Purification,Characterization And Its Application Research Of Biosurfactants Produced By A Bacillus Sp.ZG0427

Biosurfactants produced by microorganisms are attracting attention and widely used in petroleum chemical industry,medicine and health care,food processing,washing industry and environmental governance for their several advantages such as good surface activity,low toxicity and good biodegradability.In this paper,we extracted the biosurfactants from Bacillus sp.ZG0427 and studied their physical and chemical properties.The bacterial cells of Bacillus sp.ZG0427 and the extracted biosurfactants were added into the crude oil media with oil-degradation bacteria Pseudomonas sp.YM15,Rhodococcus erythropolis KB1 and Rhodococcus ruber JC106 respectively to study their effect on the oil degrading bacteria.The antimicrobial activities of the extracted biosurfactant were also studied.The main research results are as follows:The biosurfactant was partially purified from the supernatant of the bacterial culture by acid precipitation method,with the product yield of 2.91 g/L.The emulsification of the biosurfactant was also determined.The results showed that the emulsifying index reached 58.0% when the biosurfactant solution concentration was100 mg/L.Its properties were determined by thin layer chromatography and electric property determination.The biosurfactant was preliminary identified as a lipopeptide anionic biosurfactant.The biosurfactant still showed stable emulsification after being kept for 36 h.By using preparation-silicone plate,we separated out six components.High performance liquid chromatography(HPLC)was used to further purify the third major component,the purified component could reduce the surface tension of distilled water to 25.4 m N/m.Its properties were further studied by Fourier transform infrared spectroscopy(FT-IR),and were finally determined as a kind of lipopeptid,which containing several amino acids such as Glu,Ala,Gly,Val,Leu and Asp.In order to study the effect of the biosurfactant on oil degrading bacteria,the bacterial cells of Bacillus sp.ZG0427 were added into the media of oil-degradation bacteria Pseudomonas sp.YM15,Rhodococcus erythropolis KB1 and Rhodococcus ruber JC106 respectively.It was found that when Bacillus sp.ZG0427 and Pseudomonas sp.YM15 co-cultured for seven days,the oil degradation rate increased from 32.70% to 39.33%,with a increasing rate of 6.63%.When Bacillus sp.ZG0427 and Rhodococcus erythropolis KB1 were co-cultured,the degradation rate increased from 19.98% to 34.33%,with a increasing of 14.35%.When Bacillus sp.ZG0427 were co-cultured with Rhodococcus ruber JC106,the ZG0427 inhibited the growth of JC106 in the early time,but the colony count is growing fast in the third day,and the oil degradation rate increased from 25.87% to 38.93%,with a increasing rate of13.06%.When added into the media of oil-degradation bacteria Pseudomonas sp.YM15 with a concentration of 20mg/L,the extracted biosurfactants promoted growth of YM15,and enhanced the oil degradation rate by 12.53%.But same concentration of the biosurfactant inhibited growth of R.erythropolis KB1 and the degradation rate was reduced by 7.62%.When the extracted biosurfactant was added into the media of oil-degradation bacteria R.ruber JC106,the degradation rate was reduced by 20.10%and the bacteria cells growth was seriously inhibited the phenomenon of cell death was observed.In a certain range of concentrations,the effects of the biosurfactant on Pseudomonas sp.YM15 were positively correlated with the concentrations.The extracted biosurfactants inhibited growth of R.erythropolis KB1 and R.ruber JC106 to some extent,resulted in the decreasing of oil degradation rates.The Oxford cup method was used to study the bacteriostatic activities of the extracted biosurfactant.The results showed that the extracted biosurfactant has obvious bacteriostatic effect on some Gram-positive bacteria,such as R ruber JC106,R.KB1,Cellulomonas sp YM40,and some Gram-negative bacteria,such as E.coli,Serratia sp.YM19,Vibrio alginolyticus,V.harveyi,V.anguillarum.The minimum dilution inhibitor effect concentrations against R.ruber JC106,S.sp.YM19,V.alginolyticus and V.harveyi were 0.20 g/L,0.04g/L,0.008g/L and 0.008g/L respectively.But biosurfactants did not show obvious inhibitory activity against Pseudomonas sp.YM15 and some fungi.