Evaluation Of Efficacy And Safety On Nanosized Cationic Amylose-encapsulated Bovine Hemoglobin

Background：Development of effective and safe blood substitutes is the basic way to solve theblood shortage and other world-wide problems. Nanoscale artificial red blood cell isconsidered to be the most prospective of hemoglobin-based oxygen carriers which is madeby biodegradable polymer materials and have structure similar to natural red blood cell.Objective：A novel comprehensive strategy for the preparation of nanosized cationicamylose-encapsulated bovine hemoglobin （NCAHbs） as a nanosized oxygen carrier wasdeveloped. The stability and quality were controled by the physical and chemicalproperties; A lethal hemorrhagic shock model in rats was used to evaluate the efficacy ofthe NCAHbs. The tests of cytotoxicity, abnormal toxicity, acute toxicity and bloodcompatibility were used to evaluate the safety of the NCAHbs. These results are prepared for the further preclinical safety evaluation and human clinical trials.Methods：The morphology of particle, entrapment efficiency （EE%）, drug loading efficiency（DL%）and the stability of different batches NCAHbs were measured by Transmissionelectron microscopy and Malvern Nano-zs90analyzer. Comparing the changes ofhemodynamic parameters and24h survival rate with different resuscitation liquid onhemorrhagic shock rats model. The degree of SOD and MDA were also measured. ED₅₀and T1/2were measured by sequential method and Cyanide-methemoglobin method.Following the state standard, the cellular proliferation of HFbs cells was examined usingmitochondrial function（MTT）assay with different concentration of NCAHbs. Evaluatingthe effects of PLT,PT on NCAHbs. The clinical manifestations,the pathological changes ofthe important target organs and LD₅₀were observed within14d.Results：The average diameter of the NCAHbs was97.13±3.04nm, with a narrowpolydispersity index of0.039showed by TEM. The EE%was78.25%±0.98%, and theDL%was60.72%±1.14%. The average size of the NCAHbs was163.57nm, with a narrowpolydispersity index of0.03showed by Malvern Nano-zs90analyzer. The average degreeof Zeta potential was6.79mv, and the PH was7.34±0.32.The P50was29mmHg and theHill index was2.5.There was no free hemoglobin founded within a month. NCAHbsmaintain the mean arterial pressure effectively on hemorrhagic shock model, alsolightening the degree of lung injury caused by ischemia-reperfusion. The ED₅₀was1.13g/kg,95%confidence interval was0.64～1.69g/kg; The T1/2was19.4～21.5hinvivo. NCAHbs shows no obvious cytotoxicity and negative effect on PLT and PT. Theundue toxicity was qualified. The damage were obvious observed in lung and kidneyabove certain dose. LD₅₀ was5.35g/kg,95%confidence interval was2.37～6.85g/kg. Conclusion：The NCAHbs can improve the prognosis of ischemia reperfusion and providescientific data for further clinical trials with excellentphysical and chemical properties,stable and controllable quality and good ability of carrying-releasing oxygen，whichwould became ideal blood substitute.