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UM E-Theses Collection (澳門大學電子學位論文庫)

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Title

野黃芩素納米混懸液製備及其作為野黃芩苷活性前體的體內研究 / Formulation development of scutellarein nanosuspensions as an in-vivo active, rapidly absorbed precursor of its glycoside scutellarin / by Xiao Yang.

English Abstract

The aims of this study were to synthesize the scutellarin (SG) aglycone scutellarein (S) by an acidic hydrolysis, and to formulate and characterize the S nanosuspension for improved dissolution in vitro and enhanced reduction effect for cerebral hemorrhage on zebrafish in vivo. An optimized successive 4 steps preparation of S powder with high chemical purity (HPLC purity>95%) has been established including hydrolysis, crystallization, concentrate and lyophilization. Hydrolysis reaction took 2 hours and the yield was 52.5% (theoretical value is 61.9% with completely reaction). S nanosuspension containing poloxamer 188 (F68) as stabilizer was prepared by anti-solvent precipitation approach and then dried S nanoparticles with D-trehalose as lyophilizing protectant were prepared with fine re-dispersibility. The average particle size, PdI and Zeta potential of dried S nanoparticles were 324.8±6.4 nm, 0.211 ± 0.013 and 28.7±1.9 mV after re-dispersion. The dissolution rate of the S nanosuspension was superior compared to physical mixture in mimic gastric (pH 1.2) and intestinal (pH 6.8) media. Following oral administration of SG, S or S nanosuspension to SD rats, they were transformed into the same two major metabolites SG (M1) and its isomer (M2). Meanwhile, the pharmacokinetic parameters of M1 and M2 indicated that S had an enhance bioavailability than SG, and nanosizing could significantly change the in vivo behavior of S. After oral administration, the pharmacokinetic profiles revealed that S nanosuspension afforded higher peak and short Tmax. The Cmax was increased to 740% of SG and 362% of S, respectively, and the Tmax was only 0.54±0.48 h (8.00 ±2.45 h of SG, 4.08±5.41 h of S). The results of zebrafish tests showed that S nanosuspension had a further improvement of therapeutic effect on cerebral hemorrhage than free S which was better than SG. At each treatment dosage (50, 100, and 200 µM), it supposed that S and SG groups effectively rescued the hemorrhage phenotype, and by 72 hpf approximately 10.32±9.02%和 10.37±10.02% in 100µM and 200µM of the embryos showed evident cerebral hemorrhages, compared with 89.68±9.02% of control embryos receiving atorvastatin and the vehicle Overall, S nanosuspension were considered to be used as a potential delivery formulation with enhanced dissolution rate and rescue activity on zebrafish cerebral hemorrhage for S.

Chinese Abstract

黃酮類化合物野黃芩苷(Scutellarin, SG),又名燈盞花乙素,是傳統中藥材 燈盞花的主要活性成份,被廣泛的應用於心血管疾病的臨床治療。目前已有多種 野黃芩苷類製劑上市,比如燈盞花素注射液,燈盞花素分散片和燈盞花素滴丸等。 其中燈盞花素注射液被 2010 版中國藥典收錄,并作為心腦血管科室常規處方用 藥,臨床上用於治療腦血栓、腦出血及其後遺症、冠心病、心絞痛、心律失常、 高粘血症、頑固性頸周綜合征、椎基底動脈供血不足及其他缺血及伴有微循環障 礙等疾病。但是水溶性差和口服生物利用度低等問題成為限制它應用的因素。SG 在水中溶解度僅為 0.16 mg/mL,在 Beagle 犬和大鼠上口服絕對生物利用度分別 為 0.40%和 10.67%。 野黃芩素(Scutellarein, S),是 SG 的苷元,其透过细胞单层膜的渗透能力比 SG 好,在体内 SG 需要被水解成为 S 才能在胃肠道被吸收.在吸收过程中和入 血后,S 被迅速代谢成其醛酸 SG 和 SG 的同分异构体 SG’的形式在体内循环, 而 S 原形在体内含量很低.本研究欲通過水解的方法大量製備 S,并製備 S 納米 混懸液,考察 S 納米混懸液的在大鼠體內的藥物代謝和動力學行為,以及考察在 斑馬魚腦出血模型上的藥效作用。 本研究中首先成功建立了一個從SG原药中大量製備S固体的四步法-酸水解,結 晶,沉澱,凍乾。這個方法中,水解時間2小時,產物固体粉末HPLC純度在95% 以上,回收率達52.5%(理論回收率61.9%)。在此基礎上,本研究以水解得到的 S为模型药,利用反溶劑沉澱法製備S納米混懸液。通過單因素實驗设计,考察 了穩定劑種類、穩定劑用量、有機相比例以及有機相中藥物含量等因素對粒徑、 粒徑分佈(多分散係數,PdI)、Zeta電位和穩定性的影響。篩選出合適的凍乾保 護劑,應用冷凍乾燥的方法對S納米混懸液進行固化,獲得凍乾粉末,同時研究 凍乾保護劑種類對S納米混懸液在不同分散介質中重分散能力的影響。最終製備 工藝如下:50mg S溶解於2 mL甲醇和DMF比例為3:1的混合混合有機相,超聲條 件下用注射器注入到体积为20 mL含有0.2%(w/v)F68的水相中,保持超聲2 min。 以2%(w/v)D型海藻糖或者5%(w/v)羥丙基β環糊精作為凍乾保護劑,置於 -80℃ 冰箱預凍12 h后,在真空冷凍乾燥機中乾燥24 h,獲得S納米凍乾粉末(SP-DT或 者SP-HP-β-CD)。SP-DT在室溫保存條件下具有至少兩個月的短期穩定性,重分 散后的粒徑為324.8±6.4 nm, PdI為0.211 ± 0.013,Zeta電位為-28.7±1.9 mV。透 射電鏡和X射線衍射結果表明,重分散后,含有D型海藻糖的S納米混懸液 (SN-DT)中的S納米粒以無定型的形式存在。溶出實驗中,以模擬胃液(pH1.2) 和模擬腸液(pH6.8)作為溶出介質,以D型海藻糖作為凍乾保護劑的S納米凍乾粉 末(SP-DT)在5 min內釋放87.2%和77.2%,10 min內釋放完全。和SP-DT類似, 和以羥丙基β 環糊精做誒凍乾保護劑的S納米凍乾粉末(SP-HP-β-CD)在5 min 內分別釋放90.7%和88.9%,10 min內釋放完全。相比之下由於晶體粒徑較大(9.2 ±4.1 µm),原藥在10 min時在模擬胃液和模擬腸液中釋放分別為16.8%和 13.9%,以上結果表明納米化之後S溶出速率加快。 體內實驗以雄性 SD 大鼠為模型動物,建立了 S 和 SG 在大鼠血漿中的分析 方法,研究了 SG,S 和 S 納米混懸液口服給藥之後的藥物動力學特徵。實驗發 現口服給藥 SG,S 或者 S 納米混懸液(含有凍乾保護劑 HP-β-CD)之後,血漿中 均無法檢測到 S 原型的存在,同時發現 SG 和 S 在體內均轉化成相同的代謝產物 M1 和 M2。實驗證明 M1 即為 SG 原型,和 M2 互為同分異構體。S 的口服生物 利用度比 SG 提高至 142%,達峰時間(Tmax)分別為 4.08±5.41 h 和 8.00±2.45 h,并且兩者的藥物動力學特徵接近,血藥濃度-時間曲綫中血藥濃度平穩,無明 顯藥峰。S 經過納米化之後,藥物動力學特徵發生明顯改變,達到藥物濃度峰時間快,僅為 0.54±0.48 h,藥峰濃度分別比 SG 和 S 組提高至 740%和 362%,消 除半衰期 t1/2 僅為 2.68±0.88 h,顯著低於 SG(12.20±2.98 h)和 S(7.29±5.75 h)組(p< 0.01)。結果表明 S 降低斑馬魚腦出血率的作用强 於 SG,經過納米化之後 S 作用效果進一步增強。 結論:本研究成功從 SG 中製備出 S 並將 S 納米化。S 納米混懸液可以提高 體外溶出度。S 經過納米化之後改變在大鼠體內的藥物動力學特徵,主要體現在 消除半衰期縮短,血藥達峰濃度提升和達峰時間提前,同時 S 納米混懸液顯著提 高斑馬魚腦出血后救治作用。S 納米混懸液是一種有潛力被開發成針對腦出血后 救治效果的口服納米製劑。

Issue date

2014.

Author

楊瀟

Faculty

Institute of Chinese Medical Sciences

Degree

M.Sc.

Subject

Cardiovascular system -- Diseases -- Treatment

心血管系統 -- 疾病 -- 治療

Pharmacokinetics

藥代動力學

Supervisor

Zheng, Ying

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Location
1/F Zone C
Library URL
991005761089706306