Adsorption of proteins onto non-soluble polysaccharides matrixes: a friendly strategy to isolate enzymes with potential application for downstream processes


Adsorption of proteins onto non-soluble polysaccharides matrixes: a friendly strategy to isolate enzymes with potential application for downstream processes

Nadia Woitovich Valetti, M. Emilia Brassesc, Guillermo Alfredo Picó

Polysaccharides with electrically charged groups, polyelectrolytes (PE), have the capacity to form hydrogels under different experimental conditions, which act as ionic exchanges with high affinity to adsorb proteins. Presented in this chapter is a description of the current state of the technic on the use of these matrixes. Alginate, carrageenan, chitosan, pectin, etc. are the most used PE to make beds with the capacity of proteins adsorption. The presence of electrically charged residual groups in these PE allows their use as beds for the ion exchange chromatography in stirred tank of packed bed or expended bed. We demonstrated the adsorption of lysozyme, a model protein positively charged, onto Alginate-Guar gum matrixes cross-linked with epichlorohydrin (negatively charged). Their physical characterization, equilibrium isotherms and adsorption kinetics were carried out. Successive cycles of adsorption-washing-elution were performed. The results demonstrate the reversibility of the process and the capacity of this enzyme purification method.

Chromatography, Adsorption, Alg-guar gum, Polyelectrolytes, Bioseparation

Published online 4/25/2017, 23 pages
Copyright © 2016 by the author(s)
Published under license by Materials Research Forum LLC., Millersville PA, USA

Citation: Nadia Woitovich Valetti, M. Emilia Brassesc, Guillermo Alfredo Picó, ‘Adsorption of proteins onto non-soluble polysaccharides matrixes: a friendly strategy to isolate enzymes with potential application for downstream processes’, Materials Research Foundations, Vol. 15, pp 84-106, 2017


The article was published as article 4 of the book Applications of Adsorption and Ion Exchange Chromatography in Waste Water Treatment

[1] R. K. Scopes, Protein purification: principles and practice / Robert K. Scopes, Springer- Verlag: New York, (1994).
[2] M. -S. Chiou, P.-Y. Ho, H.-Y. Li, Adsorption of anionic dyes in acid solutions using chemically cross-linked chitosan beads, Dyes and Pigments, 60 (2004) 69-84.
[3] AL-Othman ZA, Naushad M, Inamuddin (2011) Organic–inorganic type composite cation exchanger poly-o-toluidine Zr(IV) tungstate: Preparation, physicochemical characterization and its analytical application in separation of heavy metals. Chem Eng J 172:369–375.
[4] N. W. Valetti, G. Picó, Adsorption isotherms, kinetics and thermodynamic studies towards understanding the interaction between cross-linked alginate-guar gum matrix and chymotrypsin, Journal of Chromatography B, 1012–1013 (2016) 204-210.
[5] Nabi SA, Bushra R, Naushad M, Khan AM (2010) Synthesis, characterization and analytical applications of a new composite cation exchange material poly-o-toluidine stannic molybdate for the separation of toxic metal ions. Chem Eng J 165:529–536.
[6] T. Gotoh, K. Matsushima, K.-I. Kikuchi, Preparation of alginate–chitosan hybrid gel beads and adsorption of divalent metal ions, Chemosphere 55 (2004) 135-140.
[7] A. R. Kulkarni, K.S. Soppimath, T.M. Aminabhavi, A.M. Dave, M.H. Mehta, Glutaraldehyde crosslinked sodium alginate beads containing liquid pesticide for soil application, Journal of Controlled Release 63 (2000) 97-105.
[8] M. P. Klein, C. R. Hackenhaar, A. S. Lorenzoni, R. C. Rodrigues, T. M. Costa, J. L. Ninow, P. F. Hertz, Chitosan crosslinked with genipin as support matrix for application in food process: Support characterization and β-d-galactosidase immobilization, Carbohydrate Polymers 137 (2016) 184-190.
[9] J. P. Paques, E. van der Linden, C. J. van Rijn, L. M. Sagis, Preparation methods of alginate nanoparticles, Advances in Colloid and Interface Science 209 (2014) 163-171.
[10] D. Spelzini, B. Farruggia, G. Picó, Purification of chymotrypsin from pancreas homogenate by adsorption onto non-soluble alginate beads, Process Biochemistry 46 (2011) 801-805.
[11] K. C. Aguilar, F. Tello, A. C. Bierhalz, M. G. G. Romo, H. E. M. Flores, C. R. Grosso, Protein adsorption onto alginate-pectin microparticles and films produced by ionic gelation, Journal of Food Engineering 154 (2015) 17-24.
[12] K. Y. Lee, D. J. Mooney, Alginate: Properties and biomedical applications, Progress in Polymer Science 37 (2012) 106-126.
[13] K. Dos Santos, J. Coelho, P. Ferreira, I. Pinto, S. G. Lorenzetti, E. Ferreira, O. Z. Higa, M. Gil, Synthesis and characterization of membranes obtained by graft copolymerization of 2-hydroxyethyl methacrylate and acrylic acid onto chitosan, International journal of pharmaceutics 310 (2006) 37-45.
[14] A. G. Sullad, L. S. Manjeshwar, T. M. Aminabhavi, Novel pH-Sensitive Hydrogels Prepared from the Blends of Poly(vinyl alcohol) with Acrylic Acid-graft-Guar Gum Matrixes for Isoniazid Delivery, Industrial & Engineering Chemistry Research 49 (2010) 7323-7329.
[15] T. Gotoh, K. Matsushima, K. -I. Kikuchi, Adsorption of Cu and Mn on covalently cross-linked alginate gel beads, Chemosphere 55 (2004) 57-64.
[16] J. López-Morales, D. Sánchez-Rivera, T. Luna-Pineda, O. Perales-Pérez, F. Román-Velázquez, Entrapment of Tyre Crumb Rubber in Calcium-Alginate Beads for Triclosan Removal, Adsorption Science & Technology 31 (2013) 931-942.
[17] W. W. Ngah, L. Teong, M. Hanafiah, Adsorption of dyes and heavy metal ions by chitosan composites: A review, Carbohydrate Polymers, 83 (2011) 1446-1456.
[18] F. Tello, R. N. Falfan-Cortés, F. Martinez-Bustos, V. M. da Silva, M. D. Hubinger, C. Grosso, Alginate and pectin-based particles coated with globular proteins: Production, characterization and anti-oxidative properties, Food Hydrocolloids 43 (2015) 670-678.
[19] E. Rodrigues, B. Bezerra, B. Farias, W. Adriano, R. Vieira, D. Azevedo, I. Silva, Adsorption of Cellulase Isolated from Aspergillus niger on Chitosan/Alginate Particles Functionalized with Epichlorohydrin, Adsorption Science & Technology 31 (2013) 17-34.
[20] D. R. Gondim, N. A. Dias, I. T. Bresolin, A. M. Baptistiolli, D. C. Azevedo, I. J. Silva Jr, Human IgG adsorption using dye-ligand epoxy chitosan/alginate as adsorbent: influence of buffer system, Adsorption 20 (2014) 925-934.
[21] X. Vecino, R. Devesa-Rey, J. Cruz, A. Moldes, Study of the physical properties of calcium alginate hydrogel beads containing vineyard pruning waste for dye removal, Carbohydrate Polymers 115 (2015) 129.
[22] Z. Zhang, R. Zhang, L. Zou, D. J. McClements, Protein encapsulation in alginate hydrogel beads: Effect of pH on microgel stability, protein retention and protein release, Food Hydrocolloids 58 (2016) 308-315.
[23] J. Venkatesan, I. Bhatnagar, P. Manivasagan, K. -H. Kang, S. -K. Kim, Alginate composites for bone tissue engineering: A review, International Journal of Biological Macromolecules 72 (2015) 269-281.
[24] A. S. Hoffman, Hydrogels for biomedical applications, Advanced Drug Delivery Reviews 64 (2012) 18-23.
[25] J. Patil, M. Kamalapur, S. Marapur, D. Kadam, Ionotropic gelation and polyelectrolyte complexation: the novel techniques to design hydrogel particulate sustained, modulated drug delivery system: A Review, Digest Journal of Nanomaterials and Biostructures 5 (2010) 241-248.
[26] J. Yang, J. Chen, D. Pan, Y. Wan, Z. Wang, pH-sensitive interpenetrating network hydrogels based on chitosan derivatives and alginate for oral drug delivery, Carbohydrate Polymers 92 (2013) 719-725.
[27] M. George, T. E. Abraham, pH sensitive alginate–guar gum hydrogel for the controlled delivery of protein drugs, International Journal of Pharmaceutics 335 (335) 123-129.
[28] N. Fiol, I. Villaescusa, Determination of sorbent point zero charge: usefulness in sorption studies, Environmental Chemistry Letters 7 (2009) 79-84.
[29] E. Daneshvar, M. Kousha, M. Jokar, N. Koutahzadeh, E. Guibal, Acidic dye biosorption onto marine brown macroalgae: Isotherms, kinetic and thermodynamic studies, Chemical Engineering Journal 204 (2012) 225-234.
[30] C. Zhou, Q. Wu, T. Lei, I. I. Negulescu, Adsorption kinetic and equilibrium studies for methylene blue dye by partially hydrolyzed polyacrylamide/cellulose nanocrystal nanocomposite hydrogels, Chemical Engineering Journal 251 (2014) 17-24.
[31] S. -J. Yoon, D. -C. Chu, L. Raj Juneja, Chemical and Physical Properties, Safety and Application of Partially Hydrolized Guar Gum as Dietary Fiber , Journal of Clinical Biochemistry and Nutrition 42 (2008) 1-7.
[32] M. Doğan, Y. Özdemir, M. Alkan, Adsorption kinetics and mechanism of cationic methyl violet and methylene blue dyes onto sepiolite, Dyes and Pigments 75 (2007) 701-713.
[33] K. Mazur, R. Buchner, M. Bonn, J. Hunger, Hydration of Sodium Alginate in Aqueous Solution, Macromolecules 47 (2014) 771-776.
[34] J. C. Lutter, T. -Y. Wu, Y. Zhang, Hydration of Cations: A Key to Understanding of Specific Cation Effects on Aggregation Behaviors of PEO-PPO-PEO Triblock Copolymers, The Journal of Physical Chemistry B 117 (2013) 10132-10141.
[35] Y. Zhang, P. S. Cremer, Interactions between macromolecules and ions: the Hofmeister series, Current Opinion in Chemical Biology 10 (2006) 658-663.
[36] T. Oncsik, G. Trefalt, M. Borkovec, I. Szilagyi, Specific Ion Effects on Particle Aggregation Induced by Monovalent Salts within the Hofmeister Series, Langmuir 31 (2015) 3799-3807.
[37] S. Rangabhashiyam, N. Anu, M. G. Nandagopal, N. Selvaraju, Relevance of isotherm models in biosorption of pollutants by agricultural byproducts , Journal of Environmental Chemical Engineering 2 (2014) 398-414.
[38] V. Vadivelan, K. V. Kumar, Equilibrium, kinetics, mechanism, and process design for the sorption of methylene blue onto rice husk, Journal of Colloid and Interface Science 286 (2005) 90-100.
[39] G. P. Jeppu, T. P. Clement, A modified Langmuir-Freundlich isotherm model for simulating pH-dependent adsorption effects, Journal of contaminant hydrology 129 (2012) 46-53.
[40] A. Crhribi, M. Chlendi, Modeling of Fixed Bed Adsorption: Application to the Adsorption of an Organic Dye, Asian J. Textile 1 (2011) 161-171.
[41] H. Nouri, A. Ouederni, Modeling of the Dynamics Adsorption of Phenol from an Aqueous Solution on Activated Carbon Produced from Olive Stones, Journal of Chemical Engineering & Process Technology, 4 (3) (2013). (DOI: 10.4172/2157-7048.1000153)
[42] B. Zhao, W. Xiao, Y. Shang, H. Zhu, R. Han, Adsorption of light green anionic dye using cationic surfactant-modified peanut husk in batch mode, Arabian Journal of Chemistry, (2014). (