| Magnetic modification of cell is a new type of immobilization method to obtainactive cell with magnetic response by attaching magnetic nanoparticles to cell surface.Magnetically fixed bed (MFB) is able to retain magnetically modified cell(MNP@Cell) by ferromagnetic material packed under extra magnetic field, whenactivity of retained MNP@Cell is remained. In this work, magnetic modification ofLactobacillus reuteri, Escherichia coli and Saccharomyces cerevisiae was studied indetail. Furthermore, the retention properties of MNP@Cell in MFB and itsbio-conversion of glycerol were explored.Firstly, the magnetic responses of MNP@cells prepared from three typicalmicroorganisms under different conditions were studied. L. reuteri, E. coli and S.cerevisiae were magnetically modified in fresh medium, fermentation broth andphosphate buffer (or deionized water) respectively to investigate the effect ofconditions such as cell/particle mass ratio (MRc/p), pH, particle concentration andNaCl concentration on magnetic separation of MNP@Cell. Three microorganisms canbe modified successfully. The modified L. reuteri had better magnetic response thanthe other two at any conditions. The best magnetic response for the modified L.reuteri was reached at pH3, the MRc/pof1and the particle concentration of14.46mg/mL without NaCl added. E. coli and S. cerevisiae could not be modified in freshmedium or fermentation broth, wheras they could be modified in water (pH=2-4orNaCl concentration above0.4mol/L). The following was surgested to obtainMNP@Cell with high magnetic response:1) the interaction between groups on thesurfaces of cell and nanoparticle;2) opposite surface charges of cell and nanoparticleat pH between their isoelectric points;3) particle load on cell surface, dispersity ofnanoparticle and aggregation of MNP@Cells are critical to magnetically separation.Secondly, a novel MFB was built to investigate the effect of coil current, flowrate, bed void and MRc/pon retention of MNP@Cell. MFB was packed with stainless steel wire to retain MNP@Cell under extra magnetic field generated by solenoid. Theoptimum conditions were as following: current of3.0A, flow rate of9mL/min, bedvoid of0.88and MNP@Cells are modified at MRc/pof1.5. Three methods generallywas used to improve the cell retention of MFB:1) increase current to enhanceexternal magnetic field, but current increasing cause a higher degree thermal inactivatingof MNP@Cell;2) decrease bed void or use pack with better permeability to enhanceinternal magnetic field, but decreasing bed void may result in bed blocking;3)increase the particle load on cell surface to enhance the magnetic response ofMNP@Cell, overload certainly reduce cell activity.Finally, an improved MFB reactor was used to retain magnetically modified L.reuteri and convert glycerol to3-Hydroxypropionaldehyde (3-HPA). Batchconversion conditions (glycerol concentration, circulation rate and pH), reactivationconditions (reactivation substrate, first conversion time and circulation rate) andreutilization of MNP@Cell were investigated in detail. In batch conversion, theconcentration of3-HPA reached180.3mmol/L, molar yield of3-HPA to glycerolreached75.1%when240mmol/L glycerol was converted with circulation rate of10h-1for2h at pH7. The MNP@Cell used for1h conversion of glycerol wassequentially reactivated in reactivation medium (containing10g/L glucose,20g/Lyeast extract, pH=7) with circulation rate of10h-1for8h to recover its highestactivity.3-HPA concentration after1h conversion decreased from113.1mmol/L to10.6mmol/L with5batch reutilization. Glycerol conversion yield remained70%ofits previous batch. The results showed that:1) glycerol of excessive concentrationreduce the activity of GDHt;2)3-HPA of high concentration make MNP@Cellinactivated during bio-conversion, bio-conversion ability of inactivated cell could bepartially recovered by cell reactivation. |
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