磷脂包覆银-石墨烯量子点多功能纳米粒的制备与体外评价
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摘要
本文制备了磷脂包覆银-石墨烯量子点多功能纳米粒(ADG-DDPC)并对其进行了体外评价。通过正负电荷之间的相互吸引作用,使得阳离子磷脂1,2-二油烯氧基-3-三甲氨基丙烷(1,2-dioleoyl-3-trimethy-lammoniumpropane)(DOTAP)优先吸附在银纳米粒(AgNPs)内核的表面,利用相转换原理及疏水作用使得二硬脂酰基磷脂酰乙醇胺-聚乙二醇-环肽(DSPE-PEG_(2000)-cRGD)自组装到DOTAP表面,形成稳定的多功能纳米制剂,并对其紫外吸收特性、粒径分布性质、形貌、释放行为、杀灭癌细胞能力及细胞摄取情况进行研究。合成的纳米制剂的最大紫外吸收峰在400 nm左右,马尔文粒径仪及透射电子显微镜均证明了该纳米制剂粒径约为30～40 nm,分布均一。纳米制剂的释放与H_2O_2浓度呈正相关。与AgNPs对肿瘤细胞的IC50值[(347.78±0.06) ng·mL~(-1)]比较, ADG-DDPC的IC50值为(209.68±0.09) ng·mL~(-1),具有更强的细胞毒性。通过细胞摄取实验,证实了该纳米制剂能被肿瘤细胞摄取,且能在肿瘤部位发光。本文成功制备了ADG-DDPC,该制剂体外具有肿瘤细胞标记的作用及较高体外抗肿瘤活性。
In this paper, multifunctional silver-graphene quantum dot nanoparticles coated with phospholipids(ADG-DDPC) were prepared and their properties were evaluated in vitro. Cationic phospholipids 1,2-diolefinoxy-3-trimethy-laminopropane(DOTAP) was absorbed first onto the surface of the core of silver nanoparticle(AgNPs)through the mutual attraction between the positive and negative charge. Based on the principle of phase trans‐formation and hydrophobic interaction, dstearyl-phosphatidylglycolamine-polyethylene-glycol-cyclic-cRGD peptide(DSPE-PEG_(2000)-cRGD) self-assembled onto the outlayer of DOTAP of AgNPs. A stable multifunctional nano-preparation was formed and its ultraviolet absorption, particle size distribution, morphology, in vitro release behavior, ability to kill cancer cells and cell uptake were studied. The maximum UV absorption of the synthesized nanometer preparation was about 400 nm. Malvern particle size meter and transmission electron microscope showed that the particle size of the nano-preparation was about 30-40 nm and its particle size distribution was uniform. The in vitro release of nano-preparation was positively correlated with the concentration of H_2O_2. The IC50 value of AgNPs for tumor cells was(347.78 ± 0.06) ng · mL~(-1), and the IC50 value of ADG-DDPC for tumor cells was(209.68 ± 0.09) ng · mL~(-1), indicating that ADG-DDPC possessed a stronger cytotoxicity than that of AgNPs. Cell uptake experiment showed that ADG-DDPC could be absorbed by tumor cells and exhibited fluoresce inside those cells. In conclusion, ADG-DDPC was successfully prepared, and in vitro characterization study pointed to that the nano-preparation exhibits a higher antitumor activity than AgNPs.
In this paper, multifunctional silver-graphene quantum dot nanoparticles coated with phospholipids(ADG-DDPC) were prepared and their properties were evaluated in vitro. Cationic phospholipids 1,2-diolefinoxy-3-trimethy-laminopropane(DOTAP) was absorbed first onto the surface of the core of silver nanoparticle(AgNPs)through the mutual attraction between the positive and negative charge. Based on the principle of phase trans‐formation and hydrophobic interaction, dstearyl-phosphatidylglycolamine-polyethylene-glycol-cyclic-cRGD peptide(DSPE-PEG_(2000)-cRGD) self-assembled onto the outlayer of DOTAP of AgNPs. A stable multifunctional nano-preparation was formed and its ultraviolet absorption, particle size distribution, morphology, in vitro release behavior, ability to kill cancer cells and cell uptake were studied. The maximum UV absorption of the synthesized nanometer preparation was about 400 nm. Malvern particle size meter and transmission electron microscope showed that the particle size of the nano-preparation was about 30-40 nm and its particle size distribution was uniform. The in vitro release of nano-preparation was positively correlated with the concentration of H_2O_2. The IC50 value of AgNPs for tumor cells was(347.78 ± 0.06) ng · mL~(-1), and the IC50 value of ADG-DDPC for tumor cells was(209.68 ± 0.09) ng · mL~(-1), indicating that ADG-DDPC possessed a stronger cytotoxicity than that of AgNPs. Cell uptake experiment showed that ADG-DDPC could be absorbed by tumor cells and exhibited fluoresce inside those cells. In conclusion, ADG-DDPC was successfully prepared, and in vitro characterization study pointed to that the nano-preparation exhibits a higher antitumor activity than AgNPs.
引文
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[2]Zhao XX,Liu HR,Hu YB,et al.A novel gelatin-AgNPs coating preparing method for fabrication of antibacterial and no inflam‐mation inducible coatings on PHBV[J].React Funct Polym,2016,107:54-59.
[3]Huang WD,Fang XL,Wang HD,et al.Biosynthesis of AgNPs by B.maydis and its antifungal effect against Exserohilum turcicum[J].I ET Nanobiotechnol,2018,12:585-590.
[4]Ernest V,Gajalakshmi S,Mukherjee A,et al.Enhanced activity of lysozyme-AgNP conjugate with synergic antibacterial effect without damaging the catalytic site of lysozyme[J].Artif Cells Nanomed Biotechnol,2014,42:336-343.
[5]Li BK,Wang CL,Chen P,et al.The synthesis and characteriza‐tion of silver nanoparticle-loaded mesoporous silica material and its antibacterial performance[J].Acta Sci Nat Uni Nakai(南开大学学报(自然科学版)),2018,51:31-38.
[6]Lu L,Zheng LP,Zhao PF,et al.Preparation and antifungal activity of Ag-SiO2core-shell nanoparticles[J].Chin J Bio Eng(生物加工过程),2014,12:51-55.
[7]Subbiah RP,Lee H,Veerapandian M,et al.Structural and bio‐logical evaluation of a multifunctional SWCNT-AgNPs-DNA/PVA bio-nanofilm[J].Anal Bioanal Chem,2011,400:547-560.
[8]Ritter KA,Lyding JW.The influence of edge structure on the electronic properties of graphene quantum dots and nanoribbons[J].Nat Mater,2009,8:235-242.
[9]Wang H,Wang XM.In vitro nucleus nanoprobe with ultra-small polyethylenimine functionalized graphene quantum dots[J].RSC Adv,2015,5:75380-75385.
[10]Wang X,Sun X,He H,et al.A two-component active targeting theranostic agent based on graphene quantum dots[J].J Mater Chem B,2015,3:3583-3590.
[11]Su XQ,Chan CY,Shi JY,et al.A graphene quantum dot@Fe3O4@SiO2based nanoprobe for drug delivery sensing and dual-modal fluorescence and MRI imaging in cancer cells[J].Biosens Bioelectron,2017,92:489-495.
[12]Qin J,Zhang RX,Li JL,et al.CRGD mediated liposomes enhanced antidepressant-like effects of edaravone in rats[J].Eur J Pharm Sci,2014,58:63-71.
[13]Liu PF,Qin LB,Wang Q,et al.CRGD-functionalized mPEG-PLGA-PLL nanoparticles for imaging and therapy of breast cancer[J].Biomaterials,2012,33:6739-6747.
[14]Li JZ,Yuang ZQ,Yan M,et al.pH-sensitive micelles loaded paclitaxel using carboxymethyl chitosan-palmitic acid mediated by cRGD[J].Acta Pharm Sin(药学学报),2016,51:642-649.
[15]Cabral H,Matsumoto Y,Mizuno K,et al.Accumulation of sub-100 nm polymeric micelles in poorly permeable tumours depends on size[J].Nat Nanotechnol,2011,6:815-823.
[16]Sukirtha R,Priyanka KM,Antony JJ,et al.Cytotoxic effect of green synthesized silver nanoparticles using Melia azedarach against in vitro HeLa cell lines and lymphoma mice model[J].Proc Biochem,2012,47:273-279.
[17]Yang Y,Luo L,Li HP,et al.Separation and determination of silver nanoparticle in environmental water and the UV-induced photochemical transformations study of AgNPs by cloud point extraction combined ICP-MS[J].Talanta,2016,161:342-349.
[18]Roman M,Rigo C,Castillo-Michel H,et al.Hydrodynamic chromatography coupled to single-particle ICP-MS for the simultaneous characterization of AgNPs and determination of dissolved Ag in plasma and blood of burn patients[J].Anal Bioanal Chem,2016,408:5109-5124.
[19]Mei Q,Zhang Z.Photoluminescent graphene oxide ink to print sensors onto microporous membranes for versatile visualization bioassays[J].Angew Chem Int Ed,2012,51:5602-5606.
[20]Wang LL,Zheng J,Li YH,et al.AgNP-DNA@GQDs Hybrid:a new approach for sensitive detection of H2O2and glucose via simultaneous AgNP etching and DNA cleavage[J].Anal Chem,2014,86:12348-12354.
[21]Li FF,Zhang XX,Guo SY,et al.Preliminary study on pHsensitive lipid bilayer-coated mesoporous silica nanoparticles as a novel drug carrier for antitumor drug[J].Acta Pharm Sin(药学学报),2013,48:291-297.
[22]Weinstain R,Savariar EN,Felsen CN,et al.In vivo targeting of hydrogen peroxide by activatable cell penetrating peptides[J].JAm Chem Soc,2014,136:874-877.
[23]Tu LX,Xu YH,Tang CY,et al.In vivo imaging in tumorbearing animals and pharmacokinetics of PEGylated liposomes modified with RGD cyclopeptide[J].Acta Pharm Sin(药学学报),2012,47:646-651.