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Breakthrough Model of Recombinant Human-Like Collagen in Immobilized Metal Affinity Chromatography

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Abstract

The adsorption of recombinant human-like collagen by metal chelate media was investigated in a batch reactor and in a fixed-bed column. The adsorption equilibrium and kinetics had been studied by batch adsorption experiments. Equilibrium parameters and protein diffusivities were estimated by matching the models with the experimental data. Using the parameters of equilibrium and kinetics, various models, such as axial diffusion model, linear driving force model, and constant pattern model, were used to simulate the breakthrough curves on the columns. As a result, the most suitable isotherm was the Langmuir–Freundlich model, and the ionic strength had no effect on the adsorption capacity of chelate media. In addition, the pore diffusion model fitted very well to the kinetic data. The pore diffusivities decreased with increasing the initial protein concentration, however had little change with the ionic strength. The results also indicated that the models predict breakthrough curves reasonably well to the experimental data, especially at low initial protein concentration (0.3 mg ml−1) and low flow rate (34 cm h−1). By the results, we optimized the experimental conditions of a chromatographic process using immobilized metal affinity chromatography to purify recombinant human-like collagen.

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Abbreviations

a :

Specific surface area of particle (m2/m)

C :

Protein concentration in bulk fluid phase (mg/ml)

C 0 :

Initial protein concentration in bulk fluid phase (mg/ml)

C * :

Equilibrium concentration in bulk fluid phase (mg/ml)

C p :

Protein concentration in pore (mg/ml)

D e :

Effective diffusivity (m2/s)

D 0 :

Molecular diffusivity in free solution (m2/s)

D p :

Pore diffusivity (m2/s)

D z :

Axial dispersion coefficient (m2/s)

F :

Volumetric ratio of solid phase to liquid phase

K b :

Association constant in Langmuir isotherm

k :

Constant in Freundlich isotherm

K f :

Total transfer coefficient (m/s)

k f :

Liquid film mass transfer coefficient on the particle surface (m/s)

L :

Column length (m)

Mr:

Molecular weight (kDa)

n :

Constant in Freundlich isotherm

q :

Adsorbed protein density (mg/ml)

\(q_0^* \) :

q in equilibrium with C 0 (mg/ml)

q m :

Adsorption capacity in Langmuir isotherm (mg/ml)

\({\bar q}\) :

Adsorbent particle average concentration (mg/ml)

R :

Mean particle radius (m)

r g :

Radius of gyration of proteins (m)

T :

Temperature (K)

t :

Time (min)

u :

Interstitial velocity (cm/h)

V b :

Bed volume (ml)

V L :

Volume of solution (ml)

V S :

Volume of wet gel (ml)

V v :

Bed void volume (ml)

ɛ c :

Column void fraction

ɛ p :

Effective intraparticle porosity for protein

ρ :

Liquid phase density (kg/m2)

μ L :

Viscosity of feed solution (Pa·s)

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Acknowledgments

This work was supported by grants from the National Science and Technology Key Funds (2003DA901A32) and the National Natural Science Foundation (20476085 and 20606026).

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Authors and Affiliations

  1. Department of Chemical Engineering, Northwest University, Xi’an, 710068, China

    Xiao-Jun Wang, Dai-Di Fan & Yan-E Luo

  2. Department of Environmental and Chemical Engineering, Xi’an Polytechnic University, Xi’an, 710048, China

    Xiao-Jun Wang

  3. Key Laboratory of Resource Biology and Biotechnology in Western China, Northwest University, Ministry of Education, Xi’an, 710068, China

    Dai-Di Fan

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  1. Xiao-Jun Wang
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Correspondence to Dai-Di Fan.

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Wang, XJ., Fan, DD. & Luo, YE. Breakthrough Model of Recombinant Human-Like Collagen in Immobilized Metal Affinity Chromatography. Appl Biochem Biotechnol 158, 262–276 (2009). https://doi.org/10.1007/s12010-008-8351-8

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  • DOI: https://doi.org/10.1007/s12010-008-8351-8

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