龙血竭纳米粒的制备及其对皮肤损伤的愈合作用研究

发布时间：2018-08-28 16:00

【摘要】：龙血竭是由百合科植物剑叶龙血树Dracaena cochinchinensis (Lour.) S. C. Chen的树脂经加工制成,含有黄酮、挥发油等成分,具有活血散淤、定痛止血和敛疮生肌的功能,龙血竭在水中溶解度小,难以吸收,生物利用度低。因此,如能制备出水溶性好的新剂型,将有助于提高其生物利用度,为其广泛应用创造条件。纳米给药系统(nanoparticle drug delivery system, NDDS)是指药物或药物与药用材料一起形成的粒径为1-1000nm的纳米级药物输送系统,为近年来药剂学领域颇为活跃的一系列超微小给药系统的统称。由于纳米粒比人体内最小的毛细管内径(4000nnm)和红血球(6000nm-9000nm)小得多,它可以在血中自由运动到达人体的各部位。药剂学中的纳米粒可分为两类：纳米载体药物和纳米药物。纳米载体药物为药物与药用材料一起形成的纳米给药系统,可通过载体提高药物的吸收,并可使其具有细胞亲和性、药物缓释性、靶向性,具有提高药物疗效、降低毒副作用的特征；纳米药物则是指直接将原料药加工成纳米粒,难溶性药物制备成纳米粒后,可有效提高药物的吸收,提高生物利用度,且和纳米载体相比,药物含量更高。根据脂质纳米载体药物较强细胞转运的特点,首先开展了龙血竭脂质纳米粒的研究。以硬脂酸为脂质材料,采用水性溶剂扩散法制备龙血竭脂质纳米粒；微粒粒度及表面电位仪测定其粒径和表面电位；透射电镜观察其形态；以龙血素A为指标,建立HPLC测定龙血竭脂质纳米粒中龙血竭含量的方法学；计算龙血竭脂质纳米粒的包封率和载药量。结果表明,本研究所制备的1 mg·ml-1龙血竭脂质纳米粒(药物与纳米载体比例为1：1)呈球形,数均粒径为29.4nm,电位为-20.2+1.9Mv；龙血素A于0.135-5.4μg·ml-1浓度范围内有良好线性关系,标准曲线方程为Y=33.694X+1.5898(r=0.9999)。所制备的1 mg·ml-1龙血竭脂质纳米粒中龙血素A的浓度为1.43μg·ml-1,平均包封率为89.59%。由于龙血竭中其有效成分龙血素A的含量较低,而一般脂质纳米粒的载药量仅为5%左右,所以我们针对龙血竭这类中药,开展了药物纳米粒研究,以提高纳米粒中的主药含量。采用水性溶剂扩散法制备龙血竭纳米粒；微粒粒度及表面电位仪测定其粒径和表面电位；透射电镜观察其形态；以龙血素A为指标,建立HPLC测定龙血竭纳米粒中龙血竭含量的方法学；计算龙血竭纳米粒载药量；以透析袋法对龙血竭纳米粒体进行外释放行为的研究。结果表明,本研究所制备的1 mg·ml-1龙血竭纳米粒呈球形,数均粒径为101.7nm,电位为-19.9±1.6mV；龙血素A于0.135.5.4μgvml-1浓度范围内有良好线性关系,标准曲线方程为Y=33.694X+1.589 8(r=0.9999),所制备的1 mg·ml-1龙血竭纳米粒中龙血素A的浓度为2.87μg·ml-1;体外释放行为的研究结果表明,与龙血竭原料药粉混悬液相比,龙血竭纳米粒分散液在72h中药物释放了约68%,而龙血竭原料药粉混悬液仅释放了约40%,促进释放的效果明显。在以上研究的基础上,我们进一步制备以龙血竭为主药的外用凝胶。以1%卡波姆为凝胶基质,制备龙血竭纳米粒凝胶、龙血竭粉凝胶、龙血竭醇溶液凝胶；考察其流动性,粘性及外观性能；以透析袋法对其进行体外释放行为的研究。结果表明,龙血竭纳米粒凝胶、龙血竭粉凝胶流动性小、粘性大,龙血竭醇溶液凝胶流动性小、粘性小；体外释放行为的研究中,相比较龙血竭粉所制备的凝胶,龙血竭纳米粒凝胶72h中药物释放可达约83%,而龙血竭粉所制备的凝胶72h中药物释放约为52%,龙血竭纳米粒凝胶促进释放的效果明显；而相比较龙血竭醇溶液制备的凝胶,龙血竭纳米粒凝胶在4h中药物释放了约55%,龙血竭醇溶液凝胶在4h中药物释放了约76%,龙血竭纳米粒凝胶则表现出缓释的效果。以大鼠背部剃毛区开方形全层皮肤切除创面的方法,建立动物皮肤损伤模型,以云南白药粉制备的凝胶为对照,通过肉眼观察、拍照、测量伤口大小,电子显微镜观察新生皮肤HE染色切片的方法,考察龙血竭纳米粒凝胶对皮肤损伤模型的促进愈合作用。结果表明,三种龙血竭凝胶制剂均能缩短愈合时间,其中龙血竭纳米粒凝胶促进促进皮肤愈合效果最显著。在实验初期(3d,6d),龙血竭醇溶液凝胶组愈合率较其他组显著高；但到实验后期(14d,20d),大鼠创面愈合速度进入减慢阶段后,龙血竭纳米粒凝胶组优势明显,20d时只有龙血竭纳米粒凝胶组创面完全愈合。创面新生皮肤切片经HE染色后观察,可见4组皮肤内均有丰富的毛细血管,说明新生皮肤为有组织活性的皮肤。
[Abstract]:Dracaena cochinchinensis (Lour.) S. C. Chen is a Liliaceous plant Dracaena cochinchinensis (Lour.) S. C. Chen resin processed, containing flavonoids, volatile oils and other components, has the function of activating blood circulation and dispersing stasis, relieving pain and stopping bleeding, astringent sores and muscle formation, Dracaena cochinchinensis (Lour.) Dracaena Nanoparticle drug delivery system (NDDS) is a kind of nanoparticle drug delivery system with a diameter of 1-1000 nm formed by drugs or drugs together with medicinal materials. It is an active series of ultramicro-particles in the field of pharmaceutics in recent years. Because nanoparticles are much smaller than the smallest capillary diameter (4000nnm) and red blood cells (6000nm-9000nm) in the body, they can move freely in the blood to various parts of the body. Pharmaceutical nanoparticles can be divided into two categories: nanocarrier drugs and nanodrugs. Nanocarrier drugs are drugs and medicinal materials together. The nano-drug delivery system can enhance the absorption of drugs through carriers, and make them have cell affinity, drug sustained release, targeting, with the characteristics of improving drug efficacy and reducing side effects; nano-drug refers to the direct processing of raw materials into nanoparticles, insoluble drugs into nanoparticles, can effectively improve the drug Lipid nanoparticles of Dragon's Blood were prepared by water-based solvent diffusion method with stearic acid as lipid material. The particle size and surface potential of Dragon's Blood Lipid Nanoparticles were determined; the morphology of Dragon's Blood Lipid Nanoparticles was observed by transmission electron microscopy; the content of Dragon's Blood Lipid Nanoparticles in Dragon's Blood Lipid Nanoparticles was determined by HPLC; the entrapment efficiency and drug loading of Dragon's Blood Lipid Nanoparticles were calculated. The carrier ratio was 1:1) spherical, the number average diameter was 29.4 nm, and the potential was - 20.2 + 1.9 Mv. There was a good linear relationship between the concentration of Loureirin A and the concentration of Loureirin A in the range of 0.135 - 5.4 ug. ML - 1. The standard curve equation was Y = 33.694 x + 1.5898 (r = 0.9999). The concentration of Loureirin A in the prepared nanoparticles was 1.43 ugh. ML - 1, and the average entrapment rate was 89.5 ugh. 9%. Because the content of Dragon's Blood Active Component Dragon's Blood A is low, and the drug loading capacity of the general lipid nanoparticles is only about 5%, we have carried out the research on the drug nanoparticles to improve the content of the main drug in the nanoparticles. The particle size and surface potential of Dragon's blood nanoparticles were measured by potentiometer; the morphology was observed by transmission electron microscopy; the content of Dragon's blood nanoparticles was determined by HPLC with Dragon's blood A as the index; the drug loading of Dragon's blood nanoparticles was calculated; the release behavior of Dragon's blood nanoparticles was studied by dialysis bag method. Dragon's blood A nanoparticles were spherical with an average diameter of 101.7 nm and a potential of - 19.9 (- 1.6 mV); there was a good linear relationship between the concentration of Dragon's blood A and the concentration of Dragon's blood A in the range of 0.135.5.4 ugvml-1. The standard curve equation was Y = 33.694 X + 1.589 8 (r = 0.9999), and the concentration of Dragon's blood A in the prepared nanoparticles was 2.87 UG (- ml-1); the release behavior in vitro was studied. The results showed that compared with the suspension of Dragon's blood powder, the drug release rate of Dragon's blood nanoparticles was about 68% at 72 h, while the suspension of Dragon's blood powder only released about 40% and the effect of promoting release was obvious. Dragon's blood nanoparticle gel, Dragon's blood powder gel and Dragon's blood alcohol solution gel were prepared with M as gel matrix, and their fluidity, viscosity and appearance properties were investigated. Compared with the gel prepared by dragon's blood powder, the drug release rate of dragon's blood nanoparticle gel reached about 83% after 72 hours, while that of dragon's blood powder gel reached about 52% after 72 hours, and the effect of dragon's blood nanoparticle gel on promoting drug release was obvious. The prepared gel, Dragon's blood nanoparticle gel and Dragon's blood alcohol solution gel were released about 55% and 76% respectively in 4 h and 4 h. Dragon's blood nanoparticle gel showed sustained release effect. The effect of dragon's blood nanoparticles gel on the healing of skin injury model was investigated by means of naked eye observation, photography, measurement of wound size and electron microscopy. The results showed that the three kinds of dragon's blood nanoparticles gel could shorten the healing time, and dragon's blood nanoparticles gel could promote the healing of skin injury model. At the beginning of the experiment (3d, 6d), the healing rate of Dragon's Blood Solution Gel group was significantly higher than that of other groups, but at the end of the experiment (14d, 20d), the healing rate of the wound in the Dragon's Blood Nanoparticle Gel group was slowed down. At the 20th day, only Dragon's Blood Nanoparticle Gel group healed completely. Skin sections stained with HE showed that there were abundant capillaries in the skin of the four groups, indicating that the newly formed skin was an organized and active skin.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R943

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