| Summary: | 碩士 === 明志科技大學 === 生化工程研究所 === 102 === In this work, polyacrylonitrile nanofiber membrane (PAN) was prepared by using electrostatic spinning technique. After a series of treatment for the PAN membrane, the modified membrane was finally functionalized with bromoacetic acidic (BrA), and iminodiacetic acidic (IDA) groups, respectively. The modified membranes wre chelated by Cu(II), Co(II), Ni(II), or Zn(II) as immobilized metal affinity nanofibrous (IMAN) membrane (abbrev. BrA-M or IDA-M). Recombinant E. coli cells containing enhanced green fluorescent protein (EGFP) was disrupted by ultrasonic technique. The isotherm and kinetic adsorption experiments were carried out by using clarified feedstock in a bath stirred tank system. The isotherm data for general protein and EGFP were well correlated by the Freundlich model for both IMAN membranes chelated by these metal ions.
In kinetic experiments, the kinetic data for IMAN membrane were tested by using pseudo-first-order reaction and pseudo-second-order models. Generally speaking, the kinetic studies for general protein and EGFP showed that the adsorption followed a pseudo-second-order reaction. Its rate constant k2 increased with increasing concentrations of adsorption, indicating that the increased concentration of E. coli sample accelerated the adsorption rate. In membrane adsorption chromatography for both IMAN membranes, the dynamic binding capacity (DBC) was evaluated by adsorption breakthrough curve and calculated under different chelated metal ions and flow rates, respectively. The DBC for IMAN membrane was found to be lower at higher operating flow rate. For IDA-M adsorption process at a flow rate of 0.1 ml/min, the order of DBC for the general protein was Co(II) (40.0 mg/g)= Zn(II) (40.0 mg/g )> Cu(II) (16.2 mg/g)> Ni(II) (16.2 mg/g); the order of DBC for EGFP was Co(II) (5.77×106 AU/g)> Zn(II) (4.77×106 AU/g)> Cu(II) (3.47×106 AU/g)> Ni(II) (2.73×106 AU/g). However, for BrA-M adsorption process at the same flow rate, the order of DBC for the general protein was Ni(II) (31.6 mg/g)> Cu(II) (27.4 mg/g )> Zn(II) (21.4 mg/g)> Co(II) (10.6 mg/g); the order of DBC for EGFP was Ni(II) (8.25×106 AU/g)> Cu(II) (4.90×106 AU/g)> Co(II) (3.60×106 AU/g)> Zn(II) (3.41×106 AU/g). Based on the adsorption breakthrough curves, the results showed that IDA-M or BrA-M as IMAN membrane was not suitable for use in the purification of EGFP by using membrane chromatography.
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