Study On Protein Purification And Bacterium Separation For The Modified Magnetic Nanoparticles

With special physical and chemical properties, Magnetic nanoparticles（MNP） wereapplied widely in medicinal and biochemistry field.MNP with functional group on the sarfacewere usually used to protein and nucleic acid separation、cell separation、bacteria separationdetection、targeted drug etc. For these nanocomposite better application in actual life, the nanomicrospheres must have multifunctional properties of biocompatibility、abundant functionalgroup、hige magnetic responsiveness and easy to magnetic separation. In the work, the severalof functional nanocomposite microspheres were prepared.The modified microspheres wereused to purified protein and separate bacteria.1.Magnetic nanoparticles were modifed with capric acid（MNP-CPAD） by the improved chemistrycoprecipitation method,and were characterized with transmission electron microscopy（TEM）, Xraydiffraction （XRD） and Fourier transform infrared spectrophotometry（FTIR）. The effects ofpH,contact protein concentration,adsorption time,temperatuer and ionic strength onadsorption of lysozyme（Lys） were studied. The results showed that MNP-CPAD wassphere,and its average size was about10nm.When the pH was at10.7,the adsorption capacitywas35.0mg·g^-1, after3consecutive, MNP-CPAD remain can maintain good adsorptionproperties.The adsorption process fitted the Langmuir monolayer adsorption model.Desorption of the lysozyme was achieved by using1.0mol·L^-1NaCl, the elution efficiency was80％after twice elution. Purification lysozyme from egg white was carried out withpurification fold30.9,and the average recovery of enzyme activity72.9％.2. The modified Magnetic nanoparticles（MNP-TDGA） were preparated with Diethylthioglycolate, and were characterized with TEM, XRD and FTIR.The effct of the adsorptioncondtions, including pH, contact protein concentration,adsorption time, ionic strength andadsorbent dosage on adsorption behavior of lysozyme（Lys） and ovalbumin（Ova）. The resultsshowed that the average magnetite microspheres size was about12nm. When the pH was at10.7,the adsorption capacity of lysozyme was120.0mg·g^-1, desorption of the lysozyme wasachieved with1.0mol·L^-1NaCl,the elution efficiency was up to74.56％. When the pH was at4.5,the adsorption capacity of ovalbumin was95.5mg·g^-1, desorption of the ovalbumin wasachieved were investigatde by using pH11.0Tirs-NaOH buffer solution, the elution efficiency was52.3％. MNP-TGAD can be used to extract lysozyme and ovalbumin from eggwhite.Tthe purification fold of lysozyme was29.3,and the average recovery of enzymeactivity was54.1％.3. Magnetic Nanoparticles were prepared in aqueous solution using chemicalcoprecipitation method， and modified with （3-Aminopropyl）triethoxysilane on thissuface,then coated with Teicoplain,which can inhibit gram-positive bacteria.The preparatedMNP-APTES-Tei were characterized with TEM, XRD and FTIR.The microspheres were usedto adsorb and separate Staphylococcus aureus and Escherichia coli,The adsorptionconditions,such as pH, adsorption time, the contact concentration of bacteria, temperature,ionic strength and adsorbent dosage were investigated for the adsorption behavior ofbacterium. It was indicated that the MNP-APTES-Tei can selectively adsorb Staphylococcusaureus, the adsorption capacity was1.20×1011cfu·g^-1,and it scarcely ever adsorbedEscherichia coli.4. Magnetic Nanoparticles Fe₃O₄were first synthesized by chemical coprecipitationmethod, followed by the modification on their surfaces with tetraethylorthosilicate（TEOS）and （3-Aminopropyl）triethoxysilane（APTES）.The surface-modified Fe₃O₄nanoparticles werethen linked to mercaptoacetic acid（SA）,to form the MNP-TEOS-APTES magneticparticles,and Fuchsin basic（BF） was immobilized onto it using the1-ethyl-3-（3-dimethylamin-opropyl）-carbodii-midehy drochloride （EDC） and N-hydroxysuccinmide（NHS）.MNP-TEOS-APTES-SA-BF were characterized with TEM, XRD and FTIR. The microsphereswere used to adsorb and separate Mycobacterium and Escherichia coli,The effects of pH,adsorption time, the contact concentration of bacteria, temperature, ionic strength andadsorbent dosage on the adsorption behavior of bacterium were studied. It was showed thatthe Modified microspheres can selective adsorb Mycobacterium,with the adsorption capacity3.25×1010cfu·g^-1, MNP-TEOS-APTES-SA-BF was applied for the separation and detecitionof mycobacterium in hospital wastewater samples,The results indicated that the content ofmycobacterium was1.37×106cfu·mL^-1.