Investigating Culture Media for Obtaining Lipolytic Biocatalysts Based on Rhizopus oryzae Fungi

Abstract

: Rhizopus oryzae is widely distributed in nature and can be isolated from different sub-strates such as decomposing vegetables, fruits and various soils. It is generally classified as GRAS filamentous fungi and commonly used in the production of oriental traditional food such as tempeh or peka. This microorganism has a great industrial potential due to the capability to synthesize enzymes (glucoamylases, cellulases and lipases) and organic acids (lactic acid, fumaric acid). The most studied enzymes of the fungi are lipases (ROL). Therefore, the aim of the study was the selection of growth medium content and initial pH rate, which would provide high lipase synthesis yield in 5 days shaken cultures. Two fractions of lipases were investigated in order to obtain lipase biocatalysts: extracellular enzymes present in supernatant and cell-bound lipases in biomass. There were used nutrient-rich media: YPG (10 g/L yeast extract, 20 g/L peptone, 20 g/L glucose), YPO (10 g/L yeast extract, 20 g/L peptone, 20 g/L olive oil), YMG (3 g/L yeast extract, 3 g/L malt extract, 5 g/L peptone, 20 g/L glucose), YMO (3 g/L yeast extract, 3 g/L malt extract, 5 g/L peptone, 20 g/L olive oil) and mineral media: SMG (10 g/L peptone, 14 g/L KH 2 PO 4 , 2.4 g/L K 2 HPO 4 , 0.4 g/L MgSO 4 , 20 g/L glucose) and SMO (10 g/L peptone, 14 g/L KH 2 PO 4 , 2.4 g/L K 2 HPO 4 , 0.4 g/L MgSO 4 , 20 g/L olive oil). Fungi biomass and supernatant were separated and used to measure lipase activity by a spectrophotometric method based on the hydrolysis of p -nitrophenyl laurate. The results showed that the highest lipase activity after 5 days of cultivation was reached in YPO medium for biomass (from 7- to 60-fold higher results depending on compared variant of culture media) and YMG for supernatant (from 3- to 6.5-fold higher results depending on used variant of culture media). The addition of citric acid resulted in two times increase of the activity of produced lipases after 5 days of cultivation.