金屬離子螯合劑，例如：diethylene triamine pentaacetic acid (DTPA)，可藉由螯合二價金屬離子，達到打開細胞間緊密連接蛋白(tight junction)及抑制酵素活性的功效。本研究以DTPA修飾聚麩胺酸[poly(γ-glutamic acid), γ-PGA]，與幾丁聚醣(chitosan, CS)形成離子鍵結型奈米微粒，作為胰島素的口服釋放載體。由於CS具有黏膜吸附性，可以使DTPA集結在小腸黏膜，於釋放藥物的局部發揮抑制酵素活性及促進paracellular pathway通透性的功能。由體外(In vitro)實驗結果證實，所合成出的γPGA-DTPA確實可表現出促進細胞間滲透性及抑制酵素活性的效力；γPGA-DTPA可和CS形成具有pH值敏感性的奈米微粒，其粒徑大小會隨pH值增加而逐漸膨脹，且在pH 7.0以上的環境中崩解。此外，本研究以雷射掃描共軛焦顯微鏡及單光子放射斷層掃描(SPECT)觀察CS/γPGA-DTPA奈米微粒在生物體內的分布情形。由實驗結果得知，CS/γPGA-DTPA奈米微粒可促進胰島素在整個小腸區段中被吸收，且可在實驗鼠的腎臟及膀胱中清楚地觀察到被吸收的胰島素。此外，藥效學及藥物動力學實驗結果直接地證實：藉由此功能性奈米微粒所投遞的胰島素，確實可降低血糖值並延長低血糖持續的時間(與皮下注射胰島素溶液相比)；在投藥後四小時，胰島素可在實驗鼠體內達到最高的濃度，且藥物生體可用率(bioavailability)大約為20%。綜合本研究的實驗結果，此CS/γPGA-DTPA功能性奈米微粒相當具有潛力做為胰島素的口服釋放載體。

Complexing agents such as diethylene triamine pentaacetic acid (DTPA) are known to disrupt intestinal tight junctions and inhibit intestinal proteases by chelating divalent metal ions. This study attempts to incorporate these benefits of DTPA in functional nanoparticles (NPs) for oral insulin delivery. To maintain the complexing agent concentrated on the intestinal mucosal surface, where the paracellular permeation enhancement and enzyme inhibition are required, DTPA was covalently conjugated on poly(γ-glutamic acid) (γPGA). The functional NPs were prepared by mixing cationic chitosan (CS) with anionic γPGA-DTPA conjugate. The γPGA-DTPA conjugate inhibited the intestinal proteases substantially, and produced a transient and reversible enhancement of paracellular permeability. The prepared NPs were pH-responsive; with an increasing pH, CS/γPGA-DTPA NPs swelled gradually and disintegrated at a pH value above 7.0. Additionally, the biodistribution of insulin orally delivered by CS/γPGA-DTPA NPs in rats was examined by confocal microscopy and scintigraphy. Experimental results indicate that CS/γPGA-DTPA NPs can promote the insulin absorption throughout the entire small intestine; the absorbed insulin was clearly identified in the kidney and bladder. In addition to producing a prolonged reduction in blood glucose levels, the oral intake of the enteric-coated capsule containing CS/γPGA-DTPA NPs showed a maximum insulin concentration at 4 h after treatment. The relative oral bioavailability of insulin was approximately 20%. Results of this study demonstrate the potential role for the proposed formulation in delivering therapeutic proteins by oral route.

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