| Congresso Brasileiro de Microbiologia 2023 | Resumo: 621-1 | ||||
Resumo:L-asparaginase is an important enzyme in the treatment of Acute Lymphoid Leukemia due to its ability to hydrolyze the amino acid L-asparagine. The clinical preparations of this enzyme are derived from a bacterial source and its use is often associated with serious adverse reactions. Therefore, the search for new sources of L-asparaginase is relevant. Enzyme extraction is an indispensable step in enzyme processing. The aim of this work was to perform a screening of the enzymatic extraction methods of L-asparaginase by P. cerradense. The fungus is a new isolate from Cerrado soil. The culture medium was prepared according to modified Czapek Dox (L-proline 1.71 %, L-asparagine 1.38 %, NaNO3 1.99 %, glucose 0. 65 %, KH2PO4.7H2O 0.0152 %, KCl 0.52 %, MgSO4.7H2O 0.52 %, traces of CuNO3.3H2O, ZnSO4.7H2O, FeSO4.7H2O), 50 mL for four days, 120 rpm and 30 °C. Three mechanical methods were evaluated for enzyme release comparing activity in the biomass: i) maceration of the frozen biomass with the aid of a mortar and pestle until a fine powder was obtained for cell suspension (1:2 m/v); ii) agitation with glass beads in cell suspension (1:3 m/v), five cycles 60:45 s (pulse/no pulse); and iii) by using an ultrasonic Bath Sonicator 750 W/20 kHz/amplitude 40% with cell suspension (1:20 m/v), five cycles 60:30 s (pulse/no pulse). L-asparaginase activity was assayed according to β-hydroxamate aspartate. The results obtained showed that the use of glass bead agitation (0.49 U/gcell) was more effective in the L-asparaginase enzyme extraction from biomass than maceration (0.09 U/gcell) and sonicator (0.03 U/gcell) methods. Future studies will be conducted to optimize the L-asparaginase enzyme extraction method by agitation with glass beads. This work presented the most effective method of enzyme extraction for L-asparaginase from P. cerradense which can support the improvement of this enzyme production. Palavras-chave: L-asparaginase, Penicillium cerradense, Leukemia, Enzyme extraction Agência de fomento:FAPDF, FAPESP and CAPES | |||||