【作者】 黄优生；

【导师】 谢明勇；

【作者基本信息】 南昌大学 ， 食品科学， 2007， 硕士

【摘要】 山楂为蔷薇科植物，是一种重要的药食两用资源，在我国种植栽培面积广阔，产量可观。现代研究证实，山楂具有抗动脉粥样硬化，降血压，降血脂，增加冠脉血流量，抗氧化，防癌、抗癌，以及提高机体免疫力等生理活性，研究表明，这些生理活性与山楂中所含的黄酮、原花青素以及三萜酸等活性成分有关。但现有研究主要停留在一些简单的提取工艺及粗提物的加工上，这极大的影响了山楂及其活性成分的有效利用。为此，本研究构建了山楂总黄酮的分析方法；将超临界CO₂萃取新技术应用于山楂活性成分的提取中并优化了山楂总黄酮提取工艺，对各种提取物抗氧化功能进行了比较研究；结合HPLC-DAD-MS联用技术系统研究了山楂活性成分的指纹图谱，以期有效控制山楂药材质量。主要研究概述如下：1．将改进后的聚酰胺吸附与NaNO₂-Al（NO₃）₃比色法用于山楂总黄酮测定，减少了杂质的干扰，并且优化了显色时间，结果表明该方法精密度高，重现性好，显色稳定，加样回收率高，该方法能更准确的测定山楂总黄酮的含量。2．采用乙醇回流法提取山楂总黄酮，以测得山楂样品中总黄酮含量为指标，选用正交表L₉（3⁴）对山楂总黄酮提取工艺进行研究。试验结果表明：最佳提取条件为用70％乙醇，固液比为1：20，在70℃下提取2h，重复提取2次，此时总黄酮得率为3.97％。3．研究了超临界CO₂萃取技术提取山楂总黄酮的最佳工艺。通过考察萃取压力、萃取温度和萃取次数等影响因素，运用中心组合试验设计和响应面分析法分析得到最佳的萃取工艺条件为：萃取压力36.6Mpa，萃取温度47.8℃，萃取4次（40mL 95％乙醇夹带剂，萃取2h），最优工艺条件下的总黄酮得率为0.59％。4．通过研究山楂不同提取物对DPPH自由基和O₂^-·自由基的清除作用考察其抗氧化活性。结果表明，山楂各种不同提取方法获得的山楂提取物均有抗氧化活性，不同提取物抗氧化活性强弱为：醇提物＞超临界CO₂萃取物＞水提物，提取物量与抗氧化活性呈正相关。通过测定山楂提取物乙酸乙酯、正丁醇和萃余物三个萃取部分清除DPPH·自由基的能力，发现乙酸乙酯萃取部分清除DPPH·自由基能力和抗氧化活性最强。5．建立了不同山楂样品的HPLC指纹图谱，并采用HPLC-DAD-MS联用技术获得了指纹图谱的多维信息，参照现有文献，对指纹图谱中的主要色谱峰进行了结构归属的初步判断，鉴定了7个共有峰。对色谱指纹图谱进行相似度计算，获得了令人满意的结果，并生成了对照指纹图谱，该法为山楂样品的鉴定提供了较全面的信息，有助于提高山楂药材的质量控制水平，同时深化了HPLC-DAD-MS联用技术在中草药复杂组分分析和鉴定中的应用。更多还原

【Abstract】 Hawthorn （Crataegus pinnatifida）, referring to the Rosaceae family, has beenused as herbs and edibles for a long time and has been planted widely in China. Manyresearches showed that Hawthorn has many biological activities such as reducingblood pressure and total plasma cholesterol, treating the early stages of congestiveheart failure, anti-arteriosclerosis, anti-oxidation, anti-cancer and improvingimmunity of corporeity. These benefits were attributed to several effectivecomponents such as flavonoids, procyanidins and triterpene acids etc. However, theprevious researches mainly focused on the simple extraction techniques and theprocessing of crude extracts, as a result, the utilization of hawthorn and its effectivecomponents was badly affected. In this paper, An analytical method of totalflavonoids was established; SFE-CO₂ was used to extract the effective components ofhawthorn, and the extraction conditions of total flavonoids were optimized; Theantioxidant activities of different extracts were studied and compared; A systematicclassification method was developed for the quality assessment of hawthorn based onchemical fingerprints and HPLC-DAD-MS. The main results in the paper areconcluded as follows:1. The NaNO₂-Al（NO₃）₃ colorimetric method combined with the improvedpolycaprolactam purification was developed for the determination of totalflavonoids in hawthorn. A fast analytical method has been established by optimizingthe conditions for colorimetry and reducing the interference of impurity. The resultsshowed that the developed method could eliminate the interference of impurity andwas a better method for the determination of total flavonoids in hawthorn with highprecision and good reproducibility.2. Optimum conditions of the extraction of the total flavonoids from hawthornwith ethanol reflux were studied by using orthogonal experiment design. The resultsshowed that the optimum conditions for the extraction with water bath was 2h at70℃, the ratio of solvent to material of 20:1, 70％ethanol as the solvent, extraction 2 times. The yield of total flavonoids was 3.97％at the optimum condition.3. The SFE-CO₂ technique used to extract total flavonoids in hawthom wasoptimized. The optimization study involved a central composite rotatabledesign（CCRD） and response surface methodology（RSM）; the design was used tostudy the three factors with a potential influence on the extraction process, namely:extraction pressure, extraction temperature and extraction times. The results showedthat the optimum conditions were as follows: 36.6Mpa, 47.8℃and 4 times（40mL95％aqueous-ethanol and 2 hours）. The yield of total flavonoids was 0.61％at theoptimum conditions.4. The anti-oxidant effects of different Hawthom were studied in vitro models byusing DPPH and pyrogallol autooxidation methods. The results showed that they allhad anti-oxidant effects, and the anti-oxidant effects had a positive correlation withconcentration, and the anti-oxidative activities of the extract decreased in the order ofethanol bath, SFE-CO₂, water. The ethyl acetate part possesses the strongestscavenging effects on DPPH radical by compared with other polarity parts.5. The HPLC-DAD-MS method was used to study the HPLC fingerprint ofHawthorn. Meanwhile, seven common peaks were identified by compared their UVand MS spectra with the references. Satisfactory results were obtained by calculatingthe similarity. This method provides complete information for the identification ofHawthorn, and benefits for the quality control of Hawthorn. In addition, the study canpromote the utility of HPLC-DAD-MS in the analysis and identification of complexcomponents in herbal materials.更多还原