Issue
Korean Journal of Chemical Engineering,
Vol.28, No.3, 867-874, 2011
Immobilization and stabilization of Pseudomonas aeruginosa SRT9 lipase on tri(4-formyl phenoxy) cyanurate
Lipase was extracted and purified from Pseudomonas aeruginosa SRT9. Culture conditions were optimized and highest lipase production amounting to 147.36 U/ml was obtained after 20 h incubation. The extracellular lipase was purified on Mono QHR5/5 column, resulting in a purification factor of 98-fold with specific activity of 12307.81U/mg. Lipase was immobilized on tri (4-formyl phenoxy) cyanurate to form Schiff’s base. An immobilization yield of 85% was obtained. The native and immobilized lipases were used for catalyzing the hydrolysis of olive oil in aqueous medium. Comparative study revealed that immobilized lipase exhibited a shift in optimal pH from 6.9 (free lipase) to 7.5 and shift in optimal temperature from 55 ℃ to 70 ℃. The immobilized lipase showed 20-25% increase in thermal stability and retained 75% of its initial activity after 7 cycles. It showed good stability in organic solvents especially in 30% acetone and methanol. Enzyme activity was decreased by ~60% when incubated with 30% butanol. The kinetic studies revealed increase in KM value from 0.043mM (native) to 0.10 mM for immobilized lipase. It showed decrease in the Vmax of immobilized enzyme (142.8 μmol min.1 mg.1), suggesting enzyme activity decrease in the course of covalent binding. The immobilized lipase retained its initial activity for more than 30 days when stored at 4 ℃ in Tris-HCl buffer pH 7.0 without any significant loss in enzyme activity.
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