Preparation And Evaluation Of Liposomes Constructed By D-α-tocopheryl Poly(2-ethyl-2-oxazoline) Succinate For Docetaxel And Ginsenoside Rg3 Co-delivery

Docetaxel（DOC）is one of the most widely used anticancer drugs,exhibitting high-active to a variety of solid tumors.Ginsenoside Rg3（G-Rg3）is a kind of tetracyclic triterpenoid saponins in ginseng,which has certain anti-tumor effect.For the purpose of attenuating toxicty and targeting anti-tumor efficacy synergistically,this article adopts a pH-sensitive polymer,D-α-tocopheryl poly（2-ethyl-2-oxazoline）succinate（TPOS）was constructed into acid-sensitive liposomes for the combined delivery of both drugs.High Performance Liquid Chromatography（HPLC）method was established for the content analysis of DOC and G-Rg3.The detection wavelengths were determined（228 nm and 205 nm）,and the specificity,linear range,precision and recovery rate of the method were also examined.The results showed a good linear relationship between the two drugs within the set concentration range.The precision and recovery results showed that the method met the requirements of content analysis providing the basis for the subsequent quality evaluation.DOC,G-Rg3 and DOC/G-Rg3 loaded liposomes were prepared by the lipid film hydration method.Utilizing the“post-insertion”method,TPOS coated DOC/G-Rg3 loaded liposomes were prepared.The optimal phospholipids type and ratio of phospholipids to lipids of DOC/G-Rg3-L were screened and optimized according to the encapsulation efficiency and loading efficiency.The optimized ratio is:6:1（SPC to cholesterol,w/w）and 1:1:60（DOC/G-Rg3/SPC,w/w/w）.The encapsulation efficiency and loading efficiency of DOC in the optimized DOC/G-Rg3-L was（92.71±1.58）%and（0.94±0.02）%,respectively.The encapsulation efficiency and loading efficiency of G-Rg3 in the optimized DOC/G-Rg3-L was（93.52±1.64）%and（3.20±0.43）%,respectively.The average particle sizes of DOC/G-Rg3-L and TPOS-DOC/G-Rg3-L were less than 100 nm.The liposomes were spherical and had obvious bimolecular structure through scanning electron microscope（SEM）and transmission electron microscopy（TEM）.X-ray powder diffraction（XRD）analysis showed that two drugs were both encapsulated into liposomes.The stability was evaluated by determining the light transmittance of the liposomes incubating in 10%fetal bovine serum（FBS）for 24 h.The results showed that the transmittance of DOC/G-Rg3-L and TPOS-DOC/G-Rg3-L were 78%and 90%,respectively,indicating that the stability of TPOS-DOC/G-Rg3-L was better than that of DOC/G-Rg3-L.The in vitro releasing condition of the liposomes was investigated by dialysis method.Compared with DOC/G-Rg3-L,TPOS-DOC/G-Rg3-L showed a sustained low release under the condition of pH 7.4.The release amount of TPOS-DOC/G-Rg3-L at pH 6.4 was significantly much higher than that of TPOS-DOC/G-Rg3-L at pH 7.4,which showed obviously rapid release,indicating TPOS-DOC/G-Rg3-L has pH-sensitivity.The intracellular uptake of He La and MCF-7 cells was determined by HPLC.The highest uptake of TPOS-DOC/G-Rg3-L was observed in both kinds of cell lines,especially at pH 6.4,which showed the pH-sensitivity of TPOS-DOC/G-Rg3-L.At the same time,the cell inhibition of each formulation was evaluated by MTT method.The results showed that TPOS-DOC/G-Rg3-L had higher cell inhibition.With the decrease of pH,only TPOS-DOC/G-Rg3-L showed enhanced inhibition on cells and showed pH-sensitivity.The calculated IC₅₀ values showed that TPOS-DOC/G-Rg3-L was the lowest,which was consistent with the results of cell inhibition.The synergistic effect of combined administration was evaluated by combination index（CI）.The results showed that,compared with DOC+G-Rg3,DOC/G-Rg3-L showed moderate synergistic effect in He La cells,superpositional effect in MCF-7 cells,which showed that DOC and G-Rg3 co-loaded liposomes had synergistic antitumor effect on He La cells in vitro.