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BACKGROUND: The free enzyme has the problems of poor stability and inability to be reused during application. Using magnetic polymer microspheres as the carrier of the binding enzyme to prepare immobilized enzyme can maintain the natural activity of the enzyme, and can be reused, but also provide convenient conditions for automatic production management. OBJECTIVE: Magnetic chitosan microspheres as carrier of binding enzyme were used to prepare the immobilized lactase, which is easy to recycle, can be reused, and has high enzyme activity and stability. METHODS: By inputting a certain amount of magnetic chitosan microspheres into the phosphoric acid buffer for swelling for 2 hours, the swelling magnetic chitosan microspheres were collected with a magnet and added to a certain concentration of lactase phosphate buffer. They were shocked in a constant temperature shaker for 1 hour and preserved in refrigerator at 4 °C. The microspheres were precipitated with a magnet to pour out the supernatant. After full washing with buffer solution, the immobilized lactase was obtained. The properties of the magnetic microspheres (the amount of glutaraldehyde used in the preparation of microspheres was 2, 4, 6, 8, 10, 12, 14 mL), the amount of enzymes (0.5, 1, 1.5, 2 g/L) added, the pH (6.4, 6.8, 7.0, 7.2) of the buffer, and the immobilization time (1, 2, 5, 10, 15, 20 hours) were tested to determine the optimal immobilization conditions. RESULTS AND CONCLUSION: (1) The optimum conditions for immobilized lactose with magnetic chitosan microspheres were as follows: magnetic chitosan microspheres prepared with 10 mL of glutaraldehyde were selected as the immobilized carrier of lactase. The amount of enzyme added was 0.3 g/L, pH 7.0 and the immobilization time was 5 hours. (2) Compared with the free enzyme, immobilized lactase showed a wider range of reaction temperature and pH value. (3) The ability of immobilized enzyme binding substrate was enhanced. (4) After repeated use of the immobilized enzyme five times, the enzyme activity remained 65%. (5) The storage stability of lactase was also improved after immobilization.
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BACKGROUND: As carriers of enzymes, cells and drugs, magnetic polymer microspheres have been widely used in the fields of bioengineering, cytology, and biomedicine. OBJECTIVE: To prepare the magnetic polymer microspheres characterized by small particle size, good dispersion, strong magnetic response, safety, and non-toxicity. METHODS: Magnetic chitosan microspheres were prepared by reverse phase suspension process using Fe3O4 as core, paraffin as dispersed medium, Span-80 as emulsifier, and glutaraldehyde as cross-linking agent. The effects of factors including crosslinking time (0, 20, 40, 60, 80, 100, 120, 150 and 180 minutes), reaction temperature (20→50 °C, 30→60 °C, 40→70 °C, 50→80 °C), the concentration of chitosan (0. 01, 0. 02, 0. 03, 0. 04, 0. 05 g/mL), Fe3O4/chitosan mass ratio (1:1, 1:2, 1:3, 1:4), the amount of glutaraldehyde (8-10 mL), the amount of liquid paraffin (40, 60, 80, 100 mL), and stirring speed (0-2 000 r/min) on the properties of magnetic chitosan microspheres. The morphology, particle size, dispersion, and magnetic responsiveness of magnetic chitosan microspheres were characterized. RESULTS AND CONCLUSION: The optimum conditions for preparing magnetic chitosan microspheres were as follows: Starting with glutaraldehyde as crosslinking agent, the reaction was performed at 40 °C for 1 hour and then at 70 °C for 120 minutes. The concentration of chitosan was 0. 02 g/mL, the mass ratio of Fe3O4/chitosan was 1∶2, the dosage of liquid paraffin was 80 mL, the stirring speed was 1 200 r/min, and the dosage of glutaraldehyde was 8-10 mL. Magnetic chitosan microspheres had strong magnetic properties under the applied magnetic field and had good suspension stability in the natural state. The Fe3O4/chitosan composites were spherical, and the nanoparticles were encapsulated in the microspheres, which were core-shell structure. The surface of the microspheres was smooth and monodisperse. The magnetic chitosan microspheres prepared had a diameter of 1-15 μm, which is beneficial to the dispersion and magnetic separation of the microspheres in the reaction system.
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SuperparamagneticSiO2@Fe3O4microspherewithcore-shellstructurewaspreparedbythe method of hydro-thermal and St?ber method, and modified by (3-aminopropyl) triethoxysilane ( APTES) . The obtained amino-Fe3 O4@SiO2 microsphere with controlled surface charge properties was characterized by SEM and TEM, and the surface electric property of amino-microsphere was investigated by Zeta potential measurement. The microsphere was used to extract DNA from human blood, and the solid phase extraction method by microspheres was developed. The mechanism of action between Aminated-Fe3 O4@SiO2 microsphere and DNA was explored, and the gel electrophoresis and PCR test were done on the extraction product. The result showed that genomic DNA with high purity was successfully extracted from whole blood by using amino-Fe3 O4@SiO2 microsphere, with the extraction rate about 70%, the saturated adsorption capacity for DNA was about 40 ng/μg, and the elution could be directly used for further bio-analysis.
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Objective To investigate pathologic changes and method of magnetic microsphere(Fe3O4) embolization for tumor.Methods 18 cases with tumors were treated by embolized using Fe3O4 under fluoroscopy through catheters by Seldinger’s technique,pathological examinations were done in 15 cases before operation,including primary hepatic carcinoma in 8,renal carcinoma in 6,renal myolipoma in 3 and breast carcinoma in 1.Results Pathological examination after embolism showed that necrosis of neoplastic tissue was 100 percent.Conclusion Fe4O4 is a good embolic substance for treating tumors and there is no damage to the other tissues.
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Objective To investigate the clinical applied value of magnetic mitomycin microspheres and magnetic Fe 3O 4 in treating tumors and vascular diseases by embolism.Methods The embolic therapy in 106 cases useing microspheres or drugs of Fe 3O 4 under TV-guided through catheterization using Seldinger's technique were performed.Results In 94 malignant tumors and 12 benign vascular diseases.The most of tumors became small remarkably on CT,USG or BSA one month later after embolism.Pathology showed that necrosis of cancer tissue was 100 percent.Examination of blood biochemistry showed that the functions of liver and kidney had some changes,however these functions could recover to normal or better after therapy.AFP decreased significantly after embolism in most primary hepatic cancer.Changes in density of serum Fe before and after embolism showed that embolism of Fe 3O 4 microspheres could maintain serum Fe in normal condition.DSA showed that microspheres of Fe 3O 4 were moved from the near portion to the peripheral embolic arteries after embolism one month later and the clinical symptoes improved.Conclusion Fe 3O 4 is good embolic substance and can avoid some complications by embolism.
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Objective To investigate the mechanism and the clinical value of magnetic microsphere(Fe_3O_4) in embolizing tumor through artery catheter.Methods 94 patients with tumor were treated by embolized using Fe_3O_4 under fluoroscopy through catheters by Seldinger's technique.Of them,5 cases of hepatocellular carcinoma underwent magnetic field putting on outside the target area,pathological examination was done in 15 cases,25 cases underwent DSA examination one month later and the DSA images before and after operation were analysed statistically.Results Pathological examination after embolism showed that necrosis of neoplastic tissue was 100 percent.The statistical analysis of DSA images was in P