聚电解质修饰氧化石墨烯用于抗癌药物口服转运的研究

发布时间：2018-05-15 05:05

本文选题：氧化石墨烯 + 阿霉素　；参考：《辽宁大学》2017年硕士论文

【摘要】：氧化石墨烯(GO)是一种新型药物载体,GO近些年被广泛应用于静注药物的口服吸收载体,从而提高难溶性药物的生物利用度。本文通过采用改良的新方法制备GO,并利用电荷吸引的原理在其表面进行聚电解质的包裹修饰。研究聚电解质修饰GO用于抗癌药物口服转运的作用及其机制,并通过电镜表征,粒度分析,体外释放,药动学研究,MTT实验等多方面进行考察。本文采用改良后的Hummers法制得氧化石墨,经超声剥离后制得GO。采用扫描电子显微镜(SEM)、透射电子显微镜(TEM)、傅里叶红外光谱扫描仪(FTIR)、紫外光谱(UV)等对其形貌和化学结构进行表征。通过表征结果证实了制备的GO是纳米级的具有片层结构的载体。相较于以前的制备GO的方法,经我们改良之后不仅简化了实验操作步骤,也获得了安全性方面的提升。随后研究了其对小分子量的阿霉素(DOX)和大分子量的平阳霉素(PYM)的包载和体外释放行为。通过载药实验数据分析,GO对DOX的包封率达87.58%;对PYM的包封率达到82.05%。这是由于GO结构中含有更多的含氧官能团,所以很大程度上增加了GO的载药能力。对GO-DOX和GO-PYM的制剂组体外释放结果进行分析。制剂组在不同的pH条件下表现出不同的释放效果,两组的实验结果又表现了相同的释放趋势,且两组都于60h时在pH2.3的条件下达到最高的累计释放量。说明其释放行为表现出一定的pH依赖性,但是未表现出明显的缓释性。通过单因素考察法对聚丙烯酸(PAA)结合半胱氨酸(CYs)的制备工艺的最优方法进行了研究。通过Ellman法检测游离巯基含量,结果表明反应时加入的EDC含量、pH和温度等都会影响PAA与CYs的结合,影响高分子聚合物PAA-CYs上二硫键的含量。因此筛选出适宜的方法,使得PAA-CYs的合成产率相对提高。通过电荷吸引的方法将GO-DOX和GO-PYM与巯基聚电解质相结合并做了一系列的表征。将PAA-CYs/PAH-GO-DOX和PAA-CYs/PAH-GO-PYM进行了透射电镜的测定,结果呈现出相对稳定的结构。通过粒径考察,两种药物的制剂组的粒径分别为700nm与750nm左右。通过Zeta电位的考察可以看出Zeta电位为-39.8mV与-34.1mV,一方面表明出制剂的稳定性好,另一方面也证明制剂组带负电荷,与之前的理论分析相符合;之后又通过检测自由巯基可看出两种制剂组都不含有游离的巯基,可以从侧面的证实PAA与CYs结合完全。最后将DOX组,GO-DOX组和PAA-CYs/PAH-GO-DOX组与PYM组、GO-PYM组和PAA-CYs/PAH-GO-PYM组分别进行了体外释放考察,通过数据分析可以看出,制剂组全都表现出了极高的耐强酸稳定性,证明通过两层聚电解质的包裹,可使药物在强酸性环境得到保护;最后本课题进行了大鼠在体肠吸收实验、药动学研究以及MTT考察。在肠吸收实验中,从分析结果来看,PAA-CYs/PAH-GO-PYM制剂组相比于PYM组和GO-PYM组表现出了更好的肠道吸收效果。主要是由于表面有PAA-CYs的包裹,使PCPGP与肠道内壁相粘附,所以达到了更好的肠吸收效果。PCPGP制剂组在药动学研究方面,其AUC值较比于原料药提高65.57mg/L·h,并且T1/2也延长了0.519h,表现出一定的缓释效果。通过MTT法考察了制剂组的细胞活性研究。结果表明,在浓度20μM-100μM的条件下,PCPGP制剂组对于癌细胞A549的致死率相比于PYM组降低了2.40%。说明在24h内,PCPGP制剂组对细胞的杀伤效果没有PYM原料药组好。PCPGP制剂组的IC50值较之PYM组提高了24.28μM,较之GO-PYM组提高了29.42μM,说明PCPGP制剂组在相同浓度24h的时间内表现出了一定的缓释效果。最终说明PCPGP制剂组有效地提高了抗癌药物的口服生物利用度。
[Abstract]:Graphene oxide (GO) is a new drug carrier. In recent years, GO has been widely used as an oral carrier for intravenous drug injection so as to improve the bioavailability of insoluble drugs. In this paper, a modified new method was used to prepare GO, and the polyelectrolyte package modification was carried out on its surface by the principle of charge attraction. The effect and mechanism of GO used for oral transport of anticancer drugs was investigated by electron microscopy, particle size analysis, in vitro release, pharmacokinetics and MTT experiments. The modified Hummers method was used to obtain graphite oxide, and GO. was prepared by scanning electron microscope (SEM), transmission electron microscope (TEM), and Fu after ultrasonic peeling. The morphology and chemical structure of the inner leaf infrared spectrum scanner (FTIR) and the ultraviolet spectrum (UV) were characterized. The result of characterization proved that the prepared GO was a nanoscale carrier with lamellar structure. Compared with the previous method of preparing GO, we not only simplified the procedure of the experiment, but also obtained the security. The encapsulation and in vitro release of the small molecular weight of adriamycin (DOX) and the large molecular weight of Pingyang mycin (PYM) was investigated. The encapsulation efficiency of GO to DOX was 87.58% through the drug loading experiment data, and the encapsulation efficiency of PYM was 82.05%., which was due to the presence of more oxygen functional groups in the GO structure, so the GO was greatly increased. The release results of GO-DOX and GO-PYM in vitro were analyzed. The preparation group showed different release effects under different pH conditions. The results of the two groups showed the same release trend, and the two groups all reached the highest cumulative release under the conditions of pH2.3 at the time of 60H. A certain pH dependence, but not showing obvious slow release. The optimum method of the preparation process of polyacrylic acid (PAA) binding cysteine (CYs) was studied by single factor investigation. The content of free sulfhydryl group was detected by Ellman method. The results showed that the content of EDC, pH and temperature in the reaction would affect the combination of PAA and CYs. The content of the two sulfur bond on the polymer PAA-CYs is rounded. Therefore, a suitable method is selected to make the synthesis yield of PAA-CYs relatively higher. A series of characterization of GO-DOX and GO-PYM with sulfhydryl polyelectrolytes is made by the method of charge attraction. The transmission electron microscopy of PAA-CYs/PAH-GO-DOX and PAA-CYs/PAH-GO-PYM has been measured. The results showed a relatively stable structure. The particle size of the preparation group of the two drugs was 700nm and 750nm respectively through the particle size. The Zeta potential was -39.8mV and -34.1mV through the Zeta potential investigation. On the one hand, the stability of the preparation was good, on the other hand, the negative charge of the preparation group was proved to be in conformity with the previous theoretical analysis. After the detection of free sulfhydryl group, we can see that all two kinds of preparation groups do not contain free sulfhydryl groups. It can be confirmed from the side of the combination of PAA and CYs. Finally, the release of DOX, GO-DOX and PAA-CYs/PAH-GO-DOX group with PYM group, GO-PYM group and PAA-CYs/PAH-GO-PYM group were examined respectively. All the groups showed high resistance to strong acid stability. It was proved that the drug could be protected by two layers of polyelectrolytes in the strong acid environment. Finally, the rat intestinal absorption experiment, pharmacokinetic study and MTT investigation were carried out in the rat. In the intestinal absorption experiment, the PAA-CYs/PAH-GO-PYM preparation group was compared to PYM The group and the GO-PYM group showed a better intestinal absorption effect, mainly due to the PAA-CYs parcel on the surface, which made the PCPGP adhere to the inner wall of the intestines, so it reached a better intestinal absorption effect, so the.PCPGP preparation group had a higher AUC value than the 65.57mg/ L. H compared to the API, and T1/2 also extended 0.519h. The cell activity of the preparation group was investigated by MTT method. The results showed that, under the condition of concentration of 20 mu M-100 M, the lethal rate of A549 in the PCPGP preparation group was lower than that of the PYM group. The killing effect of the PCPGP preparation group on the cells was no longer than the IC50 value of the PYM raw material group and the.PCPGP preparation group. The PYM group increased 24.28 mu M and increased 29.42 M compared with the GO-PYM group, indicating that the PCPGP preparation group showed a certain slow release effect in the time of the same concentration of 24h. Finally, the PCPGP preparation group effectively improved the oral bioavailability of the anticancer drug.

【学位授予单位】：辽宁大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R943

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