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Chitosan-alginate immobilized lipase based catalytic constructs: Development, characterization and potential applications
- Rashid, Robina, Anwar, Zahid, Zafar, Muddassar, Rashid, Tayyba, Butt, Iqra
- International journal of biological macromolecules 2018 v.119 pp. 992-1001
- Aspergillus, alginates, ammonium sulfate, carboxylic ester hydrolases, chitosan, cross-linking reagents, dehairing, detergents, fractionation, gel chromatography, gels, glutaraldehyde, leather, metal ions, molecular weight, pH, polyacrylamide gel electrophoresis, tanneries, temperature
- In this study, lipase (LIP) was isolated from Aspergillus crevinus, statistically optimized and purified via ammonium sulfate fractionation (ASF), and Sephadex G-100 gel permeation chromatography. LIP was 2.26-folds purified with a specific activity of 223.60 U/mg. The molecular mass was estimated to be 60 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (5% stacking and a 12% resolving gel) (SDS-PAGE). The active LIP fraction was immobilized onto chitosan-alginate (CTS-ALG) beads developed in a uniform size, i.e., 2.0 ± 0.25 mm diameter using ultrasonically dispersed 2.0% (w/v) chitosan and alginate along with 0.5% (w/v) glutaraldehyde as a macromolecular crosslinking agent. Prior to exploit for detergent compatibility and dehairing purposes, various parameters including pH, thermal, Michaelis-Menten kinetic constants and influence of organic/inorganic and metal ions on PF-LIP and CST-ALG-LIPs fractions were investigated. The immobilized fractions were optimally active and stable over a broader pH (5–11) and temperature (75 °C) as compared to the free counterpart pH (7–8) and temperature (35 °C), respectively. However, the negligible difference between the KM and Vmax values of PF-LIP i.e., 0.133 ± 0.05 mg/mL and 255.0 ± 11.8 U/mL/min and CST-ALG-LIPs revealed that the conformational flexibility of LIP was retained as such. Comparative to PF-LIP, the CTS-ALG-LIPs were found much stable and retained most of their activity up to 80% in the presence of inhibitory molecules. After 75 min incubation, CTS-ALG-LIP3 retained >95% activity at pH 9.0 which was reduced to 80% at pH 10.0 and 44% at pH 11.0. Among all three samples, 100% dehairing was observed when the sheepskin was dipped for 30 min in CTS-ALG-LIP3. The dehairing of leather (sheepskin) was greatly affected by CTS-ALG-LIP3 rendering its potential candidatures for leather and tannery industry.