic and chemical methods, respectively. Surface morphology, infrared spectra, proximate composition and monosaccharide composition of these fibers were analyzed. And the swelling capacity, water holding capacity, oil holding capacity, glucose adsorption capacity and glucose retardation index were futher studied. The results showed that the dietary fiber prepared by enzymic method had the more loose morphology, higher content of total fiber (81.10%), more abundant monosaccharide composition (rhamnose : arabinose : xylose : mannose : glucose : galactose of 1.84 : 1.53 : 0.29 : 0.26 : 1.63 : 1) and higher swelling capacity (26.17 mL/g), water holding capacity (16.29 g/g), oil holding capacity (5.79 g/g), glucose adsorption capacity (905.91μmol/g) and glucose retardation index (24.59%).(3) Methods for macamide synthesis and high performance liquid chromatography (HPLC) detection were developed and the extraction conditions were optimized.Nine macamides were synthesized by the EDC/HOAt method. The structures and purities of these synthetic macamides were verified by HPLC-UV, HPLC-MS2 and NMR. The results showed that high yields (≥ 90%) and purities (> 95%) of macamides were obtained by this synthetic method. Furthermore, a method for ultrasonic-assisted extraction of macamides by petroleum ether was optimized based on orthogonal degign. The optimal extraction conditions were as follows: ultrasonic power, 200 W; temperature, 50 oC; time, 15 min; solvent to sample ratio, 40 : 1. Besides, an isocratic HPLC method was developed and validated for the detection of macamides. The HPLC conditions were C18 column eluted with acetonitrile: water (containing 0.2% formic acid) of 90:10 at 0.6 mL/min. Calibration curves of all analytes showed good linearity (r2 ≥ 0.9990). The intra-day and inter-day relative standard deviations (RSDs) of the nine macamides were ≤ 2.12% and ≤ 3.21%, respectively, which indicated that the method had good repeatability and precision.(4) Macamide composition and content of different maca samples were investigated.Macamide composition and content of maca with different colors, parts, sizes, sources, product types and dryinng methods were analyzed. The results showed that the compositions were similar, but the contents showed significant variance among all tested maca samples. For the total content of macamides, black root (871.72 μg/g) > yellow root (704.58 μg/g) > purple root (676.83 μg/g), fibrous root (1942.90 μg/g) > main root (704.58 μg/g) > leaf (40.54 μg/g) > seed (0), small root (928.92 μg/g) > big root (435.79 μg/g), domestic (41.24-502.97 μg/g) > imported (578.42-2773.92 μg/g), essential tablet (1235.67 μg/g) > dry slice (1036.94 μg/g) > fresh root (264.15 μg/g), air-dried root (280.78μg/g) > freeze-dried root (58.70 μg/g). The total contents of macamides increased obviously with the storage time in different dried maca.(5) The Local food safety standard of Yunnan Province of maca dried products was established.According to Food Safety Law of China, sensorial, physicochemical and microbial properties of 79 maca samples from Yunnan were analyzed. The content of macamides was used as the quality indicator for maca dried products (root, slice and powder). “Local food safety standard of Yunnan province of maca dried products” was established and enacted on Sept. 27, 2015.Key words:Maca, Identification, Dietary fiber, Macamides, Quality evaluatation 

语种中文
文献类型学位论文
条目标识符http://ir.ipe.ac.cn/handle/122111/21346
专题研究所(批量导入)
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陈金金. 玛咖真伪、有效成分研究及质量评价[D]. 北京. 中国科学院研究生院,2015.
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