Construction Of Hemoglobin-based Oxygen Carrier By Template Technique And Evaluation For Its Ability Of Oxygen Carry

Objective?1?Synthesis of highly loaded hemoglobin?Hb?microcapsules to study their electrochemical properties and oxygen carrying capacity.?2?Construction of polydopamine?PDA?encapsulated Hb microspheres to explore their oxygen carrying ability.?3?Preparation of biconcave Hb microcapsules to study their morphological characteristics,electrochemical properties and oxygen-carrying capability.Methods?1?Using spherical manganese carbonate?MnCO₃?particles as the template to fabricate hollow microcapsules,?Hb/GA?₄microcapsules were synthesized using LBL technology.?2?MnCO₃ was chosen as the template and glutaraldehyde?GA?was used as the cross-linker.PDA-Hbmicrocapsuleswerepreparedby self-polymerization of dopamine on the Hb spheres.?3?Biconcave Hb-Ca?OH?₂ microcapsules were synthesized by introducing dextran into the coprecipitation process to modulate the morphology of Hb-Ca?OH?₂particles.?4?Scanning electron microscopy?SEM?,transmission electron microscopy?TEM?,and X-ray photoelectron imaging were used.Spectra?XPS?,X-ray diffraction?XRD?,confocal laser scanning microscopy?CLSM?,and energy dispersive spectroscopy?EDS?were used to characterize the morphology of the functional interface;Fourier transform infrared spectroscopy?FT-IR?were used to characterized the chemical structures oxygen carriers based on hemoglobin?HBOCs?.The ability of Hb in oxygen carriers to bind and release oxygen was evaluated by UV-vis spectroscopy.Cyclic voltammetry?CV?,differential pulse voltammetry?DPV?,time current method and other electrochemical methods were used to study the ability of oxygen carriers.The biosafety of synthetic oxygen carriers was studied using hemolysis experiments and cytotoxicity tests.Results?1?It was confirmed by CLSM that the surface of?Hb/GA?₄microcapsules contained Hb.By comparing the cyclic voltammograms of?Hb/GA?n microcapsules from 1 to 5 layers,it was found that the current values of the 4 layers are almostly consistent with the 5 layers.Oxygen-containing?Hb/GA?₁microcapsulesand?Hb/GA?₄microcapsules were released into an oxygen-free environment.It takes2400 s for?Hb/GA?₄ microcapsules modified electrode to release oxygen,which is longer than?Hb/GA?₁ microcapsules.The hemolysis rate of the?Hb/GA?₄ microcapsules is far below than 5%.?2?EDS analysis results showed that the MnCO₃ template was completely dissolved in the obtained Hb microspheres.The PDA-Hb microcapsules can keep an acceptable stability over a period of several days.FT-IR spectroscopy shows that encapsulation process does not affect the chemical structure of Hb in PDA-Hb microcapsules.UV-vis spectroscopy and electrochemical methods were used to demonstrate that PDA-Hb microcapsules can bind and release oxygen reversibly.The hemolytic rate of PDA-Hb microcapsules was within 1%,and there was no significant cytotoxicity on human kidney cells?HEK293T?.?3?XPS and CLSM graphs demonstrated the presence of Hb in Hb-Ca?OH?₂ microcapsules.FT-IR spectroscopy proved the encapsulation process had no obvious effect on the chemical structure of Hb-Ca?OH?₂ microcapsule Hb.XRD indicated that the coprecipitation of Hb in the microcapsules reduced the crystallinity of Hb-Ca?OH?₂ microcapsules.Electrochemical experiments further confirmed that the reversible oxygen carrying capacity of Hb-Ca?OH?₂ microcapsules.When the ambient temperature was 37°C,the oxygen binding/releasing ability of Hb-Ca?OH?₂ microcapsules was higher than other temperature.Conclusions?1??Hb/GA?₄ microcapsules have good blood compatibility and have the capability of binding and releasing oxygen.These studies can lay a theoretical foundation for HBOCs research and provide technical support.?2?The deposition of polymer membranes to templates by single step technology provides direct benefits to the manufacturing of microcapsules,thereby minimizing labor,cost and assembly complexity.The obtained microcapsules showed good stability,and no adverse effects to blood composition and the cell vitality,which makes them suit for oxygen carrier and other promising candidates in the field of biological medicine.?3?Hb-Ca?OH?₂ microcapsules have high oxygen affinity and are suitable for carrying oxygen,which is expected to be a potential artificial oxygen carrier in the future.In addition,it should be noted that this work provides an alternative to assessing the oxygen carrying capacity of Hb oxygen carriers,paving the way for the design of advanced oxygen carriers.