【作者】 黄敬洪；

【导师】 曹德英；

【作者基本信息】 河北医科大学 ， 药剂学， 2011， 硕士

【摘要】 目的:结肠癌为胃肠道中常见的恶性肿瘤,近年来结肠癌发病率呈明显上升趋势,对其诊治引起了各方关注。5-氟尿嘧啶为结肠癌治疗的首选化疗药物,但其口服吸收差,到达结肠的药物较少,且具有显著的不良反应及较大的毒副作用。口服结肠靶向释药系统（OCDDS）是通过适当的方法,避免药物口服后在胃、十二指肠、空肠和回肠端释放,运送至回盲部后再释药的靶向给药系统。将5-氟尿嘧啶制成pH-时间双依赖型口服结肠靶向微丸用于结肠癌的治疗,将5-氟尿嘧啶定位于结肠局部释放,不仅提高局部5-氟尿嘧啶浓度,同时降低其全身吸收和毒副作用。方法:采用微晶纤维素（MCC）为辅料,挤出滚圆法制备5-氟尿嘧啶载药丸芯,以含致孔剂（HPMC E5）的乙基纤维素水分散体为时间依赖层包衣材料,丙烯酸树脂S100（Eudragit S100）乙醇溶液为pH依赖层包衣材料,流化床底喷制备pH-时间双依赖型5-氟尿嘧啶结肠靶向微丸。以微丸的成球形（圆整度和收率）作为评价指标,挤出转速、滚圆转速、滚圆时间为工艺因素,分别选取3个不同水平,通过L₉（3³）正交试验确定制备丸芯的优化工艺。以进风温度、喷雾压力以及喷液速度为单因素考察流化床包衣工艺,确定包衣优化工艺。同时以释放度为指标,对致孔剂用量、时间依赖层和pH依赖层包衣增重进行单因素考察,确定了pH-时间双依赖型5-氟尿嘧啶结肠靶向微丸的优化工艺。采用高效液相色谱法（HPLC）对pH-时间双依赖型5-氟尿嘧啶结肠靶向微丸进行含量和释放度测定。色谱条件:色谱柱:Kromasil C₁₈,250 mm×4.6 mm, 5μm;流动相:0.01mol·L^-1磷酸二氢钾水溶液;柱温:25℃;流速:1.0 ml·min^-1;检测波长:265 nm;进样量:20μl。采用中国药典2005版二部附录XD第一法,转速100 r·min^-1,温度（37±0.5）℃,模拟人工胃液（pH1.2）2h后,换成人工小肠液（pH 6.8磷酸盐缓冲液）。6h（人工小肠液4h）后,再换成人工结肠液（pH 7.8-8.0磷酸盐缓冲液）至24h。在规定的时间点取样,在波长265nm处HPLC法测定吸光度,代入标准曲线计算药物浓度,求得累积释放度。考察高温、高湿和强光照射等因素对pH-时间双依赖型5-氟尿嘧啶结肠靶向微丸稳定性的影响,观察微丸的性状,测定含量和释放度,并进行加速试验。采用SD大鼠为试验动物,轻度乙醚麻醉,通过聚乙烯管给大鼠口服pH-时间双依赖型5-氟尿嘧啶结肠靶向微丸（试验组）,以口服5-氟尿嘧啶载药丸芯作为对照组,剂量均为30mg·kg^-1。采用HPLC法测定不同时间点血液、胃、小肠、盲肠及其内容物、结肠及其内容物的药物浓度。结果:根据综合评分,确定制备5-氟尿嘧啶载药丸芯的最优工艺为挤出转速30r·min^-1,滚圆转速40r·min^-1,滚圆时间4min。时间依赖层包衣工艺为:进风温度40℃,喷雾压力0.05MP,喷液速度1ml·min^-1。pH依赖层包衣工艺为:进风温度50℃,喷雾压力0.05MP,喷液速度1ml·min^-1。包衣处方通过单因素考察,确定致孔剂用量为聚合物用量5%,时间依赖层包衣增重为5%,pH依赖层包衣增重为55%。pH-时间双依赖型5-氟尿嘧啶结肠靶向微丸HPLC条件下保留时间为7.5 min左右,方法回收率为98.65%¹00.28%,日内精密度RSD（%）为0.86%¹.74%,日间精密度RSD（%）为0.92%¹.13%,符合中国药典标准。此方法能够准确测定包衣微丸中5-氟尿嘧啶的含量。相对湿度92.5%条件下放置5天后,吸湿增重为6.2%,大于5%。相对湿度75%、60℃高温、强光（4500lx）照射和加速试验后微丸外观、含量和释放度均无明显变化。大鼠口服给药后,pH-时间双依赖型5-氟尿嘧啶结肠靶向微丸（试验组）在胃和小肠几乎不释药,而于盲肠和结肠部位释放大部分药物;而未包衣微丸（对照组）在胃和小肠释放大部分的药物。试验组大鼠盲肠组织、盲肠内容物、结肠组织以及结肠内容物中5-氟尿嘧啶平均峰浓度分别为64.764±22.535μg·g^-1、618.949±201.641μg·g^-1、44.271±10.765μg·g^-1和458.147±93.821μg·g^-1。血药浓度相对低而平稳,达峰时间为12h,平均峰浓度为0.120±0.025μg·ml^-1;对照组平均峰浓度分别为4.537±1.548μg·g^-1、30.027±12.843μg·g^-1、1.824±0.615μg·g^-1和17.425±5.233μg·g^-1。血药浓度相对较高,达峰时间为1h,平均峰浓度为1.383±0.215μg·ml^-1。试验组和对照组的AUC值分别为1.042±0.132μg·h^-1·ml^-1和1.838±0.217μg·h^-1·ml^-1。试验组的相对生物利用度为56.69%。结论:pH-时间双依赖型5-氟尿嘧啶结肠靶向微丸具有良好的体外释药特征,重现性好;未包衣5-氟尿嘧啶微丸主要在大鼠的胃和小肠释药。而包衣微丸,药物在胃和小肠几乎不释放,定位于盲肠和结肠部位释放药物,不仅提高了5-氟尿嘧啶对结肠癌的治疗作用,也降低了其毒副作用。更多还原

【Abstract】 Objective: Colon cancer is common in gastrointestinal cancer. recently, the incidence of colon cancer is rising,so its treatment has attracted wide attention.5-Fluorouracil is one of the most widely used agents in the first-line chemotherapy of colon cancer.After oral administration, the absorption of the drug is poor,the drug which reach the colon is few.It also has serious adverse effects and toxic side effects.Oral colon-specific drug delivery system（OCDDS） is the appropriate ways to prevent drugs release in the stomach,duodenum,jejunum and ileum end,but effect an abrupt onset of drug release upon entry into the ileocecal.pH-time dependent pellets for colon-specific delivery of 5- fluorouracil release drugs located in the colon,not only improving the local therapeutic concentrations, but aslo reducing the systemic absorption and toxicity.Methods:Pellet cores containing 5-fluorouracil and microcrystalline cellulose were prepared by the process of extrusion-spheronization.The mixture of pore former（HPMC E5） and surelease was used as time dependent film coat.Eudragit S100 of ethanol solution was used as pH-dependent film coat.Coating was performed using fluidized bed coater.The optimization of technique was definite with the colligation evaluation of formation, the orthogonal experiment was designed to screen technique in which the speed of extrusion, speed of spheronization and the time of spheronization were taken as three influential factors and three different levels were selected to part, each conformation of technique was selected refer to the L₉（3³） orthogonal design table.The single factor experiment was designed to screen technique in the concentration of coating liquor ,pressure of spray ,temperature of in air.Single-factor analysis was carried out to find out the main factors（amount of pore former,weight gain of time dependent layer and pH dependent layer）affecting the drug release.HPLC method was established to determine the content and the release of 5-fluorouracil in the coated pellets.HPLC conditions:Chromatogram column:KromasilC₁₈（5μm,250mm×4.6mm）;mobilephase:0.01mol·L^-1 potassium dihydrogen phosphate buffer;column temperature:25℃; flow rate: 1ml·min ^-1;detection wavelength:265nm;injection volume:20μl.Drug release studies carried out using C.P2005 XC dissolution rate test apparatus（apparatus 1, 100rpm ,37℃）.Initial studies were carried out in 1000 ml of 0.1mol·L ^-1 HCl for 2h, then turn to pH6.8 phosphate buffer saline for 4h ,after this turn to pH7.8-8.0 phosphate buffer saline to 24h.At different time intervals, 5ml of the sample withdrawn and replaced with 5 ml of fresh dissolution medium.The samples were analysed by HPLC as described previously.The chemical and physical stability of pH-time dependent pellets for colon-specific delivery of 5-fluorouracil was investigated under the conditions of strong light ,high temperature and high humidity.Then the formulation was observed for change in physical appearance ,drug content and drug release rate.SD rats were administered orally uncoated and coated pellets via a polyethylence cannula under light ether anesthesia, respectively, at a dosage of 30mg·mg^-1.The 5-fluorouracil concentrarions in the gastrointestinal tract,cecum tissue and its contents,colon tissue and its contents,plasma were determined using HPLC method described previously.Results:The pellet core were prepared by the process of extrusion- spheronization.:speed of extrusion 30r·min^-1;speed of spheronization 40r·min^-1;time of spheronization 4 min.Time lag layer coating condition: temperature of in air 40℃,pressure of spray 0.05MPa,spray speed 1mL·min^-1. pH dependent layer coating condition:temperature of in air 50℃,pressure of spray 0.05MPa,spray speed 1ml·min^-1.The optimization of the colonic coating prescription was 5% HPMC E5 of solid content, 5% weight gain of time dependent layer and 55% weight gain of pH dependent layer .The reserve time of 5-fluorouracil was about 7.5min,the recoveries were 98.65¹00.28%, the within-day precision was 0.86¹.74%, the between-day precision was 0.92¹.13%.The method could determine the content of 5-fluorouracil accurately and precisely.Under the condition of RH92.5% for 5 days, the weight increasing ratio of pellets was 6.2%.There was no significant difference in the physical appearance and content and percent of release of the pellets at the end of the strong light,high humidity（75%） and the accelerated experiment.pH-time dependent pellets for colon-specific delivery of 5-fluorouracil （tested group）released little drug in the stomach and small intestine and most in cecum and colon tissues.But the uncoated pellets（controlled group）released most drug in the stomach and small intestine.The mean peak 5-fluorouracil concentrations in cecum tissue,cecal contents,colon tissue and colon contents,respectively,were 64.764±22.535μg·g^-1,618.949±201.641μg·g^-1, 44. 271±10.765μg·g^-1 and 458.147±93.821μg·g^-1 for 5-fluorouracil released from the coated pellets,and 4.537±1.548μg·g^-1,30.027±12.843μg·g^-1, 1.824±0.615μg·g^-1 and 17.425±5.233μg·g^-1 for the administration of the uncoated pellets.In plasma, the relatively low and flat drug concentration profile from coated pellets was in contrast to the high and sharp pharmacokinetic profile obtained from drug released from uncoated pellets.The mean Cmax from the coated pellets group (0.120±0.025μg·ml^-1) was lower than that of the uncoated pellets group (1.383±0.215μg·ml^-1).The mean Tmax of the coated pellets group（12h） was longer than that of the uncoated pellets group（1h）.The AUC values of coated pellets and uncoated pellets were 1.042±0.132μg·h^-1·ml^-1 and 1.838±0.217μg·h^-1·ml^-1.The relative bioavailability of the coated pellets from the uncoated pellets is 56.69%.Conclusions: pH-time dependent pellets for colon targeted delivery of 5- fluorouracil may provide the necessary protection to a drug in stomach and small intestine while allowing drug release in the colon.In contrast to uncoated pellets dispersal along stomach and small intestine, 5-fluorouracil was largely released from the the coated pellets in cecum and colon,not only improving the local therapeutic concentrations, but aslo reducing the systemic absorption and toxicity.更多还原