Isolation and characterization of organic-solvent tolerant bacteria

Abstract

Screening and isolation of 19 samples collected from Thailand contaminated soil yield 130 bacteria isolates. Among them isolates A102, A103 and B202 isolated from engine oil contaminated soil at Burirum province and P2 and P3 isolated from fuel oil in Bangkok province which were subsequently classified in the genus Enterobacter and Burkholderia are the best biosurfactant producer. Thus, two isolates grew in mineral salt medium containing glucose as a carbon source since partially purified biosurfactants were extracted from these isolates. The chemical structures elucidated by FTIR and NMR analysis of partially purified products upon cultivation in broth were composed of O-H stretch, ester bond and hydrocarbon chain. Results from MS indicated that the compound possesses mass estimated to be 550 m/z for both isolates. Analysis of the glycone fraction by TLC revealed one major component with Rf value 0.23 ± 0.036 and 0.23 ± 0.045, respectively. These were found near that of glucose standard. The biosurfactant of Enterobacter sp. and Burkholderia cepacia could lower surface tension down to 26.0 ± 0.52 and 25.0 ± 0.52 mN.m-1 with CMC values of 3.3 and 1,995 mg.l-1. They showed a maximum emulsion index (E24) of 88.88 ± 0.77% and 88.71 ± 0.58%, respectively for diesel oil with emulsion stability for 5 months at 37˚C. The products were stable at 75˚C for 5 hours. Moreover, the best biosurfactant stability of Enterobacter sp. and Burkholderia cepacia occurred at pH 7. Optimum condition for cultivation of Enterobacter sp. and Burkholderia cepacia to give high yield of biosurfactant was obtained by the use of 44.4 mM glucose as carbon source and 75.0 mM NaNO3 as nitrogen source. Additionally, 2% vv-1 olive oil and sunflower oil increased the production of biosurfactant for Burkholderia cepacia but not for Enterobacter sp. Furthermore, the effects of temperature and concentration of salt were studied in these isolates. The results showed that optimum production of biosurfactant occurred at 37˚C and salt was inhibitory to biosurfactant production.