【作者】 俞卓裕；

【导师】 张英； 吴晓琴；

【作者基本信息】 浙江大学 ， 食品科学与工程， 2003， 硕士

【摘要】 目前国际社会对安全有效、制剂均一、品质稳定、质量可控的药用植物提取物的需求呈急剧增长的态势，并开始关注竹叶黄酮。本课题结合近年来国内在中药现代化方面所做的大量探索性工作，依托导师在竹叶有效成分研究开发领域长期的工作积累，以天然来源的刚竹属(淡)竹叶为原料，将微波协助萃取(MAE)技术应用于竹叶有效成分的提取，通过超滤(UF)和纳滤(NF)的组合式膜分离技术分离药用活性部位，选择安全性好、对类黄酮选择性高的聚酰胺作为吸附材料进一步富集目标产物，得到了符合中药Ⅱ类新药原料药要求(总黄酮糖苷≥50％)的高精度的竹叶总黄酮制剂。同时，在小试基础上进行了放大试验，在工艺技术研究的同时整合现代质量控制手段，使研究成果具备产业应用的基础。研究内容和结果分述如下： 1．将MAE技术应用于竹叶有效成分的提取，以总黄酮(Total Favonoids，TF)含量作为响应值，进行四因素三水平的正交试验。结果表明，微波功率在700～1400W的范围内，优化的工艺参数为：乙醇浓度为40％、固液比1／20、萃取时间40min。而在模拟体系的单因素试验中，对芦丁增溶作用明显的β-环糊精，在竹叶黄酮提取的正交试验中对方差的贡献不显著。与经典的索氏抽提法和有机溶剂热回流提取法相比，MAE可使竹叶总黄酮的萃取率提高150％～160％，同时，在缩短提取时间、减少有机溶剂用量等方面体现了独特的优越性。 2．通过对纯芦丁(黄酮醇糖苷)体系的超滤行为的系统研究，探讨了在超滤过程中，料液的压力、温度、流速和pH值与渗透通量之间的关系，揭示了溶质的扩散效应和料液流动的剪效应对透过率的联立效应。对于一个可在膜表面形成可逆覆盖层的横流体系来说，在起始时间段由于覆盖层的形成，渗透通量会逐渐下降，此后体系将会达到某个比较稳定的状态，一般来说，稳态时膜通量的大小随温度和进料溢流速度的升高而增加，但会随着进料浓度的增加而下降。浙江大学硕士学位论文俞卓裕竹叶铸用有获钾位伪板取.令离、化化二艺枚术肠研究 以竹叶粗提物(EOB一f04)为起始原料，采用二段超滤(UFI和UF川和一段纳滤(NF)的组合进一步分离活性部位。在运用比色法测定总黄酮含量的同时，采用HPLC法定向检测目标产物、定量四种主要的碳昔黄酮，对工艺过程的有效性进行实时监控。UFI、UFn和NF三段分级产物中总黄酮干基含量分别达到31.20%、40.20%和25.50%，相应的截留率为12.31%、36.61%和22.64%;总固形物的截留率分别为10.90%、25.13%和24.51%;四个碳普黄酮合计的截留率为10.49%、37.86%和30.44%。其中UFll截留物的干基总黄酮含量超过了40%，是一种中、高精度的竹叶总黄酮制剂，可直接用作膳食补充剂、保健功能因子、高档护肤因子、食品添加剂等，也可经进一步精制成为H类新药原料药。表明，该组合式膜分离过程起到了分离杂质、富集有效部位和浓缩料液的三重作用。 3.以竹叶提取物(Ebl971)为原料，比较大孔树脂和聚酞胺两种吸附材料对竹叶黄酮的精制效果，得出二者对总黄酮的最大吸附容量均在14%一15%之间，最适吸附pH值在7.5左右，理想的乙醇洗脱浓度为50%。聚酞胺对竹叶黄酮的选择吸附性能优于大孔树脂，对有效部位的富集效果显著强于大孔树脂，如在聚酞胺柱的50%乙醇洗脱部位中，干基总黄酮含量达到了65.19%，回收率为30.95%;而大孔树脂柱相应部位中干基总黄酮含量仅为42.78%，回收率为27.95%。 在中型柱的放大试验中，选用UFH截留液为起始物料，用聚酞胺作为吸附剂，水洗除去大部分酚类物质，用50%的乙醇一步洗脱，洗脱液经减压干燥后，比色法测得干基总黄酮含量为53.9%，HPLC法测得其中鱿草昔、异药草普、牡荆昔和异牡荆昔的含量合计为10.49%。聚酞胺精制前后，总黄酮含量仅提高了13.6%，而四种碳昔的干基含量从2.05%上升到10.49%，增加了5倍。 综上所述，微波协助萃取能显著提高竹叶有效成分的得率及生产效率，粗提物经组合式膜分离技术除杂，结合聚酞胺吸附一解析技术精制，可获得符合中药H类新药原料药要求的、有效部位含量不低于50%的竹叶总黄酮制剂。表明微波协助萃取、膜分离和吸附~解析三种高新技术的有机组合，在竹叶药用活性部位的提取、分离和精制中收到了理想的效果，研究结果对植物类黄酮的生产具有普遍的指导意义。更多还原

【Abstract】 Nowadays, more and more herb-plant extracts with high safety, availability, efficiency, uniformity and stability have received broad attention from all over the world. Among of those, the total flavonoids extracted from bamboo leaves emerged recently. Based on a lot of domestic researches in the field of Chinese traditional medicine modernization, and combined with a long-term groundwork conducted by my supervisor in the R&D of effective fractions in bamboo leaves, this paper was designed to do next series work: taking the leaves of Phyllostachys bamboo as raw material, using microwave assisted extraction (MAE), membrane separation(MS) and Asorption - Desorption (AD) technologies, a kind of refined preparation of bamboo-leaf-total- flavonoids (BLTF) was given out, which contained total flavone glycosides in above 50% in dry base and could be used as raw-drug directly. At the same time, modern technology means was combined with advanced analysis methods; experiment capacity was scaled-up step by step; and so that the test results can be cited in practical production. The main research contents and results are as follows.Applying the MAE in the extraction of bamboo leaf effective components, and taking the total flavonoids (TF) as a responsive index, a orthogonal test with four factors and three levels, table L9(34), was conducted. Results showed that in the microwave power range of 700 to 1400W, optimum technologic parameters were ethanol concentration 40%, solid-liquid ratio 1 to 20, and extracting time 40min. However, -cyclodextrin which displayed significant solubility-enhancing effect on rutin in a mimic system had not behaved same action in this orthogonal test. Compared with by a classical Soxhlet’s extracting and a industrialized heat reflux, the extracting yield of BLTF using by MAE was elevated to 150% to 160%. Furthermore, MAE exhibited particular advantages to cut extracting time, reduce organic solvent dosage and so on.A systematic study on the ultrafiltration (UF) process of pure rutin solution wasmade to discover the relationship between permeation flux and pressure, temperature, bulk velocity or pH of treated solution. Results showed that membrane permeation flux depended on the combinative effect of solute diffusion and brush action of solution flow. As regards a cross-flow system that a reversible cover layer can be formed on the dial of membrane, due to the cover-layer, the permeation is decreasing gradually in the initial stage, and then keeping at a more stable level. Generally, the permeation flux of system on this stage will increase with the enhancement of temperature and fluid speed, and decrease with the elevation of solution concentration.The effective components in bamboo-leaf-crude-extract was further condensed using by a complex membrane separation technology, which includes two stage UF and one NF operation stages. At the same time of measuring TF by spectrometry, using HPLC to detect objective products and quantify four flavone C-glycosides, in order to assure the efficiency of technology process. TF contents in three graded fractions (UF1, UF2 and NF cut off parts) were individually 31.20%, 40.20% and 25.50% in dry base, and corresponding TF reserve ratios were 12.31%, 36.61% and 22.64%; the reserve ratios on solvent were 10.90%, 25.13% and 24.51%; and the reserve ratios on four flavone-C-glycosides were10.49%, 37.86% and 30.44%. Among of those, the part of UF2 cut-off contained TF in above 40%, which was a much refined total-flavonoid preparation and can be used as dietary supplement, functional factor, cosmetic and food additives, as well as further purification for drug material. Results showed that this combined membrane separation process exhibited multiple advantages: isolation impurity, enrichment effective components and concentration liquid volume.Comparing the purifying effect of absorption, and desorption between macroreticular resin and polyamide on BLTF, the adsorption capacity for both two sorbents was 14% to 15%, the optimum action parameters were pH 7.更多还原