N-乙酰半胱氨酸活性炭缓释微囊的制备及其抗大鼠肝纤维化作用的研究

发布时间：2018-02-23 17:34

本文关键词： N-乙酰半胱氨酸活性炭缓释微囊口服肝纤维化　出处：《浙江大学》2017年硕士论文　论文类型：学位论文

【摘要】：本研究利用乳化交联法制备N-乙酰半胱氨酸活性炭缓释微囊(NCSM),优化制备工艺,考察其制剂学和药代动力学参数,进一步研究微囊口服给药对大鼠肝纤维化的治疗作用,并与N-乙酰半胱氨酸溶液和市售经典抗肝纤维化药物水飞蓟宾口服给药进行比较。目的：N-乙酰半胱氨酸(NAC)作为谷胱甘肽的前体,其抗氧化性对肝纤维化病变的治疗作用已日渐受到关注,目前临床给药途径为静脉注射,但因其在体内的快速代谢,无法形成稳定持久的血药浓度,且临床注射给药依从性较差。N-乙酰半胱氨酸活性炭缓释微囊通过消化道的缓释作用,形成较为稳定的血药浓度,延长药物作用时间,开辟N-乙酰半胱氨酸口服治疗肝纤维化给药途径,增加患者依从性。方法：①处方前研究。建立N-乙酰半胱氨酸高效液相色谱测定方法,制备活性炭载药体,选择明胶为囊材,(去除了聚丙烯酸树脂Ⅱ/Ⅲ)采用乳化交联法建立N-乙酰半胱氨酸活性炭缓释微囊制备方法。②优化N-乙酰半胱氨酸活性炭缓释微囊制备工艺。利用正交试验L9(34),考察药载比、明胶浓度、搅拌速度和乳化剂浓度4因素对活性炭载药囊芯的载药量、包封率和粒径分布的影响,得出最佳微囊制备工艺,评价微囊质量。③考察N-乙酰半胱氨酸活性炭缓释微囊体外释放情况及体内血药浓度变化,评价NCSM缓释效果。④考察NCSM口服给药对四氯化碳(CC14)所致大鼠肝纤维化的治疗作用。建立CCl4致大鼠肝纤维化模型,以NCSM高中低三个剂量：80mg·kg-1·D-1、40mg·kg-1·D-1、20mg·kg-1·D-1(按照乙酰半胱氨酸计算),NAC、水飞蓟宾作为阳性对照药物,进行灌胃给药。治疗8周后处死大鼠,比较其不同给药剂量与NAC和水飞蓟宾对大鼠肝纤维化的治疗作用,探讨其对肝纤维化大鼠肝功能指标、肝纤维化病理学及大鼠肝纤维化成纤细胞信号通路(TGF-β1/TβR-Ⅰ/Smad2/3)的影响,评价口服NCSM对肝纤维化模型大鼠的治疗效果。结果：①通过试制,确定成囊效果较好的明胶作为囊材进行N-乙酰半胱氨酸活性炭缓释微囊的制备工艺的研究。②正交试验L9(34)结果表明,成囊效果影响由大到小依次为药载比(药物与明胶的比例)搅拌速度明胶浓度乳化剂浓度。药载比为1：1,明胶浓度为15%,搅拌速度为1000 r·min-1和乳化剂(Span-80)浓度为2%的条件下,制备的NCSM粒径均匀,载药量,包封率高。验证试验结果表明,该工艺成囊质量稳定,确认为N-乙酰半胱氨酸活性炭缓释微囊的最佳制备工艺。制备三批NCSM,平均粒径为110.4μm±13.59μm,粒径分布在80-140 μm的微囊占总数81.9%,平均包封率为78.1%,平均载药量为15.9%。③体外释放结果表明,NCSM在1h内累计释放为20.4%,24h为69.2%,符合一级动力学方程。小鼠体内药代动力学结果表明,口服微囊的t1/2为NAC的3.32倍,AUC(o-24)为NAC1.18倍,具有明显缓释作用。④CCl4致大鼠肝纤维化模型制备成功,其大鼠肝组织病理切片HE染色和网状纤维染色表明其肝纤维化程度为Ⅲ-Ⅳ期,免疫组化显示胞内TGF-β1/TβR-Ⅰ和Smad2/3大量表达。NCSM各治疗组大鼠肝组织病理切片表明肝纤维化好转,免疫组化显示胞内TGF-β1/TβR-Ⅰ和Smad2/3表达减少,NCSM高剂量组(80mg·kg-1·D-1)表达具显著性差异(p0.05)。结论：采用乳化交联法制备的N-乙酰半胱氨酸活性炭缓释微囊,粒径均匀,载药量和包封率稳定。通过口服给药途径,与NAC比较,血药浓度维持时间显著增加,生物利用度显著提高,达到缓释作用。NCSM对大鼠肝纤维化有较好地治疗作用,且较等剂量的NAC及对照药物水飞蓟宾有更显著的疗效,其作用机制可能与抑制大鼠肝纤维化胞内成纤信号分子TGF-β/TβR-1和Smad2/3有关。
[Abstract]:The study on Preparation of activated carbon N- acetylcysteine microcapsules by emulsion crosslinking method (NCSM), to optimize the preparation process, the preparation and pharmacokinetic parameters, further study on microcapsule oral administration on hepatic fibrosis in rats and the therapeutic effect of N- acetylcysteine solution and commercially available classic anti fibrosis drug silymarin Philippines oral administration were compared. Objective: N- acetylcysteine (NAC) as a precursor for glutathione and its antioxidant role in treatment of liver fibrosis has been increasingly concerned, the current clinical administration for intravenous injection, but because of its rapid metabolism in vivo, unable to form a stable blood concentration and lasting. Clinical injection poor adherence of.N- acetylcysteine activated carbon microcapsules by sustained release of digestive function, forming a relatively stable plasmaconcentration, prolong drug action time, open N- B Cysteine oral treatment of liver fibrosis and route of administration, increase the compliance of patients. Methods: the study before prescription. To establish a method for determination of N- acetylcysteine, HPLC, preparation of activated carbon loaded, gelatin was selected as the wall materials (removal of polyacrylic resin II / III) by emulsion crosslinking method to establish the N- acetyl cysteine activated carbon release microcapsule preparation method. The optimization of N- acetylcysteine activated carbon microcapsules preparation. By orthogonal test L9 (34), the investigation of drug loading ratio, gelatin concentration, stirring speed and loading concentration of emulsifier 4 factors on activated carbon loaded bag core, influence the encapsulation efficiency and particle size distribution the optimum preparation process of microcapsules, and evaluate the quality of microcapsule. The effects and changes of blood concentration in vivo release of N- acetylcysteine activated carbon microcapsules in vitro release effect. The evaluation of NCSM effects of NCSM given orally for four Carbon tetrachloride (CC14) treatment induced liver fibrosis in rats. Establishing a rat model of hepatic fibrosis induced by CCl4 in NCSM three intervendonal: 80mg - kg-1 - D-1,40mg - kg-1 - D-1,20mg - kg-1 - D-1 (calculated by N-acetylcysteine), NAC, silybin was used as positive control drug, respectively intragastric administration. After 8 weeks of treatment, the rats were sacrificed to compare the therapeutic effect and NAC dosage and silymarin on liver fibrosis in rats, to explore the index of liver function in rats with liver fibrosis, and liver fibrosis rat fibroblast signaling pathway in liver fibrosis (TGF- beta 1/T beta R- I /Smad2/3) effect, to evaluate the therapeutic effect of oral NCSM on liver fibrosis model rats. Results: 1. Through trial research to determine the preparation process of microcapsules better effect of gelatin as capsule material of N- acetylcysteine activated carbon microcapsules. The orthogonal test of L9 (34). The results show that the sacculation effect from high to low is drug load ratio (the ratio of drug and gelatin) stirring speed of gelatin concentration of emulsifier concentration. Drug loading ratio was 1:1, the concentration of gelatin was 15%, stirring speed was 1000 R - min-1 and emulsifier (Span-80) concentration is 2%, the preparation of NCSM uniform particle size, drug loading, encapsulation rate was high. The testing results show that the process of sacculation stable quality and confirm the optimal conditions for the preparation of activated carbon N- acetylcysteine microcapsules. The preparation of the three batch of NCSM, the average particle size of 110.4 m + 13.59 m, accounting for 81.9% of the total number in particle size distribution was 80-140 m, the average entrapment rate was 78.1%, the average drug loading was 15.9%., the in vitro release results showed that NCSM in 1H cumulative release was 20.4%, 24h was 69.2%, with a kinetic equation. In vivo pharmacokinetic results showed that oral administration of t1/2 microcapsules was 3.32 times NAC, AUC (o-24 NAC1) .18 times, has obvious sustained-release effect. The CCl4 induced rat liver fibrosis model was successfully prepared, the rat liver tissue HE staining and reticular fiber staining showed that the liver fibrosis degree III - IV period, immunohistochemistry showed that intracellular TGF- beta 1/T beta R- I and Smad2/3 expression of.NCSM in the treatment of liver group the pathological sections showed that liver fibrosis improved, immunohistochemistry showed decreased expression of TGF- beta 1/T beta R- I and Smad2/3 cells in high dose group of NCSM (80mg kg-1 D-1) expressed significant difference (P0.05). Conclusion: using emulsion crosslinking method to prepare N- acetyl cysteine activated carbon microcapsules, uniform particle size, entrapment efficiency and drug loading stability. Through the oral route of administration, compared with NAC, significantly increased the blood concentration of maintenance time, improve the bioavailability, achieve sustained release effect.NCSM has better therapeutic effect on liver fibrosis in rats, and the dose The NAC and the control drug silybin had a more significant effect. The mechanism may be related to inhibiting the intracellular fibroblast signaling molecules TGF-, /T, beta R-1 and Smad2/3 in rat liver fibrosis.

【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R943;R96

【相似文献】