Maximization of Intracellular Lipase Production in a Lipase-Overproducing Mutant Derivative of Rhizopus oligosporus DGM 31: A Kinetic Study

Tehreema Iftikhar1,2, Mubashir Niaz2, Munazza Afzal3, Ikram-ul-Haq1 and Muhammad Ibrahim Rajoka3*

1Institute of Industrial Biotechnology, Department of Botany, GC University, Lahore, Pakistan

Department of Botany, GC University, Faisalabad, Pakistan

3National Institute for Biotechnology and Genetic Engineering, P.O. Box 577, Jhang Road, Faisalabad, Pakistan

Article history:

Received July 18, 2007

Accepted April 30, 2008

Key words:

lipases, Rhizopus oligosporus, oils, casein, ammonium nitrate, ammonium oxalate, starch, glycerol


Regulation and maximization of lipase production in a mutant derivative of R. oligosporus has been investigated using different substrates, inoculum sizes, pH of the medium, temperature, and nitrogen sources in shake flask experiments and batch fermentation in a fermentor. The production of intracellular lipase was improved 3 times following medium optimization involving one-at-a-time approach and aeration in the fermentor. Interestingly, intracellular lipase was poorly induced by oils, instead its production was induced by sugars, mainly starch, lactose, sucrose, xylose, glucose and glycerol. Dependent variables studied were cell mass, lipase activity, lipase yield, lipase specific and volumetric rate of formation. It was confirmed that lipase production in the derepressed mutant is sufficiently uncoupled from catabolite repression. The results of average specific productivities at various temperatures worked out according to the Arrhenius equation revealed that mutation decreased the magnitude of enthalpy and entropy demand in the inactivation equilibrium during product formation, suggesting that mutation made the metabolic network of the organism thermally more stable. The highest magnitudes of volumetric productivity (QP=490 IU/(L·h)) and other product attributes of lipase formation occurring on optimized medium in the fermentor are greater than the values reported by other workers. The purified enzyme is monomeric in nature and exhibits stability up to 80 °C and pH=6.0–8.0. Activation energy, enthalpy and entropy of catalysis at 50 °C, and magnitudes of Gibbs free energy for substrate binding, transition state stabilization and melting point indicated that this lipase is highly thermostable.


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