摘要 脉冲给药系统(pulsed drug delivery system, PDDS)能克服传统给药模式(如片 剂、缓控释制剂)在治疗某些依据时辰生理发生变化的疾病时,复杂的给药方式造 成患者顺应性低的缺点,而根据时辰生物学和时辰药动学特点,以制剂手段使药 物在体内经过一个时滞期后在疾病发作时快速完全地释药,达到最佳的疗效。本 课题根据进餐前后肠道内胰脂肪酶浓度相差4-10倍以上的生理情况,探讨利用胰 脂肪酶和脂质成分相互作用而引起局部pH变化来设计pH敏感的新型口服脉冲制 剂,因应进餐前后血糖的变化,反馈肠道中胰脂肪酶的释放,自动调节释药速率, 在餐后加快释放降糖药物,增强降糖治疗效果,提高患者服药的顺应性。 为了研究这种新型脉冲制剂的可行性,需要研究常用脂质辅料在不同浓度的胰 脂肪酶作用下脂解后体系的pH变化规律,筛选在不同浓度胰脂肪酶作用下脂解后 体系pH差异显著的脂质辅料。这种辅料能在进食后胰脂肪酶显著增加时快速脂解 而显著降低局部pH诱发药物的释放。因此,本课题考察在进食前后不同胰脂肪酶 浓度下各种纯甘油酯的脂解行为,研究油脂类型与脂肪酸释放量、体系pH变化的 规律。此外,选取合适的pH敏感高分子材料作为药物载体是设计和制备这种智能 给药系统的另一关键。因此,本课题的主要研究工作有: (1)脂质辅料脂解规律研究:禁食/进食情况下胰脂肪酶浓度差异显著。利用体 外脂解模型,模拟进食前后的胰脂肪酶浓度,对常用脂质辅料进行脂解,结果显 示在进食后高浓度的胰脂肪酶作用下能产生更多的脂肪酸,使体系pH较禁食条件 下有显著降低。采用体外脂解模型,分别在禁食/进食条件下,对不同链长和不同 酯键数的纯甘油酯进行脂解,探索pH的变化规律,筛选出适合新型pH敏感脉冲 制剂的脂质辅料。 (2) pH敏感纳米粒的制备和表征:化学合成pH敏感的聚合物——聚13-氨基酯, 并采用核磁共振(nuclear magnetic resonance, NMR)、凝胶渗透色谱(gel permeation chromatography, GPC)对合成的聚合物进行验证,结果显示成功合成了聚义-氨基 酯。以格列吡嗪为模型药,利用自乳化溶剂扩散法制备聚13-氨基酯载药纳米粒, 采用Box-Behnken设计优化纳米粒的制备处方,结果显示优化的处方与实际检测 西南大学硕士学位报告 值接近,优化结果可靠,制备的纳米粒粒径225.6nm,多分散指数为0.206,Zeta 电位为3.27mV。采用透析法分别测定pH 5.0和pH 7.8环境下纳米粒的体外释药, 结果发现在pH 5.0时释药速率和累积释药量均大于在pH 7.8,表明聚J3-氨基酯在 pH < 7的环境下释药较快速,所合成的pH敏感高分子材料达到了预期的设定目标。 本课题是探讨胰脂肪酶触发的新型脉冲制剂可行性的基础研究,对脂质辅料的 脂解规律进行了初步探索,发现由于进食后的胰脂肪酶浓度远远高于禁食状态, 导致进食后脂质辅料脂解后产生大量脂肪酸,显著降低体系局部pH。而且脂质辅 料的甘油酯链长越短、酯键数目越多,体系的pH降低越多,对药物的释放越有利。 在此基础上,后续研究需要进一步确定最适合的脂质辅料种类及用量。本课题合 成了适宜的pH敏感高分子材料——聚J3-氨基酯,制备了载格列吡嗪的纳米粒,并 考察了其体外释药性质。下一步要将此纳米粒混悬于脂质辅料中,最终制成pH敏 感的纳米粒脂质混悬液——胰脂肪酶触发的脉冲制剂,研究其释药行为和体内药 动学特性。 关键词:脉冲制剂;脂解;pH敏感;体外脂解模型;纳米粒;聚13-氨基酯; 格列吡嗪 II ABSTRCT ABSTRACT Pulsed drug delivery systems (PDDS) could overcome the disadvantages of traditional formulations,which have low compliance resulting from complicated administration. Based on chronobiological and chronopharmacokinetic characteristics, PDDS are designed to release drug quickly and completely after a lag-time when necessary. In this issue,the feasibility of a novel oral pH-sensitive pulsed formulation was investigated. The formulation was established to improve the inconveniences and compliance of oral hypoglycemic drags according to high level of pancreatic lipase after meal enhancing lipolysis on lipid excipients which would result in the increase of lipid products, such as fatty acids,and the change of pH in the intestinal microenvironment. In order to study the feasibility of this novel pulsed formulation,it is necessary to investigate pH change during lipolysis of lipid excipients catalyzed by different concentration of pancreatic lipase,and then to select the appropriate lipid excipient which could stimulate drag release quickly by lipolysis under the condition of high level pancreatic lipase in fed state. This paper aims to investigate the relationship among lipid type, the release amount of fatty acids and pH change. Moreover,it is another key to choose a pH responsive polymer as drag carrier for this intelligent drug delivery system. Consequently, in this paper, we investigate as follows: (1) Lipolysis pattern of lipid excipient: it was found that the concentration of pancreatic lipase was significantly different between fasted and fed state. In vitro lipolysis model was used to determine title lipolysis of common lipid excipients. The results showed that more fatty acids were produced under the condition of higher pancreatic lipase in fed state resulting in the significant decrease of environmental pH. In the same way, lipolysis on pure glycerides with different chain length and number of ester bonds were investigated in fasted and fed state. The results would help to choose the appropriate lipid excipient for novel pH-sensitive pulsed formulation. (2) The synthesis and characterization of poly (fi-amino esters): poly (fi-amino esters) 西南大学硕士学位报告 was synthesized in laboratory and determined by using NMR and GPC methods. The results showed that the desired poly (6-amino esters) was synthesized successfully. Glipizide was as a model drug and the drug-loaded nanoparticles were prepared by self-emulsifying solvent diffusion method. The formulation was optimized by Box-Behnken design and results showed that the average particle size of nanoparticles was 225.6nm,the polydispersity index (PDI) was 0.206 and the zeta potential was 3.27mV. The in vitro drug release characteristics of nanoparticles were investigated by dialysis method at pH 5.0 and pH 7.8 respectively and results showed that at pH 5.0 the drug was released faster than that at pH 7.8. It suggested that poly (6-amino esters) was dissolved quickly when pH < 7 and could be solved more easily in acidic environment. This subject mainly involved the evaluation of the feasibility of the novel pancreatic lipase stimulated pulsed formulation. In present work,the change patterns of environmental pH were studied and the results showed that more fatty acids were produced from lipolysis since higher concentration of pancreatic lipase in fed state, resulting in environment pH significantly decreased. The lipid excipioit with shorter chain length and more ester bonds would be catalyzed fast and produced more fatty acids to form acidic environment, which was benefit to drug release. In future, detailed study on lipid excipients such as its type and dosage should be carried out. On the other hand,a pH-responsive polymer as drug carrier for the intelligent drug delivery system, poly (B-amino esters) was synthesized, drag-loaded nanoparticles were prepared and drag release characteristics in vitro were also investigated. In future, the pancreatic lipase stimulated pulsed formulation should be prepared by suspending pH-sensitive nanoparticles in lipid carrier suspension and drug release behavior and pharmacokinetic characteristics should be investigated. Key Words: pulsed drug delivery system; lipolysis; pH-sensitive pulsed formulation; in vitro lipolysis model; nanoparticle; poly (J3-amino esters) ; glipizide IV 文献综述 第1章文献综述 ‘ 根据生物药剂学分类系统(biopharmaceutical classification system, BCS)⑴分类, 典型的BCS II和BCS IV类药物往往在体内吸收速率慢、吸收量较少,其原因在于 这些药物在体内的溶解、膜渗透情况不理想。水难溶性药物(poorlywater-soluble drugs,PWSD) [2]在胃肠道内一般以晶态形式存在,为了提高此类药物的口服生物 利用度,常用的药剂学方法一方面是减少晶体药物的粒子大小,另一方面釆用合 适的处方设计使药物在肠内处于稳定的溶液状态,即制成无定形制剂或脂质制剂。 脂解是指脂类物质在酶消化作用下分解产生游离脂肪酸和甘油。从脂解作用被 发现至今,脂解已经广泛应用于食品、工业生产、研究方面主要着重于生物工程 技术、生物化学[3,4]等领域,其在药学领域特别是药剂、药物开发领域中的研究和 应用带来了更多的机遇和挑战。 1.1体内脂解过程 胃肠道的脂肪酶主要有唾液脂肪酶、胃脂肪酶、胰脂肪酶。其中唾液脂肪酶由 唾液腺分泌,;pH4酶活性最大,作用于甘油三醋的第3位酯键。胃脂肪酶由胃粘 膜细胞分泌,pH 36酶活性较大,作用于甘油三醋的第1、3位酯键。胰脂肪酶 由胰腺分泌,主要存在于小肠部