Immobilization of Aspergillus oryzae β galactosidase on concanavalin A-layered calcium alginate-cellulose beads and its application in lactose hydrolysis in continuous spiral bed reactors

Ansari, Shakeel; Husain, Qayyum

doi:10.2478/v10026-011-0043-4

Abstract

In this study, Aspergillus oryzae β galactosidase was immobilized on concanavalin A layered calcium alginate-cellulose beads as a bioaffinity support. Immobilized enzyme showed a remarkable broadening in temperature-activity profiles as compared to the native enzyme and exhibited 65% activity in the presence of 5% galactose. Michaelis constant (K_m) was 2.57 mM and 5.38 mM for the free and the immobilized β galactosidase, respectively. Crosslinked β galactosidase showed greater catalytic activity in the presence of Mg²⁺ and was more stable during storage at 4°C for 6 weeks. Immobilized enzyme hydrolyzed 67% lactose in milk in 8 h and 85% lactose in whey in 9 h in the stirred batch process at 50°C. The continuous hydrolysis of lactose by crosslinked β galactosidase in spiral bed reactor exhibited 93% and 88% hydrolysis of lactose at flow rate of 20 ml/h and 30 ml/h, after 1 month operation, respectively.

References

Zhang, S., Gao, S & Gao, G. (2010). Immobilization of β galactosidase onto magnetic beads. Appl. Biochem. Biotechnol. 160, 1386-1393. DOI: 10.1007/s12010-009-8600-5.10.1007/s12010-009-8600-5
Search in Google Scholar
Heyman, B. (2006). Lactose Intolerance in infants, children and adolescents. Pediatrics 118, 1279-1286. DOI: 10.1542/peds.2006-1721.10.1542/peds.2006-1721
Search in Google Scholar
Ansari, S.A. & Husain, Q. (2010). Lactose hydrolysis by β galactosidase immobilized on concanavalin A-cellulose in batch and continuous mode. J. Molec. Catal. B: Enzym. 6, 68-74. DOI: 10.1016/j.molcatb.2009.12.010.10.1016/j.molcatb.2009.12.010
Search in Google Scholar
Mateo, C., Palomo, J.M., Fernandez-Lorente, G., Guisan, J.M & Fernandez-Lafuente, R (2007). Improvement of enzyme activity, stability and selectivity via immobilization techniques. Enzyme Microb. Technol. 40, 1451-1463. DOI: 10.1016/j.enzmictec.2007.01.018.10.1016/j.enzmictec.2007.01.018
Search in Google Scholar
Ansari, S.A. & Husain, Q. (2011). Immobilization of Kluyveromyces lactis β galactosidase on concanavalin A layered Al₂O₃ nanoparticles-its application in biosensors. J. Molec. Catal. B: Enzym. 70, 119-126. DOI: 10.1016/j.molcatb.2011.02.016.10.1016/j.molcatb.2011.02.016
Search in Google Scholar
Husain, Q. (2010). β Galactosidases and their potential applications. Crit. Rev. Biotechnol. 30, 41-62. DOI: 10.4061/2010/473137.10.4061/2010/473137
Search in Google Scholar
Betancor, L., Luckarift, R., Seo, H., Brand, O. & Spain, J.C. (2008). Three-dimensional immobilization of β galactosidase on a silicon surface. Biotechnol. Bioeng. 99, 261-267. DOI: 10.1002/bit.21570.10.1002/bit.21570
Search in Google Scholar
Sun, S., Dong, L., Xu, X & Shen, S. (2010). Immobilization of β galactosidase from Aspergillus oryzae on macroporous poly GMA newly prepared. Int. J. Chem. 2, 89-96. DOI: 10.1016/ijc.2010.04.011.
Search in Google Scholar
Gurdas, S., Gulec, H.A. & Mutlu, M. (2010). Immobilization of Aspergillus oryzae β galactosidase onto Duolite A568 resin via simple adsorption mechanism. Food Bioproc. Technol. DOI 10.1007/s11947-010-0384-7.
Search in Google Scholar
Ansari, S.A., Husain, Q., Qayyum, S. & Azam, A. (2011) Designing and surface modification of zinc oxide nanoparticles for biomedical applications. Food Chem. Toxicol. 2011 Sep;49(9):2107-15.10.1016/j.fct.2011.05.025
Search in Google Scholar
Husain, Q., Ansari, S.A., Alam, F. & Azam, A. (2011) Immobilization of Aspergillus oryzae β galactosidase on zinc oxide nanoparticles via simple adsorption mechanism. Int. J. Biol. Macromol. 49(1):37-43. DOI: 10.1016/j.ijbiomac.2011.03.011.10.1016/j.ijbiomac.2011.03.011
Search in Google Scholar
Bradford, M.M. (1976). A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248-255. DOI: 10.1016/S0076-6879(09)63008-1.10.1016/S0076-6879(09)63008-1
Search in Google Scholar
Lee, C.H., Oh, E., Kim, J. & Ahn, D.J. (2008). Immobilization of polydiacetylene vesicles on cellulose acetate butyrate (CAB)-coated substrates for self-assembled supramolecular sensor arrays. Coll. Surf. A: Physicochem. Eng. Aspects. 313, 500-503. DOI: 10.1016/j.colsurfa.2007.04.174.10.1016/j.colsurfa.2007.04.174
Search in Google Scholar
Prashanth, S.J. & Mulimani, V.H. (2005). Soymilk oligosaccharides hydrolysis by Aspergillus oryzae β galactosidase im mobilized in calcium alginate. Process Biochem. 40, 1119-1205. DOI: 10.1016/j.procbio.2004.04.011.10.1016/j.procbio.2004.04.011
Search in Google Scholar
Smidsrod, O & Skjak-Braek, G. (1990). Alginate as immobilization matrix for cells. Trends Biotechnol. 8, 71-78. DOI: 10.1016/0167-7799(90)90139-O.10.1016/0167-7799(90)90139-O
Search in Google Scholar
Hennink, W.E. & Nostrum, C.F.V. (2002). Novel crosslinking methods to design hydrogels. Adv. Drug. Deliv. 54, 13-36. DOI: 10.1016/S0169-409X(01)00240-X.10.1016/S0169-409X(01)00240-X
Search in Google Scholar
Musthapa, S.M., Akhtar, S., Khan, A.A. & Husain, Q. (2004). An economical, simple and high yield procedure for the immobilization/stabilization of peroxidases from turnip roots. J. Sc. Ind. Res. 63, 540-547. DOI: 10.1016/j.bioeng.2006.07.004.10.1016/j.bioeng.2006.07.004
Search in Google Scholar
Elnashar, M.M.M. & Yassin, M.A. (2009). Lactose hydrolysis by β galactosidase covalently immobilized to thermally stable biopolymers. Appl. Biochem. Biotechnol. 159, 426-37. DOI: 10.1007/s12010-008-8453-3.10.1007/s12010-008-8453-3
Search in Google Scholar
Bayramoglu, G., Tunali, Y. & Arica, M.Y. (2007). Immobilization of β galactosidase onto magnetic poly (GMAMMA) beads for hydrolysis of lactose in bed reactor. Catalys. Commun. 8, 1094-1101. DOI: 10.1016/j.catcom.2006.10.029.10.1016/j.catcom.2006.10.029
Search in Google Scholar
Ozdural, A.R., Tanyolac, D., Boyac, I.H. Mutlu, M & Webb, C. (2003). Determination of apparent kinetic parameters for competitive product inhibition in packed-bed immobilized reactor. Biochem. Eng. J. 14, 27-36. DOI: 10.1016/S1369-703X(02)00099-2.10.1016/S1369-703X(02)00099-2
Search in Google Scholar
Zhou, Q.Z., Chen, X.D & Li, X. (2003). Kinetics of lactose hydrolysis by β galactosidase of Kluyveromyces lactis immobilized on cotton. Biotechnol. Bioeng. 81, 127-133. DOI: 10.1002/bit.10414.10.1002/bit.1041412451549
Search in Google Scholar
Sutendra, G., Wong, S., Fraser, M.E & Huber, R.E. (2007). β galactosidase (Escherichia coli) has a second catalytically important Mg₊₂ sites. Biochem. Biophys. Res. Comm. 352, 566-570. DOI: 10.1016/j.bbrc.2006.11.061.10.1016/j.bbrc.2006.11.06117126292
Search in Google Scholar
Gaur, R., Pant, H., Jain, R. & Khare, S.K. (2006) Galacto-oligosaccharide synthesis by immobilized Aspergillus oryzae β galactosidase. Food Chem. 97, 426-430. DOI: 10.1016/j.foodchem.2005.05.020.10.1016/j.foodchem.2005.05.020
Search in Google Scholar
Haider, T. & Husain, Q. (2008). Hydrolysis of milk/whey lactose by β galactosidase: a comparative study of stirred batch process and packed bed reactor prepared with calcium alginate entrapped enzyme. Chem. Eng. Proc.: Process Intens. 48, 576-580. DOI: 10.1016/j.cep.2008.02.007.10.1016/j.cep.2008.02.007
Search in Google Scholar
Diwedi, A. & Kayastha, A.M. (2009). Stabilization of β galactosidase (from peas) by immobilization onto amberlite MB-150 beads and its application in lactose hydrolysis. J. Agric. Food Chem. 57, 682-688. DOI: 10.1021/jf802573j.10.1021/jf802573j
Search in Google Scholar
Novalin, S., Neuhaus, W. & Kulbe, K.D. (2005). A new innovative process to produce lactose reduced skim milk. J. Biotechnol. 119, 212-218. DOI: 10.1016/j.jbiotec.2005.03.018.10.1016/j.jbiotec.2005.03.018
Search in Google Scholar
Li, X., Zhou, Q.Z.K. & Chena, X.D. (2007). Pilot scale lactose hydrolysis using β galactosidase immobilized on cotton fabric. Chem. Eng. Proc. 46, 497-500. DOI: 10.1016/j.cep.2006.02.011.10.1016/j.cep.2006.02.011
Search in Google Scholar
Tanriseven, A & Dogan, S. (2002). A novel method for the immobilization of β galactosidase. Process Biochem. 38, 27-30. DOI: 10.1016/S0032-9592(02)00049-3.10.1016/S0032-9592(02)00049-3
Search in Google Scholar
Roy, I. & Gupta, M.N. (2003). Lactose hydrolysis by lactozym immobilized on cellulose beads in batch and fluidized modes. Process Biochem. 39, 325-332. DOI:10.1016/S0032-9592(03)00086-4.10.1016/S0032-9592(03)00086-4
Search in Google Scholar
Mammarella, E.J. & Rubiolo, A.C (2006). Predicting the packed bed reactor performance with immobilized microbial lactase. Process Biochem. 41, 1627-1636. DOI: 10.1016/j.procbio.2006.03.009.10.1016/j.procbio.2006.03.009
Search in Google Scholar

Immobilization of Aspergillus oryzae β galactosidase on concanavalin A-layered calcium alginate-cellulose beads and its application in lactose hydrolysis in continuous spiral bed reactors

Abstract

Paradigm

My account