Delayed release drug delivery system (DDS) is a chronopharmacology and hominal physiology-based DDS, which is mainly applied for the treatment of diseases exhibiting circadian rhythms, such as ischemic heart disease and asthma, and intestinal diseases. As a multiparticulate DDS, delayed release pellets are receiving increasing attention from researchers. Double membranes system was always utilized for preparing time controlled delayed release pellets. However, there are some limitations of double membranes system, such as sole using swelling material cannot form a compact film, where a loose and porous membrane will be formed. Therefore, there is a potential risk that the water-soluble drug with low molecule weight would dissolve into the water and penetrate into the pellets through pores. In this study, above problem was solved by using swelling agent mixed with hydrophobic material as swelling layer, and its pharmacokinetics was evaluated in vivo. Danshensu is selected as the model drug, which is a small hydrophilic active compound isolated from the commonly used Chinese herb, Salvia miltiorrhiza. Its molecular weight is 198 and it can effectively cure heart diseases through multi-mechanisms, such as coronary heart disease, angina and myocardial infarction. In our previous study, the delayed release pellets which have double membranes have been established in our previous work. These double membranes are composed by swelling layer and control layer, and the swelling layer is composed by swelling agent and hydrophobic material. After screening, cross-linked carboxymethyl cellulose sodium (CC-Na) was chosen as swelling layer and Surelease as hydrophobic material. Based on this work, we increased the drug loading, characterized the mechanism of the swelling layer, evaluated the behavior of the drug in vivo, scaled-up the preparation process and applied the techniques on Danshen extraction. To carry out the in-vivo experiment, drug loading of Danshen pellets needs to be increased. Firstly, blank pellet cores were coated by Hydroxypropyl methylcellulose (HPMC) solution where sodium Danshensu was dissolved in through fluid bed coating. Secondly, drug layer was sealed by HPMC coating to isolate drug from double membranes. Thirdly, CC-Na and Surelease were mixed at a weight ratio of 1:5 and coated onto the pellets with 120% weight gain. Finally, controlled layer composed by Surelease was coated at a weight gain of 20%. Results showed the drug loading was increased from 4.1 mg/g to 12.18 mg/g, and the dissolution test indicated that tlag, tmax, and the overall drug release rate of Danshen of coated pellets were 3.50 h, 7.00 h, and 93.2%, respectively. In the stability test, the pellets still demonstrated 3 h time lag in drug release profiles after being stored in dry condition at room temperature for three months, suggesting that coated pellets are stable during short-term storage. Since blending hydrophobic material with swelling agent as swelling layer is a new approach to prepare delayed release pellets, more mechanistic investigations including the effects of Surelease in the swelling layer needs to be elaborated. The surface and cross-section of pellets were examined by scanning electron microscopy. It was found that adding Surelease would make the film more condense and uniform, which will decrease water-vapor permeability and prevent water to pass thought the film. Results also showed that the swelling degree of CC-Na will be stronger if the pH of the environment solvent becomes higher. After in-vitro evaluation, the pharmacokinetic behavior of coated pellets was evaluated in vivo. Firstly, a method to quantification Danshensu in the Beagle dog’s plasma was established using HPLC-DAD. The methodology validation demonstrated that this method is sensitive, accurate and reliable for in-vivo quantification of Danshensu. After oral administration of Danshensu uncoated pellets and delayed release pellets to beagle dogs, the tlag and tmax were increased from 0.00 h to 1.50 h and 1.08 h to 3.33 h, respectively. MRT was extended from 1.50 h to 6.36 h. No significant difference was observed for AUC (p>0.05) although the mean value was reduced from 5.22 μg*h/mL to 3.91μg*h/mL. These results indicated that these coated pellets could achieve drug delayed release and maintain the similar absorption amount in vivo. Since Chinese Medicines are generally administrated as a mixture of multiple herbs, above formulation strategy was further utilized on Danshen extraction in the following study. Firstly, the extraction condition of Danshen was optimized. Results showed that extraction yield increased 6-7 times under alkaline condition (i.c. pH 8.2) compared to that at neutral condition. Above extraction solution was dried and coated onto the blank pellet by the same method mentioned above. Using Danshensu and Ginsenoside Rb1 as markers, dissolution study demonstrated that tlag were maintained at ~4 h but the speed of drug release was reduced with tmax higher than 10 h compared to that of coated pellets with pure danshensu. Finally, the scale-up experiments were carried out. Result showed that the drug loading ratios of the pellets were from 3.48 mg/g to 3.91 mg/g after scale-up. When the weight gain of the controlled layer was 15%, tlag, tmax and the final drug release rate were 3.5 h, 11 h, and 81.7%, respectively. Overall, this study demonstrated that the delayed release system which contains Surelease and CC-Na as swelling layer could achieve the delayed release for water-soluble active compounds with small molecular weight in both in-vitro and in-vivo experiments.

遲釋給藥系統是根據時辰藥理學和人體生理學設計的新型給藥系統，主要 應用於心血管疾病、哮喘等具有時辰發作規律疾病的治療,也可以作為定位釋 藥系統來治療腸道疾病。作為多微粒給藥系統，遲釋微丸是遲釋給藥系統的研 究熱點之一，其中雙層膜時控遲釋系統經常被採用, 但是該方法有一定的局限 性,即單獨使用水不溶性溶脹材料作為溶脹層，所形成的溶脹層結構疏鬆多孔， 在控釋層被撐破之前，小分子藥物容易順著溶脹層的空隙擴散出微丸，造成藥 物釋放的時滯變短。本研究通過將溶脹材料與疏水性材料聯用作為溶脹層，有 效地解決了這一問題。本實驗的模型藥物，丹參素是常用活血化瘀中藥丹參中 的小分子水溶性活性成分之一，分子量為 198，能夠通過多種機制治療心血管 疾病，例如冠心病、心絞痛及心肌梗死。 本研究之前，本課題組已經建立了雙層膜遲釋微丸的製備工藝。雙層膜由 溶脹層與控釋層組成，而溶脹層則由溶脹材料與疏水性材料混合組成。經過篩 選，溶脹材料選用交聯羧甲基纖維素鈉（CC-Na），疏水性材料則為乙基纖維 素的水分散體 Surelease。而本文在已有的研究基礎上，對該遲釋微丸進行了載 藥量的提高、溶脹層的機理探討、藥物的體內動力學評價、流化床包衣工藝的 放大以及本技術對於中藥複方的應用。 由於需要對微丸進行體內的藥物動力學評價，為了順利的進行動物試驗， 本研究首先採用流化床法製備了具有更高載藥量的丹參素雙層膜遲釋微丸。工 藝包括丹參素原料藥通過溶於羥丙甲基纖維素（HPMC）溶液後，包於蔗糖空 白丸芯上，並且利用 HPMC 將載藥層與雙層膜遲釋系統進行隔離。CC-Na 與 Surelease 按照 1:5 的比例進行混合，進行增重為 120%的溶脹層包衣；而控釋 層的材料為 Surelease，包衣增重為 20%。載藥量由之前的 4.1 mg/g，提高至 12.18 mg/g，體外溶出試驗證明該微丸具有 3.50 h 的釋藥時滯（tlag），tmax=7.00 h 最 終釋藥率 R 為 93.2%。而在穩定性考察中，該微丸經過 3 個月的乾燥室溫儲藏 後，依然能夠保持 3.00 h 的釋藥時滯，說明該雙層膜遲釋微丸在該期間內可以 穩定儲存。 由於採用溶脹材料與疏水性材料混合後作為溶脹層材料，作為一種全新的 理念，有必要研究 Surelease 在溶脹層中所起到的作用。採用 SEM 對為微丸的橫切面進行觀察，可以明顯的觀察到加入 Surelease 對於溶脹層衣膜物理結構 上的影響，相比於沒有加入 Surelease 的溶脹層，加入 Surelease 后可以使衣膜 更為均勻；而水汽通透性試驗則說明了加入 Surelease 能夠更為有效的阻擋水 分的滲透。而 CC-Na 作為溶脹材料，其溶脹能力對於 pH 的變化也很敏感。經 過試驗證明，其溶脹度的能力會隨著環境溶液 pH 的增大而增大。 完成體外評價後，我們對微丸進行了體內的藥物動力學評價。本研究的體 內評價部分採用比格犬進行。首先建立了 HPLC-DAD 分析方法測定比格犬血 漿中的丹參素的濃度。方法學考察證明，該方法靈敏度高，操作方便，可快速、 準確地測定比格犬血漿中丹參素的含量。比格犬口服丹參素遲釋微丸後，丹參 素的釋藥時滯從 0.00 h 增加到了 1.50 h， tmax從 1.08 h 增加到了 3.33 h，MRT 從 1.50 h 延長到了 6.36 h，而 AUC 雖然相對略有減小，但是並沒有顯著性差 異（p>0.05）。說明該遲釋微丸在體內也可以達到一定的釋藥時滯，並且藥物 吸收量並沒有明顯降低。 因為傳統中藥的給藥往往是以複方的形式來進行的，所以本研究繼續考察 了在以丹參三七複方提取物為載藥層時，丹參素能否還以遲釋的狀態進行釋藥， 同時觀察其他水溶性成分的釋藥曲線。首先以丹參素為考察指標，優化了丹參 三七水提取溶液的 pH 值。當提取溶液的 pH 值為 8.2 時，丹參素的提取率為普 通水提法的 6-7 倍。採用真空旋轉蒸發以及冷凍乾燥等方法製備了丹參三七水 提取物粉末，並按照上述流化床包衣法將其製備成遲釋微丸。體外溶出實驗證 明，當以丹參素和人參皂苷 Rb1 為考察指標時，該遲釋微丸具有 4 h 左右的釋 藥時滯。但是與丹參素純品的釋藥曲線相比，釋藥速率較為緩慢，tmax 均在 10.00 h 以上。 本文最後研究了該遲釋微丸的製備規模從 50 g 放大到 500 g 後的製備工藝， 對控釋層進行了重新調整並對微丸進行了體外評價。結果表明經過製備規模放 大後的微丸，具有 3.48 mg/g - 3.91 mg/g 的載藥量。當 Surelease 控釋層的增重 為 15%時，釋藥曲線較為理想，tlag=3.50 h，tmax=11.00 h，最終釋藥率 R=81.7%。 本研究證明，將Surelease與CC-Na混合後作為溶脹層的雙層膜遲釋微丸， 在體外與體內均可以對水溶性小分子的中藥活性成分丹參素進行遲釋釋藥。並 且將該遲釋微丸大規模製備後，或者採用中藥複方提取物來進行試驗依然可以 達到遲釋效果。關鍵字：丹參素，遲釋微丸，交聯羧甲基纖維素鈉（CC-Na），乙基纖維素水分 散體（Surelease），流化床，比格犬，丹參三七提取物