1.2.3. Biosurfactant production
Initially, ICA56 was inoculated into APGE plates and in cubatedat 30◦C for 24 h. After this period, three colonies were transferred to 250 mL Erlenmeyer flasks containing 50 mL of Mineral medium(MM) and 0.1% of trace element solution [14]. The flasks wereincubated in a rotary shaker (Tecnal-TE240, Sao Paulo, Brazil) at150 rpm, 30◦C for 24 h. The optical density (OD) was adjusted withsterile MM to 0.15 at 600 nm, in order to assure the same cell con-centration in the seed culture [12].Biosurfactant production was conducted in 250 mL Erlenmeyerflasks containing 100 mL of MM. An aliquot of 10 mL (10%, v/v) ofthe seed culture was transferred to the Erlenmeyer flask and theexperiments were carried out in a rotary shaker (Tecnal-TE240) at150 rpm, 30◦C. In order to evaluate unconventional carbon sourcesfor biosurfactant production, the assay was conducted for 48 h andsamples were withdrawn at the end of cultivation. Afterwards, themost promising carbon source was selected in order to evaluatethe influence of cultivation time and abiotic variables on biosur-factant production. In this case, the assay was conducted for 72 hand samples were collected at defined intervals of time. Then, cellswere removed by centrifugation at 10,000 × g for 20 min and thecell-free supernatant was assayed to measure pH, substrate andbiosurfactant concentrations [12]. During cultivation of ICA56, theanalysis of surface tension of the cell-free fermented broth was al