目的 制备一种生物利用度良好的芍药苷-甘草蛋白自组装纳米粒（paeoniflorin-glycyrrhiza protein self-assembled nanoparticles，Pae-GP/SAN）并考察其肠吸收机制。方法 以甘草蛋白为载体，采用超声分散法制备Pae-GP/SAN，以平均粒径、多分散系数（polydispersity index，PDI）、包封率、载药量为评价指标优化处方和制备工艺，并对制备的纳米粒进行表征。建立大鼠在体单向肠灌流模型，考察并对比芍药苷溶液（Pae/Sol）、芍药苷-甘草蛋白物理混合物（Pae-GP/PM）以及Pae-GP/SAN的肠吸收行为，并应用水淬灭ACQ（aggregation-caused quenching）荧光探针，通过激光扫描共聚焦显微镜（confocal laser scanning microscope，CLSM）观察GP/SAN的肠吸收情况，初步阐明其促渗机制。结果 优化后Pae-GP/SAN粒径为（178.2±6.3）nm，PDI为0.152 1±0.011 2，ζ电位为（−14.91±1.13）mV，包封率为（36.45±2.32）%，载药量为（21.70±1.30）%，微观形态呈均一球状。肠灌流实验表明，Pae-GP/SAN在回肠中的吸收效率优于空肠（P＜0.05、0.01）；纳米粒的形成可显著促进芍药苷的吸收，且高质量浓度Pae-GP/SAN的吸收参数显著高于低质量浓度（P＜0.001），而质量浓度对Pae/Sol与Pae-GP/PM的吸收无显著影响；转运蛋白抑制剂干预实验显示，吲哚美辛和利血平对3种制剂的吸收均无显著影响；维拉帕米可显著提高Pae/Sol和Pae-GP/PM的吸收（P＜0.01、0.001），而对Pae-GP/SAN无显著影响。CLSM观察证实GP/SAN能够以完整纳米粒形式被肠道吸收。结论 Pae-GP/SAN可通过完整纳米粒形式被肠道内吞吸收，有效规避P-gp外排蛋白的屏障作用，进而显著提升芍药苷的口服生物利用度。

Objective To prepare paeoniflorin-glycyrrhiza protein self-assembled nanoparticles (Pae-GP/SAN) with good bioavailability and investigate its intestinal absorption mechanism. Methods In this study, glycyrrhiza protein (GP) was used as the carrier to prepare Pae-GP/SAN by ultrasonic dispersion method. The formulation and preparation process were optimized using average particle size, polydispersity index (PDI), encapsulation efficiency, and drug loading as evaluation indicators, and the prepared nanoparticles were characterized. A rat in situ single-pass intestinal perfusion model was established to investigate and compare the intestinal absorption behaviors of paeoniflorin solution (Pae/Sol), paeoniflorin-glycyrrhiza protein physical mixture (Pae-GP/PM), and Pae-GP/SAN. Additionally, an aggregation-caused quenching (ACQ) fluorescent probe was used to observe the intestinal absorption of GP/SAN by confocal laser scanning microscopy (CLSM), aiming to initially clarify its penetration-enhancing mechanism. Results The optimized Pae-GP/SAN had an average particle size of (178.2 ± 6.3) nm, a PDI of 0.152 1 ± 0.011 2, a ζ potential of (−14.91 ± 1.13) mV, an encapsulation efficiency of (36.45 ± 2.32)%, and a drug loading of (21.70 ± 1.30)%, with a uniform spherical micromorphology. Intestinal perfusion experiments indicated that the absorption efficiency of Pae-GP/SAN in the ileum was superior to that in the jejunum (P < 0.05, 0.01). The formation of nanoparticles significantly promoted the absorption of paeoniflorin. Moreover, the absorption parameters of high mass concentration Pae-GP/SAN were significantly higher than those of low mass concentration Pae-GP/SAN (P < 0.001), while mass concentration had no significant effect on the absorption of Pae/Sol and Pae-GP/PM. Transporter inhibitor intervention experiments showed that indomethacin and reserpine had no significant effect on the absorption of the three formulations. Verapamil significantly increased the absorption of Pae/Sol and Pae-GP/PM (P < 0.01, 0.001) but had no significant effect on Pae-GP/SAN. CLSM observations confirmed that GP/SAN can be absorbed by the intestine in the form of intact nanoparticles. Conclusion Pae-GP/SAN can be endocytosed and absorbed by the intestine as intact nanoparticles, effectively evading the barrier effect of P-gp efflux protein, thereby significantly improving the oral bioavailability of Pae.

R283.6

江西省教育厅科学技术研究项目（GJJ2403609）；国家自然科学基金项目（82260848）；江西省自然科学基金项目（20232BAB216139）