Study On Biodegradation Of Oil Biomarkers And Their Application On Quantitative Evaluation For Spilled-Oil Bioremediation

With the growing demand of petroleum and its refined products,accidental oil spills occur on a frequent basis during operations of extraction, transportation, storage, refining and distribution. Oil spills posed great threats and cause extensive damage to marine coastal environment, ecology, resource and human health. Bioremediation has become a promising technology to deal with shoreline environmental oil pollutants, due to its several advantages, such as effective, low cost and ecological sound. The composition, property and concentration of oil spilled into the marine may have a great change under in the various actions of wind, waves, currents, light, temperature and biological activity. Therefore, oil contaminated sites are often highly heterogeneous, where oil concentrations can vary greatly within a small area. This will lead to the evaluation of oil biodegradation becomes a difficult task.The research focuses on establishing quantitative evaluation method for oil-polluted bioremediation effect. Firstly, the biodegradation of biomarkers, such as pristane/phytane, hopanes, alkyl-PAHs have been studied by lab and field experiment. Secondly, appropriate biomarkers have been selected to evaluate oil bioremediation in the field study. Finally, the method based on biomarkers has been applied in the field oil-spilled shoreline bioremediation to confirm the effectiveness of bioaugmentation and biostimulation. Biodegradation are tracked by GC-MS analysis of selected components, and the measured concentrations are corrected for abiotic removal by hopane normalization. Four treatments were evaluated: a no-nutrient addition control, addition of water-soluble nutrients, and addition of slow release fertilizer nutrients, and addition of slow release fertilizer nutrients with oil-degrading microbial inoculums. The results not only have great theoretical significance in scientific evaluation of the bioremediation effect, but also in promoting wide applications of bioremediation technology in different oil-contaminated coastlines. The main results of this research are shown as follows:1) To determine several biomarker groups (include pristane/phytane, hopanes and steranes, alkyl-PAH) which are more susceptible or resistant to biodegradation, the marine oils BXPT,BZ34-1 and SZ36-1 are biodegraded in the lab. Based on GC-MS analysis, the results showed that pristane/phytane of three oils degraded obviously during 60 days experiment period. The degradation degree of pristane/phytane could reach 15%~96% and 20%~75% respectively; pristane/phytane have degraded in the early biodegradation period for heavy-gravity crude oil SZ36-1, but pristane/phytane degraded in the mid and late biodegradation period (20～60d) for light-gravity crude oils BXPT and BZ34-1. Phytane are more resistant than pristane. However, hopanes and steranes have not been degraded during whole experiment period. The relative distribution of C1-dibenzothiophene isomers have changed obviously. Pristane/phytane have degraded sharply at low oil concentration of 0.5 and 1.0 g/L, the degradation degree is more than 97%, while at high oil concentration of 5.0 g/L, 10.0 g/L and 20.0 g/L, the degradation degree is only about 60%. And there is no observable sign of alteration of hopanes and steranes during biodegradation period. The relative distribution of C1-dibenzothiophene (C1-DBT) isomers also has changed obviously. Therefore, the biomarker C30-17α(H), 21β(H)-hopane is selected to evaluate bioremediation effect in the whole field experiment.2) The biodegradation of three biomarker groups (include pristane/phytane, hopanes and steranes, alkyl-PAH) were investigated in the field oil-spilled shoreline bioremediation experiments. The result shows that pristane/phytane have degraded obviously during 120 days bioremediation period, its biodegradation degree ranked from 10% to 50%. Hopanes and steranes have not been degraded during bioremediation period. The relative distribution of C1-dibenzothiophene isomers also have changed obviously, and could distinguish biodegradation from abiotic weathering processes. These results suggest the method of using C30-17α(H),21β(H)-hopane as a conserved internal and combination of C1-DBT isomers relative distribution to evaluate biodegradation of crude oil is feasibile.3) The effectiveness of several bioremediation strateges, including adding water-soluble nutrients, adding slow release fertilizer nutrients and adding petroleum degraded inoculums, was evaluated in the field oil-spilled bioremediation experiments by the method of using C30-17α(H),21β(H)-hopane as a conserved internal and combination of C1-DBT isomers relative distribution. For ease of comparation, the first-order degrading-rate constant ( FODRC) of petroleum constituent are calculated based on C30-17α(H),21β(H)-hopane normalized hydrocarbons concentration. The result shows that plots treated with add slow release fertilizer nutrients and petroleum degraded inoculums get the best bioremediation effect weather for the coarse sand substrate or the fine sand substrate. Their FODRCs is more 2.8 and 2.5 times respectively than blank plots; the plots adding water-soluble nutrients get better bioremediation effect, their FODRCs is more 1.9 and 2.2 times respectively than that of blank plots. The plots adding slow release fertilizer nutrients get a good result for the fine sand matrix. Their FODRCs are about 1.9 times more than that of blank plots. However, the treatment of adding slow release fertilizer nutrient is no effective for the coarse sand matrix. The deficient of nitrogen and phosphorous nutrient in the pore water is the mainly factor that limits the growth of petroleum-degrading bacteria.