Protopanaxadiol (PPD) is a Darma alkanes tetracyclic triterpene with poor solubility (<50 ng/mL) and high lipophilicity. PPD exhibits strong anti-cancer activity and antidepressant-like effect, whereas its poor aqueous solubility limits its application. The aim of the study was to fabricate PPD nanocrystals to improve its oral bioavailability and delivery into the brain. In this study, single factor experiment, including various type of solvents and stabilizers, drug concentration, organic/aqueous solvent ratio and stirring time, was used to study their effects on particle size, polydispersity index (PdI), zeta potential and short-term stability. The PPD nanocrystals were prepared using anti-solvent precipitation where D-α-Tocopheryl polyethylene glycol 1000 succinate (TPGS) was optimized as the stabilizer. The filtration tube was centrifugal at 4000 rpm/min for 25 min to remove the organic phase. Then, equal volume Milli-Q water was added back in the filtration tube to maintain the PPD nanocrystals at the same concentration. The formulation was consisting of 0.5 mg/ml drug, PPD/TPGS at 3:1, Milli-Q water/acetone at 10:1, and the particle size, PdI, and zeta potential of the PPD nanocrystals were 90.46±1.45 nm, 0.149±0.016, and -21.2±0.85 mV，respectively. The fabricated nanocrystals were nearly spherical with uniform size distribution observed by transmission electron microscope. Powder X-ray diffraction analysis indicates that PPD was present in a crystalline state in the nanocrystals. In-vitro stability study showed that PPD nanocrystals can be stabilized at 4ºC for one month. In in-vitro dissolution experiment, more than 80% PPD could be rapidly released from nanocrystals within 1 min in 0.2% (w/v) sodium lauryl sulfate at 37±0.5 ºC. And the permeability value of the PPD solution was (3.05±0.74) ×10-5 cm/s, which is ~38 times higher than that of lucifer yellow and is comparable to that of propranolol, suggesting that permeability is not the rate-limiting step in oral absorption of PPD. According to the biopharmaceutics classification system (BCS), PPD is classified as a BCS II class compound. After oral administration, PPD nanocrystals were absorbed quickly into plasma with a Tmax of 0.70 h, and the plasma Cmax values of PPD nanocrystals were 4.75-fold higher than that of the physical mixture. Oral bioavailability of PPD nanocrystals (AUCo-t) was 1.81-fold superior than the physical mixture. Interestingly, there was about 1 h delay for PPD to reach to Tmax in the brain than that in the plasma, which was eliminated slowly than those in the plasma. The results suggested that PPD was rapidly absorbed into plasma, then x transported across the BBB and eliminated in the brain slowly. The intracerebral Cmax values of PPD nanocrystals were 1.84-fold higher than that of the physical mixture. Moreover, the brain AUC0-t of PPD after dosing with the PPD nanocrystals was 1.86-fold higher than that of the physical mixture. However, no brain targeting was observed, as the ratios of the plasma AUC0-t to brain AUC0-t for the two groups were similar. In summary, PPD nanocrystals are a potential oral delivery system to improve PPD’s poor bioavailability and its delivery into the brain for neurodegenerative disease and intracranial tumor therapies in the future.

原人參二醇（20(s)-Protopanaxadiol，PPD）屬於人參二醇类皂苷的皂苷元， 是達瑪烷型四環三萜類化合物。原人參二醇有良好的抗腫瘤活性，有抗疲劳，抗 抑郁等脑内活性。但其水溶性差，口服生物利用度和脑部递送差，限制了其临床 應用。 本研究的目的是針對水溶性差的 PPD，設計口服劑型提高原人參二醇的口服 生物利用度，并提高其入腦濃度。首先採用反溶劑沉澱法製備了 PPD 納米晶混 懸液。本研究通過單因素實驗，考察了有機相種類，藥物濃度，穩定劑種類等因 素對粒徑，PdI 和 zeta 電位的影響。處方優化結果如下：5 mg PPD 溶解於 1 mL 丙酮中，在 1000 rpm 搅拌條件下，将有机溶剂通过注射器快速注入到 10 mL 含 0.015%（w/v）TPGS 水相中，繼續高速攪拌 5 min。用超濾法除有機溶劑，4000 rpm/min 離心 25 min，用等體積 Milli-Q 水復溶，最終得到 PPD 納米晶混懸液。 优化的 PPD 納米晶混懸液粒徑為 90.46±1.45 nm，PdI 值為 0.149±0.016, ，zeta 電位為 -21.2±0.85 mV。透射電鏡和 X-射線衍射譜圖結果表明，PPD 以類圓形晶 體形態存在于 PPD 納米晶混懸液中。溶出實驗以 0.2% SDS 為介質，PPD 納米晶 混懸液在 1 min 內累積釋放達到 80%，20 min 釋放完全。而物理混合物中在 60 min 後僅有约 25% PPD 可釋放。以上結果說明，PPD 納米化後溶出速率加快。 溶解度實驗表明，納米化后的 PPD 溶解溶解度提高了至少 16 倍。 在細胞實驗中，PPD 溶液在 MDCK 單層細胞的 Papp為 (3.05±0.74) ×10-5 cm/s， 是熒光黃的表觀滲透係數的約 38 倍，與普萘洛爾的表觀滲透係數相近，闡明 PPD 是屬於生物药剂分类系统 II 类（biopharmaceutics classification system，BCS）的 化合物。PPD 溶液與 PPD 納米晶混懸液的細胞滲透率相比沒有顯著性差異，說 明製備成製劑後，PPD 的細胞透過率並未提高。體內動物實驗發現，PPD 製成 納米晶混懸液後，Cmax提高了 375 %，AUC0-t提高了 81%。說明 PPD 製成製劑後， 顯著提高了 PPD 的口服生物利用度。腦部递送實驗發現，PPD 製成納米晶混懸 液给大鼠口服後，Cmax提高了 81 %，AUC0-t 提高了 86%。說明 PPD 製成製劑後， 顯著提高了 PPD 的腦內濃度。計算藥物靶向率（Drug targeting efficiency，DTE） 發現，PPD 納米晶混懸液和物理混合物的 DTE 相近，說明製備成 PPD 納米晶混 viii 懸液并未提高 PPD 的腦靶向率。可能因為製備成劑型后，口服后 PPD 吸收提高， 在血中濃度提高，導致了 PPD 腦內濃度的提高。 本研究採用反溶劑沉澱法，PPD 和 TPGS 以 3:1 的质量比製備成納米晶体混 懸液。口服給藥后，PPD 納米晶体混懸液提高了 PPD 的大鼠口服生物利用度以 及腦內濃度。本研究為將來的腦內疾病和腦部腫瘤的治疗提供了一个具有前景和 参考价值的口服难溶藥物遞送系統。