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Brazilian Archives of Biology and Technology
On-line version ISSN 1678-4324
PADILHA, Giovana da Silva; SANTANA, José Carlos Curvelo; ALEGRE, Ranulfo Monte and TAMBOURGI, Elias Basile. Extraction of lipase from Burkholderia cepacia by PEG/Phosphate ATPS and its biochemical characterization. Braz. arch. biol. technol. [online]. 2012, vol.55, n.1, pp.7-19. ISSN 1678-4324. http://dx.doi.org/10.1590/S1516-89132012000100002.
This work aimed to study the partitioning of a lipase produced by Burkholderia cepacia in PEG/Phosphate aqueous two phase system (ATPS) and its characterization. Lipase was produced by B. cepacia strains in a fermenter. Enzyme partitioning occurred at pH 6.0 and 8.0, using PEG 1500 and 6000 on two tie lines. Metal ions, pH and temperature effects on enzyme activity were evaluated. Five milliliter of 7.5% olive oil emulsion with 2.5% gumarabic in 0.1M sodium phosphate buffer at pH 8.0 and 37ºC were used for the activity determinations. Results showed that crude stratum from B. cepacia was partitioned by PEG1500/phosphate ATPS at pH 6.0 or 8.0 for, which the partitioning coefficients were 108-and 209-folds. Lipase presented optimal activity conditions at 37ºC and pH 8.0; it showed pH-stability for 4 h of incubation at different pH values at 37ºC. Metal ions such as Mn2+ , Co2+, I-and Ca2+ sustained enzymatic activities; however, it was inhibited by the presence of Fe2+, Hg2+ and Al3+ . Km and Vmax values were 0.258 U/mg and 43.90 g/L, respectively. A molecular weight of 33 kDa and an isoelectric point at pH 5.0 were determined by SDS-PAGE and IFS electrophoresis, respectively.
Keywords : lipase; Burkholderia cepacia; aqueous two phase system; partitioning; characterization.